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Department of Physiology, Development and Neuroscience

 

Further information and application details

Riccardo Beltramo

Neural pathways for vision-based spatial navigation

Contacts

rb973@cam.ac.uk

https://www.beltramolab.org/

References

Beltramo R. & Scanziani M., A collicular visual cortex: Neocortical space for an ancient midbrain visual structure. Science, 2019 Jan 4;363(6422):64-69. https://doi.org/10.1126/science.aau7052

Beltramo R., A new primary visual cortex. Science, 2020 Oct 2;370(6512):46.https://doi.org/10.1126/science.abe1482

LaChance P.A., Todd T.P. & Taube, J.S. A sense of space in postrhinal cortex. Science, 2019 Jul 12;365(6449), eaax4192 https://doi.org/10.1126/science.aax4192

 

A critical requirement for survival is the ability to navigate in the environment. This skill is believed to rely on internal spatial representations of the surroundings, such as the “spatial maps” found in the hippocampus and parahippocampal cortex.

Sensory inputs are fundamental for shaping these internal spatial representations. In particular, visual cues are considered essential factors that guide navigation. However, how visual input is converted into spatial maps is still poorly understood.

Mammals have two coexisting brain structures that process visual information: the ancient “superior colliculus” and the phylogenetically newer “visual cortex”. This Ph.D. project aims to determine the roles played by these parallel visual systems in spatial navigation.

We have recently discovered that the superior colliculus has a dedicated space in the visual cortex: the postrhinal area (POR) (Beltramo & Scanziani, 2019; Beltramo, 2020). POR, whose responses rely on collicular activity and are critically involved in spatial navigation (LaChance et al., 2019), is perfectly placed at the interface between the cortical and collicular visual streams and the hippocampal formation.

Combining electrophysiological, 2photon imaging, and opto/chemogenetic approaches, we will study the interactions between ancient and modern visual pathways in sensory processing and spatial navigation.

Riccardo Beltramo

Sensory processing and fear overgeneralization in stress-induced anxiety, across the visual system

Contacts

rb973@cam.ac.uk

https://www.beltramolab.org/

References

[1] Dunsmoor J. & Paz R., Fear Generalization and Anxiety: Behavioral and Neural Mechanisms. Biol Psychiatry, 2015 Sep 1;78(5):336-43. https://doi.org/10.1016/j.biopsych.2015.04.010

[2] Beltramo R. & Scanziani M., A collicular visual cortex: Neocortical space for an ancient midbrain visual structure. Science, 2019 Jan 4;363(6422):64-69. https://doi.org/10.1126/science.aau7052

[3] Burgess C.R., Rohan N. Ramesh R.N., Sugden A.U., Levandowski K.M., Minnig M.A., Fenselau H., Lowell B.B,1, Andermann M.L., Hunger-dependent enhancement of food cue responses in mouse postrhinal cortex and lateral amygdala Neuron, 2016 Sep 7; 91(5): 1154–1169. https://doi.org/10.1016/j.neuron.2016.07.032

Anxiety disorders stem from dysfunctions in the circuits that process aversive stimuli[1]. When confronted with potential dangers, animals must select the appropriate behavioral response. In an environment full of complex stimuli, sometimes threat cues are ambiguous, making it challenging to distinguish dangerous from irrelevant stimuli. The ability to generalize across similar stimuli is an adaptive strategy that protects animals from fatally missing a potentially deadly environmental cue. In other words, “better safe than sorry”. However, an exaggerated generalization of fear to harmless stimuli (i.e. “fear overgeneralization”) is maladaptive and considered a hallmark of numerous anxiety disorders.

The cellular and circuit mechanisms mediating fear overgeneralization are largely unknown. This PhD project investigates the development of fear overgeneralization at the level of the sensory circuits that process aversive information.

Using the mouse visual system as a model, we will focus on a visual cortex selectively activated by innately fearful stimuli[2], targeted by the amygdala[3], and driven by a midbrain center that triggers avoidance behaviors[2]. We will determine how changes in the neural sensory representations of dangerous and safe visual stimuli affect the animals’ ability to discriminate them. Given the critical role of preexisting stress in the pathogenesis of numerous anxiety disorders, we will test the hypothesis that emotional distress changes how sensory stimuli are encoded during learning, leading to fear overgeneralization.

Through chronic 2photon imaging, large-scale electrophysiology, and innovative behavioral tasks, we will determine the effects of severe stressors on early sensory processing, during the development of stress-induced anxiety.

Thorsten E. Boroviak

How to build a primate: Towards an in vitro model for primate embyogenesis

Contacts

teb45@cam.ac.uk

https://boroviaklab.pdn.cam.ac.uk/

References

Primate embryogenesis predicts the hallmarks of human naïve pluripotency, Boroviak and Nichols, Development, 2017

In the Boroviak lab, we aim to generate synthetic primate embryos as a functional model system to delineate the mechanisms of early development in our own species.

This project entails bioengineering and microfluidic approaches to reconstitute late blastocysts from non-human primate epiblast-, hypoblast- and trophoblast cell lines. Assembled structures will be allowed to implant on an attachment matrix and sustained using state-of-the-art protocols for postimplantation embryo culture in vitro. Epiblast-like cells will be obtained from naïve pluripotent marmoset embryonic stem cells and we have recently established culture conditions to grow extraembryonic hypoblast and trophoblast stem cells. The student will benefit from our recent spatial embryo profiling dataset, which we will use as a blueprint for the first 25 days of marmoset development (Bergmann et al., Nature, under review). Importantly, this will allow him/her to systematically compare synthetic to in vivo developed embryos to ensure that findings are developmentally relevant. The candidate will be able to take advantage of the unique expertise at the Centre for Trophoblast Research for extraembryonic tissues and will be working in a truly multidisciplinary and stimulating environment in the Boroviak- and Hollfelder laboratories. Many of the relevant techniques for this project are already setup and include: single-cell RNA-seq and triple-omics, 4-colour confocal imaging and virtual reconstruction, microfluidics and embryo micromanipulation.

Ultimately, the results from this study will open new avenues for genome-wide functional interrogation of the transcription factor networks and underlying signalling pathways controlling primate development.

Sarah Bray and Leila Muresan

Decoding the Notch Signal
Contacts

sjb32@cam.ac.uk

https://www.pdn.cam.ac.uk/directory/sarah-bray

References

1. Bray SJ, Gomez-Lamarca M. Notch after cleavage. Curr Opin Cell Biol. (2018) 51:103-109. doi: 10.1016/j.ceb.2017.12.008.

2.Gomez-Lamarca, M. J., Falo-Sanjuan, J., Stojnic, R., Abdul Rehman, S., Muresan, L., Jones, M. L., … Bray, S. J. (2018). Activation of the Notch Signaling Pathway In Vivo Elicits Changes in CSL Nuclear Dynamics. Developmental Cell, 44(5), 611–623.e7. doi:10.1016/j.devcel.2018.01.020

3.Falo-Sanjuan J, Lammers NC, Garcia HG, Bray SJ. (2019) Enhancer Priming Enables Fast and Sustained Transcriptional Responses to Notch Signaling. Developmental Cell 50(4):411-425.e8.

Cells in our body need to communicate with each other, to make and organize the different tissues. The information they receive must then be accurately deciphered so that the appropriate actions are taken—mistakes are at the root of many inherited diseases and cancers. Our focus is on the Notch signalling pathway, which provides key information during developmental decisions and tissue homeostasis [1].  A critical end-result from Notch activity is that genes are turned on and our goal is to find-out how this occurs accurately in the developing animal.  How does the behavior of the nuclear complexes change when signaling is ON so that the information is accurately transduced? We have been using cutting-edge strategies to visualize events in real time, directly measuring dynamics within the nucleus of the key transcription complex [2] and analyzing the real-time profile of target gene transcription during different developmental processes [3]. Current approaches include Single molecule Localization Microscopy (SmLM), with which we have been able to detect and track single molecules within the nucleus with ~20nm precision, and MS2/MCP labelling of nascent transcripts to acquire a quantitative measure of transcription initiation in real-time.  The aim of the projects is to develop methods for analysis of these exciting data to extract key underlying properties and, based on this analysis, to formulate and test hypothetical models of the underlying regulatory mechanisms.

Kevin Chalut and Clare Buckley

Investigating the role of PI3K signalling in mechanics, morphogenesis and cell fate

Contacts

kc370@cam.ac.uk and ceb85@cam.ac.uk

https://www.stemcells.cam.ac.uk/people/pi/chalut and https://www.pdn.cam.ac.uk/directory/clare-buckley

References

1. Belly, H. D. et al. Membrane Tension Gates ERK-Mediated Regulation of Pluripotent Cell Fate. Cell Stem Cell 28, 273-284.e6 (2021).

2. Buckley, C. E. et al. Reversible Optogenetic Control of Subcellular Protein Localization in a Live Vertebrate Embryo. Dev Cell 36, 117–126 (2016).

This collaborative project will use live imaging and optogenetics within embryonic stem cell culture and zebrafish embryos to determine how PI3K is integrated with mechanical signalling, and how it instructs morphogenesis and cell fate through its putative role in mediating cell surface mechanics.

One of the main regulators of cell surface mechanics are phosphoinositides such as PIP2, which act as substrates for a number of factors that regulate the tension of the plasma membrane. A primary effect of PI3K is to phosphorylate PIP2, which places it as an interesting potential signal regulating cell surface mechanics. Moreover, PI3K initiates the Akt signalling cascade, ultimately regulating cell-cell adhesions. The PI3K pathway is also key for apico-basal polarity establishment during epithelial morphogenesis.

It is becoming clear that the regulation of membrane tension and cell-cell adhesions plays an important role in both morphogenesis and cell fate. For example, membrane tension has recently been shown by the Chalut lab to play a critical role in cell fate (1) and cell-cell adhesions have been found by the Buckley lab to be important in directing apico-basal polarity during epithelial morphogenesis.

The Buckley lab have adapted optogenetic techniques to allow in vivo light-mediated control of PI3K signalling in zebrafish embryos (2). In combination with 4D live imaging approaches, we will:

1) Determine the spatiotemporal dynamics of membrane phosphoinositide composition over the course of organ morphogenesis and stem cell differentiation.

2) determine how PI3K signalling affects cell surface mechanics and whether this effect is propagated between cells

3) Characterise the effect of alterations in PI3K signalling on morphogenesis and cell fate

William Colledge and Susan Jones Investigating the physiological changes in Kiss1 neurons occuring at puberty.

Contacts

whc23@cam.ac.uk

pdn.cam.ac.uk/directory/bill-colledge

References

Hwa-Yeo, S., Kyle, V.R., Morris, P.G., Jackman, S., Sinnett-Smith, L, Schacker, M., Chen, C. and Colledge, W.H. (2016). Visualization of Kiss1 neuron distribution using a Kiss1-CRE transgenic mouse J Neuroendocrinol 28(11): 10.1111/jne.12435.

Yeo, S-H and Colledge, W.H. (2018). The Role of Kiss1 Neurons As Integrators of Endocrine, Metabolic, and Environmental Factors in the Hypothalamic-Pituitary-Gonadal Axis. Front. Endocrinol. 9:188. doi: 10.3389/fendo.2018.00188. Review

Mammalian fertility is regulated by neuronal circuits within the hypothalamus that stimulate GnRH secretion. Activation of pulsatile GnRH secretion is a critical event in the onset of puberty and this is regulated by the action of kisspeptin neuropeptides encoded by the Kiss1 gene. In mice, Kiss1 expression has been mapped in the hypothalamus mainly to the arcuate nucleus (ARC) in both males and females and to the anteroventral periventricular nucleus (AVPV) of females. It is known that sex steroids can alter the expression of kisspeptins and the activity of Kiss1 neurons but very little is known about the other physiological changes that occur to Kiss1 neurons during the pubertal transition period. This is an important question, as establishing the intrinsic activity of Kiss1 neurons throughout puberty will provide knowledge into the central mechanisms that contribute to normal fertility.

This project will study the changes that take place in Kiss1 neurons in mice as they progress from the pre-pubertal stage to post-puberty. We have developed a transgenic mouse line in which Kiss1 neurons are fluorescently labelled to allow us to easily identify and study these neurons. The project will involve complementary approaches using a variety of techniques including molecular analysis of gene expression changes, immunohistochemistry and/or electrophysiological measurements of Kiss1 neuronal activity.

Angeleen Fleming

Using zebrafish models to understand childhood neurodegenerative disorders

Contacts

af425@cam.ac.uk

https://www.pdn.cam.ac.uk/directory/angeleen-fleming

References

Fleming, A. and Rubinsztein, D.C. (2020). Autophagy in Neuronal Development and Plasticity. Trends Neurosci. Oct;43(10):767-779. doi: 10.1016/j.tins.2020.07.003.

Childhood neurodegenerative disorders are characterised by both structural and functional changes in the CNS that typically result in progressive neurological dysfunction and often in early death.  Although there are over 600 of these rare diseases, a large number are caused by genetic mutations in protein clearance pathways or lysosomal digestive enzymes.  Our lab uses zebrafish models to study protein clearance pathways in health and disease.

Knockout studies of genes in the autophagy pathway have shown that this can lead to defects in neurogenesis and neuronal plasticity.  However, there are conflicting results from both experimental studies and from clinical data on patients with recessive mutations in autophagy genes.  Hence, it is unclear whether neurological defects arise as a result of deficits in protein/organelle clearance (autophagy) or from non-canonical roles of the encoded protein.  To date, most studies have been performed in vitro or from the post-mortem analysis of knockout mice.  Zebrafish offer a unique opportunity to study these processes in vivo.  We have developed a range of knockout, hypomorph and reporter lines, as well as models where we can up- and down-regulate key autophagy genes with tissue-specific drivers and with temporal control.  The project will use a range of imaging techniques as well as  biochemical and genetic analysis to investigate the role of autophagy in neurogenesis and to determine whether autophagy or the non-canonical roles of these proteins accounts for the neuronal defects associated with childhood neurodegeneration.

Angeleen Fleming

Investigating the role of autophagy in the maintenance neuronal homeostasis

Contacts

af425@cam.ac.uk

https://www.pdn.cam.ac.uk/directory/angeleen-fleming

References

Menzies F.M., Fleming A. and Rubinsztein D.C. (2015). Compromised autophagy and neurodegenerative diseases. Nat Rev Neurosci. Jun;16(6):345-57. doi: 10.1038/nrn3961.

Menzies F.M., Fleming A., Caricasole A., Bento C.F., Andrews S.P., Ashkenazi A., Füllgrabe J., Jackson A., Jimenez Sanchez M., Karabiyik C., Licitra F., Lopez Ramirez A., Pavel M., Puri C., Renna M., Ricketts T., Schlotawa L., Vicinanza M., Won H., Zhu Y., Skidmore J., Rubinsztein D.C. (2017). Autophagy and Neurodegeneration: Pathogenic Mechanisms and Therapeutic Opportunities. Neuron. Mar 8;93(5):1015-1034. doi: 10.1016/j.neuron.2017.01.022.

 

Autophagy is an intracellular clearance pathway that delivers cytoplasmic contents to the lysosome for degradation. It plays a critical role in maintaining protein homeostasis and providing nutrients under conditions where the cell is starved. It also helps to remove damaged organelles and misfolded or aggregated proteins.  Using the zebrafish as a model system, this project seeks to expand our understanding of the role of autophagy in maintaining neuronal homeostasis.

Work in the Fleming laboratory focuses on the roles of toxic aggregate-prone proteins in the pathogenesis of neurodegenerative diseases and the clearance of aggregated proteins via autophagy.  From in vitro studies, we have a good understanding of the intracellular events that occur during autophagosome formation and of the signalling pathways that control this process.  However, little work has been done to determine the rate and regulation of autophagy in different tissue and cell types in vivo, for example, how this contributes to degeneration in neurons.  The ability to quantify and manipulate autophagic flux in vivo is an essential part of our studies and to this end, we have developed a transgenic zebrafish line in which we can observe autophagic flux in vivo.  Zebrafish are an ideal model for these investigations since larvae are small and transparent hence investigations can be performed in real time, in vivo, using non-invasive confocal microscopy.  In addition, we have developed a range of genetic tools to temporally control and manipulate autophagy in different tissues.  The aim of this proposal is to investigate how manipulation (up and down-regulation) of autophagic flux affects pathology in a range of zebrafish neurodegenerative disease models using these newly developed tools.

Elisa Galliano

Activity-dependent neuronal plasticity in the mouse olfactory bulb

Contacts

eg542@cam.ac.uk

https://www.pdn.cam.ac.uk/directory/elisa-galliano

References

[1] Galliano et al (2021) ‘Brief Sensory Deprivation Triggers Cell Type-Specific Structural and Functional Plasticity in Olfactory Bulb Neurons’, J. Neurosci. Off. J. Soc. Neurosci., vol. 41, no. 10, pp. 2135–2151,.

[2] Mandairon et al (2006) ‘Broad activation of the olfactory bulb produces long-lasting changes in odor perception’, Proc. Natl. Acad. Sci. U. S. A., vol. 103, no. 36, pp. 13543–13548.

[3] Erskine et al (2019) ‘AutonoMouse: High throughput operant conditioning reveals progressive impairment with graded olfactory bulb lesions’, PloS One, vol. 14, no. 3, p. e0211571.

All living organisms must constantly sample the ever-changing environment via their sensory organs, and compute the resulting information to generate an appropriate behavioural output. How is this behavioural flexibility achieved, and which are the underlying brain computations? Neurons can modify themselves in response to environmental changes in a process called neuronal plasticity, which is thought to be the basis of adaptation and learning. The different cellular mechanisms of neuronal plasticity (e.g. structural changes in morphology, functional changes of synapses, modulation of the neuron’s intrinsic excitability) have been extensively studied, but largely in isolation. Little is known about how they combine within individual cells to influence neuronal activity and behavioural flexibility. We plan to address this by studying how mice adapt to olfactory stimulation and learn new olfactory tasks, and how this is sustained by flexible computations in the part of their brains that processes odours.

We will perturb the olfactory landscape to trigger adaptive responses by subjecting the mice to either sensory deprivation (=nose blockage, like a mild cold[1]) or olfactory enrichment (=overexposure to odours, like when humans enter a perfume shop[2]). We will take advantage of the mouse genetic toolbox to label neurons that respond to specific odours, and we will use immunohistochemistry and patch-clamp electrophysiology to investigate how neurons in the olfactory bulb plastically change their synaptic connections, shape and intrinsic excitability, after deprivation or enrichment of different durations. Using automated behavioural testing [3], we will then probe the mice ability to sense and discriminate odours to test if and how adaptive plasticity influences learning.

Elisa Galliano

Embryonic and adult neurogenesis: do different birth dates lead to functional diversity?

Contacts

eg542@cam.ac.uk

https://www.pdn.cam.ac.uk/directory/elisa-galliano

References

[1] Lledo, et al. (2006) ‘Adult neurogenesis and functional plasticity in neuronal circuits’, Nat. Rev. Neurosci., vol. 7, no. 3, pp. 179–193.

[2] Galliano et al. (2018) ‘Embryonic and postnatal neurogenesis produce functionally distinct subclasses of dopaminergic neuron’, eLife, vol. 7.

[3] Galliano et al. (2021) ‘Brief Sensory Deprivation Triggers Cell Type-Specific Structural and Functional Plasticity in Olfactory Bulb Neurons’, J. Neurosci. vol. 41, no. 10, pp. 2135–2151.

A crucial aspect of brain development and function is that neurons can structurally and functionally modify themselves and the strength of their connections with other neurons in response to certain stimulus patterns. These changes pertain to three main classes of plasticity: synaptic intrinsic, and structural. In the olfactory circuit, structural plasticity is taken to an extreme: not only neurons can change size and shape of neuronal sub compartments, but quite a few neuronal subpopulations can regenerate throughout life, adding and removing entire elements of the circuit [1]. Among these regenerating cells are olfactory sensory neurons in the nasal epithelium, dopaminergic cells and granule cells in the olfactory bulb, and interneurons in the olfactory cortex. While adult-born neurons have long been believed to be a like-for-like replacement of embryonic-born ones, recent work focusing on bulbar dopaminergic neurons has challenged this view. Indeed, embryonic and postnatally-born dopaminergic cells differ in morphology, function and activity-dependent plasticity [2], [3]. Using transgenic mouse models, immunohistochemistry, electrophysiology, and behavioural testing, this project wants to expand on these findings. Specifically it wishes to investigate (a) whether these differences based on birth date seen in the dopaminergic population can be generalized to the other regenerating populations in the olfactory system, and (b) what behavioural roles do embryonic and regenerating cells play in olfactory processing.

Dino A. Giussani

Maternal Obesity: Translatable Programmed Cardiovascular Dysfunction in Offspring

Contacts

dag26@cam.ac.uk

www.pdn.cam.ac.uk/directory/dino-giussani

References

Giussani DA.  Breath of Life: Heart Disease Link to Developmental Hypoxia.  Circulation 144(17):1429-1443, 2021

Botting KJ, Skeffington KL, Niu Y, Allison BJ, Brain KL, Itani N, Beck C, Logan A, Murray AJ, Murphy MP, Giussani DA. Translatable mitochondria-targeted protection against programmed cardiovascular dysfunction. Sci Adv. 6(34):eabb1929, 2020

Allison BJ, Brain KL, Niu Y, Kane AD, Herrera EA, Thakor AS, Botting KJ, Cross CM, Itani N, Shaw CJ, Skeffington KL, Beck C, Giussani DA.Altered Cardiovascular Defense to Hypotensive Stress in the Chronically Hypoxic Fetus.  Hypertension 76(4):1195-1207, 2020.

 

Obesity has reached epidemic proportions, including in women of reproductive age. Over half of women in the UK are now obese or overweight during pregnancy. This is of significant concern as obesity during pregnancy not only has detrimental effects on the mother, but also on her children. Evidence derived from human studies and from animal models shows that maternal obesity can markedly increase the risk of heart disease in the offspring, even when young and when the progeny are fed a healthy diet and they themselves do not become obese. This highlights that it is something about exposure of the embryo or fetus to an obesity environment during gestation itself that either triggers a fetal origin of cardiovascular dysfunction and/or increases the risk of heart disease in the adult offspring. Therefore, this PhD project will test the hypothesis that maternal obesity during pregnancy promotes a fetal origin of heart disease via adverse mechanisms triggered by mitochondria-derived oxidative stress.  Causality will be addressed directly using mitochondria-targeted antioxidant intervention with MitoQ. The hypothesis will be tested using an established ovine model of maternal obesity during pregnancy.  In contrast to rats and mice, which are born highly immature, the timing of cardiovascular developmental milestones in sheep and humans is much more similar.  Therefore, using sheep markedly improves the human clinical translation of the biomedical research.  The work will adopt an integrative approach, combining experiments of in vivo cardiovascular function with those at the isolated organ, cellular and molecular levels, as we have published before in the accompanying references. In particular, the successful PhD candidate will be trained in fetal cardiovascular surgery.

Dino A. Giussani

Embryonic Origins of Heart disease: The Impact of Intermittent Hypoxia

Contacts

dag26@cam.ac.uk

www.pdn.cam.ac.uk/directory/dino-giussani

References

Badran M, Yassin BA, Lin DTS, Kobor MS, Ayas N, Laher I. Gestational intermittent hypoxia induces endothelial dysfunction, reduces perivascular adiponectin and causes epigenetic changes in adult male offspring. J Physiol. 597(22):5349-5364, 2019.

Giussani DA.  Breath of Life: Heart Disease Link to Developmental Hypoxia.  Circulation 144(17):1429-1443, 2021.

Itani N, Skeffington KL, Beck C, Niu Y, Giussani DA. Melatonin rescues cardiovascular dysfunction during hypoxic development in the chick embryo. J Pineal Res. 60(1):16-26, 2016.

Obstructive sleep apnoea (OSA) is characterized by episodes of intermittent hypoxia (IH), which promote oxidative stress and increase the risk of heart disease in affected patients. In turn, human pregnancy is associated with OSA, which is aggravated by obesity, the rates of which in the UK, including in women of reproductive age, are reaching epidemic proportions.  However, the effects of maternal IH due to OSA during pregnancy on the cardiovascular health of the offspring are only just beginning to be addressed.  A recent study reported that IH in a mouse model of OSA during pregnancy programmed cardiovascular dysfunction in the adult offspring.  However, mechanisms remain uncertain because the partial contributions of IH on the mother, placenta and fetus are difficult to disentangle.  This PhD project will study the effects of IH in the chicken embryo, an established model system that permits isolation of the direct effects of developmental challenges on the cardiovascular system of the offspring, independent of effects on the mother and/or the placenta.

Fertilised chicken eggs will be exposed to normoxia or IH sing an Oxycycler (BioSpherix). At day 19 of the 21-day incubation period, hearts will be isolated.  In one cohort of embryos, the heart will be mounted onto a Langendorff preparation to determine effects on systolic and diastolic function during basal conditions and in response to a period of ischaemia-reperfusion (IR).  Cardiac IR injury will be determined by infarct size (tetrazolium chloride staining).  In another cohort of embryos, cardiac mitochondrial respiratory capacity and substrate preference will be determined by permeabilised cardiac muscle fibre respirometry.   In separate cohorts, experiments will be repeated at adulthood.

Courtney Hanna and Miguel Constancia

Investigating the role for epigenetic modifier MLL2 in placental development

Contacts

cwh36@cam.ac.uk

https://www.pdn.cam.ac.uk/staff/hanna

References

(1) Rugg-Gunn P, Cox B, Ralston A, Rossant J. (2010) Distinct histone modifications in stem cell lines and tissue lineages from the early mouse embryos. PNAS 107:10783-90. (2) He W, Wei Y, Gong X, Chang L, Jin W, Liu K, Wang X, Xiao Y, Zhang W, Chen Q, et al. (2020) Developmentally Delayed Epigenetic Reprogramming Underlying the Pathogenesis of Preeclampsia. BioRxiv. doi: https://doi.org/10.1101/2020.05.08.085290. (3) Denissov S, Hofemeister H, Marks H, Kranz A, Ciotta G, Singh S, Anastassiadis K, Stunnenberg HG, Stewart AF. (2014) Mll2 is required for H3K4 trimethylation on bivalent promoters in embryonic stem cells, whereas Mll1 is redundant. Development 141:526-37.

 

The placenta forms the maternal-foetal interface in pregnancy, controlling nutrient and waste exchange, hormone production and immunotolerance. Impaired placental function is linked to poor pregnancy outcomes. Yet, early regulators of placental development that lead to a functional placenta and healthy pregnancy are poorly understood.

After implantation, embryonic stem cells undergo a process of ‘epigenetic priming,’ a necessary transition that allows these pluripotent cells to be receptive to signalling for differentiation into all of the cell types of the body. A hallmark of this priming event is the deposition of bivalent chromatin at a subset of promoters, marked by active H3K4me3 and repressive H3K27me3, poising them for activation. Failure to prime the embryonic genome results in developmental defects. The role for bivalent chromatin in placenta remains contentious (1,2) and remains an important research question. The aim of this project is to characterise and test the functional importance of epigenetic priming in placental cells, linking poor priming to defects in placentation and consequences on foetal growth.

Using mouse as a model, this project will investigate the role for H3K4 methyltransferase MLL2, a critical enzyme in the establishment of bivalent chromatin (3), in priming placental cells for differentiation. Three important questions will be addressed:

1. What gene promoters are targeted by MLL2 during placental lineage specification?

2. What are the consequences of loss of MLL2 on placental gene expression during development?

3. How does loss of MLL2 impact placental differentiation and function during pregnancy?

The student will obtain training in a variety of techniques including next generation sequencing, histology, embryology and bioinformatics.

Allan Herbison

Neural synchronisation mechanisms underlying brain control of fertility

Contacts

aeh36@cam.ac.uk

https://www.pdn.cam.ac.uk/directory/allan-herbison

References

Herbison AE (2016) Control of puberty onset and fertility by gonadotropin-releasing hormone neurons. Nature Rev Endo, 12, 452-466.

Piet R, Kalil B, McLennan T, Porteous R, Czieselsky K, Herbison AE (2018) Dominant neuropeptide co-transmission in kisspeptin-GABA regulation of GnRH neuron firing driving ovulation. J Neurosci 38, 6310-6322

All mammalian females have a large surge of luteinizing hormone (LH) secretion at mid-cycle to trigger ovulation. Studies have now identified that a population of kisspeptin neurons located in the preoptic area of the forebrain are responsible for triggering a massive increase in the release of gonadotrophin-releasing hormone (GnRH) into the pituitary gland that, in turn, drives the LH surge (Herbison, 2016).  Revealing the mechanisms through which the preoptic area kisspeptin neurons drive the GnRH surge is at the heart of understanding mammalian ovulation and, potentially, ovulatory disorders.

This project will use cutting edge GCaMP imaging, optogenetic and electrophysiology approaches in acute brain slices prepared from genetically modified mice (Piet et al., 2018) to examine the mechanisms through which preoptic area kisspeptin neurons synchronize their activity to drive the GnRH neuron surge. Where appropriate studies may extend to the observation of single neuron and population kisspeptin neuron activity in freely behaving mice. The student will join a vibrant laboratory using a variety of state-of-the-art cellular and in vivo approaches to examine the neural networks controlling fertility in genetic mouse models.

Allan Herbison

Brainstem modulation of the GnRH pulse generator controlling fertility

Contacts

aeh36@cam.ac.uk

https://www.pdn.cam.ac.uk/directory/allan-herbison

References

 

Herbison AE (2016) Control of puberty onset and fertility by gonadotropin-releasing hormone neurons. Nature Rev Endo, 12, 452-466.

Clarkson J, Han SY, Piet R, McLennan T, Kane G, Ng J, Porteous R, Kim J, Colledge WH, Iremonger KJ, Herbison AE (2017) Definition of the hypothalamic GnRH pulse generator in mice. Proc Natl Acad Sci (USA) 114, E10216-E10223.

Han SY, Clarkson J, Piet R, Herbison AE (2018) Optical approaches for interrogating neural circuits controlling hormone secretion. Endocrinology, 159, 3822-3833.

After more than three decades of searching, the neurons responsible for generating pulsatile reproductive hormone secretion have been identified as the kisspeptin neurons located in the arcuate nucleus of the hypothalamus (Clarkson et al., 2017).  These cells directly control the secretion of gonadotropin-releasing hormone (GnRH) into the pituitary and are now considered to be the “GnRH pulse generator”. However, a wide range of environmental and internal homeostatic factors modulate the frequency of the GnRH pulse generator and, hence, the fertility of an individual (Herbison, 2016).  The serotonin and noradrenaline neurons of the brainstem project throughout the forebrain to convey fundamental aspects of life including stress, mood, and arousal state on neuronal activity. As such, it is very likely that they modulate the activity of the GnRH pulse generator. This project will use the latest in vivo GCaMP imaging and chemogenetic approaches (Han et al., 2018) in addition to viral tract tracing to examine the impact of brainstem noradrenaline and serotonin neurons on the activity of the kisspeptin GnRH pulse generator in a suite of genetically modified mice.  The studies are aimed at understanding the core neural network responsible for controlling reproductive hormone secretion with a view to improving the treatment of infertility in the clinic.

Randall Johnson

Role of exogenous and endogenous metabolites in immunotherapy

Contacts

rsj33@cam.ac.uk

https://randalljohnsonlab.com

References

Veliça P, Cunha PP, Vojnovic N, Foskolou IP, Bargiela D, Gojkovic M, Rundqvist H, Johnson RS. (2021) Modified Hypoxia-Inducible Factor Expression in CD8+ T Cells Increases Antitumor Efficacy. Cancer Immunology Research DOI: 10.1158/2326-6066.CIR-20-0561

Rundqvist H, Veliça P, Barbieri L, Gameiro PA, Bargiela D, Gojkovic M, Mijwel S, Reitzner SM, Wulliman D, Ahlstedt E, Ule J, Östman A, Johnson RS. (2020) Cytotoxic T-cells mediate exercise-induced reductions in tumor growth. eLife 9:e59996

Tyrakis P, Palazon A, Macias D, Kian LL, Phan AT, Veliça P, You J, Chia GS, Sim J, Doedens A, Abelanet A, Evans CE, Griffiths JR, Poellinger L, Goldrath AW and Johnson RS. (2016) S-2-hydroxyglutarate regulates CD8+ T-lymphocyte fate. Nature 540:236-241

The project will focus on how metabolites, and specifically those induced by hypoxia, are modulated by immune response and activation, particularly in cytotoxic T cells and macrophages. A secondary but important focus will be on how manipulation of metabolite levels, either genetically or pharmacologically, could modulate immune response, particularly during cancer immunotherapy.

Julija Krupic

Neuronal-glial interactions in the hippocampus during learning

Contacts

jk727@cam.ac.uk

www.krupiclab.com

References

Craddock et al 2018 Frontiers in Cellular Neuroscience; Zhou et al 2021 PNAS

The aim of the project is to investigate neuronal-glial interactions in the hippocampus during learning. The hippocampus is one of the first areas affected during Alzheimer’s disease (AD), and glial cells emerge as key players in pathogenesis. In particular, astrocytes have been implicated in altering synapse function both directly and indirectly by mediating inflammatory signalling, which can worsen hippocampal damage or dysfunction. To interrogate this possibility, the candidate will perform in vivo two-photon imaging of neurons and astrocytes visualised by different colours while the animal is performing learning tasks in virtual environments. Our unique experimental set up allows simultaneous and multimodal monitoring of cell-type-specific functions, therefore, time-dependent correlated activity between astrocytes and neurons could be assessed during spatial learning. The studies will be performed in wild type mice as well as in well-established AD models such as the APP and EC-Tau mice, which are available for the lab.

Kathy Niakan

Lineage tracing the human amnion using imaging and sequencing-based approaches

Contacts

kkn21@cam.ac.uk

https://www.pdn.cam.ac.uk/staff/niakan

References

1. Nakamura, T. et al. A developmental coordinate of pluripotency among mice, monkeys and humans. Nature 537, 57–62 (2016).

2. Strnad, P. et al. Inverted light-sheet microscope for imaging mouse pre-implantation development. Nature 13, (2016).

3. Fogarty, N. M. E. et al. Genome editing reveals a role for OCT4 in human embryogenesis. Nature 550, 67–73 (2017).

An important event in human embryogenesis that remains controversial is whether the amnion originates from the epiblast, that gives rise to the embryo proper, or extraembryonic placental progenitor cells. The amnion is essential for fetal growth and facilitates exchange of biochemical products, water and nutrients between mother and fetus. It was hypothesized that placental progenitor cells give rise to amniotic epithelium and more recent studies in monkey suggests that the amniotic cavity emerges from within the epiblast(1). The exact timing of amnion specification remains is unclear and detailed lineage tracing of the resolution required to determine the origin of human amniotic cells has not yet been performed.

The goal of this study is to utilise imaging and sequencing-based approaches to trace the lineages of human embryos in vitro up to 14 days post-fertilization to determine the origins of the human amnion. The student will utilise embryo labelling methods that we have recently optimised, light sheet microscopy and methods for cell segmentation to generate a lineage tree similar to methods that have been recently developed(2). Depending on the success of the first approach, alternative or complementary approaches will be the use for genetic labelling. The student would be trained to utilise our existing pipelines for genome editing and single-cell multi-omics analysis(3) to conduct lineage tracing using either induced heritable mutations, engineered mutagenesis with optical in situ readout or a combination of approaches with the imaging method above. The study of these cell fate decisions would provide fundamental insights into human development, stem cell biology, and has importance clinically to improve infertility treatment.

Kathy Niakan

Investigating the role of BMP signalling in the divergence of the hypoblast and epiblast

Contacts

kkn21@cam.ac.uk

https://www.pdn.cam.ac.uk/staff/niakan

References

1. Blakeley, P., Fogarty, N.M.E., del Valle, I., Wamaitha, S.E., Hu, T.X., Elder, K., Snell, P., Christie, L., Robson, P., and Niakan, K.K. (2015). Defining the three cell lineages of the human blastocyst by single-cell RNA-seq. Development 142, 3613–3613. Available at: http://dev.biologists.org/lookup/doi/10.1242/dev.131235.

2. Fogarty, N.M.E., McCarthy, A., Snijders, K.E., Powell, B.E., Kubikova, N., Blakeley, P., Lea, R., Elder, K., Wamaitha, S.E., Kim, D., Maciulyte, V., Kleinjung J., Kim, J.-S., Wells, D., Vallier, L., Bertero, A., Turner, J.M.A., Niakan, K.K. (2017). Genome editing reveals a role for OCT4 in human embryogenesis. Nature 550, 67–73. Available at: http://dx.doi.org/10.1038/nature24033.

3. Gerri C., McCarthy A., Alanis-Lobato G., Demtschenko A., Bruneau A., Loubersac S., Fogarty N.M.E., Hampshire D., Elder K., Snell P., Christie L., David L., Van de Velde H., Fouladi-Nashta A.A. and Niakan K.K. (2020) A conserved molecular cascade initiates trophectoderm differentiation in human, bovine and mouse embryos prior to blastocyst formation. Nature, 587: 443-447.

Based on preliminary data in our laboratory, we hypothesise that BMP signalling functions to regulate the second cell fate decision in humans, which is the divergence of the pluripotent epiblast and hypoblast, which give rise to the embryo proper or the yolk sac, respectively. BMP signalling pathway components are expressed in human embryos(1) and our preliminary data shows that signalling activity was detectable in hypoblast cells. Given the temporal expression and cell type-specific restriction of pSMAD1/5/9, the downstream effector of this pathway, we hypothesize that BMP signalling may be required for establishment or maintenance of the human hypoblast. To further study the function of BMP signalling the student will use specific inhibitors (DMH1 or LDN193189) and activators (exogenous BMP) of this pathway to determine the developmental window for BMP signalling requirement and whether it leads to a fate switch between epiblast and hypoblast cells or is required later in development shortly after implantation. To complement the pharmacological modulation of this signalling pathway the student will induce expression of antagonists of the BMP signalling pathway (i.e. NOGGIN or GREMLIN-1). Using established immunofluorescence and single-cell multiomics analysis(2) the student will determine the functional role of this signalling pathway in human pre- or early post-implantation development and perform comparative analysis in several species, similar to previous studies(3), to determine conserved and divergent programs that lead to a step wise acquisition of cell fate. We anticipate that this student project would provide fundamental insights about the role of the BMP signalling pathway in regulating early human development.

Ole Paulsen

Neuromodulation of spike timing-dependent plasticity in the hippocampus

Contacts

op210@cam.ac.uk

https://noggin.pdn.cam.ac.uk

References

Brzosko Z, Schultz W, Paulsen O (2015) Retroactive modulation of spike timing-dependent

plasticity by dopamine. eLife 4:e09685.

Brzosko ZA, Zannone S, Schultz W, Clopath C, Paulsen O (2017) Sequential neuromodulation

of Hebbian plasticity offers mechanism for effective reward-based navigation. eLife 6:e27756.

Brzosko Z, Mierau S and Paulsen O (2019) Neuromodulation of spike timing-dependent

plasticity: Past, present, and future. Neuron 103: 563-581.

Synaptic plasticity is the leading candidate for a cellular mechanism of learning and memory, but it has been difficult to reconcile the time scales of induction of synaptic plasticity with the time scales of learning. Neuromodulation of synaptic plasticity is a potential mechanism to explain this apparent discrepancy. In particular, different brain states are associated with different activity in cholinergic and dopaminergic inputs (Brzosko et al., 2019). We have recently found that cholinergic and dopaminergic stimulation in the

hippocampus can bias plasticity in opposite directions; acetylcholine biases plasticity towards depression, whereas dopamine biases plasticity towards potentiation (Brzosko et al., 2015, 2017). Surprisingly, the effect of dopamine can be retroactive, converting synaptic

depression into potentiation when applied after the induction of plasticity (Brzosko et al., 2015). These mechanisms are likely to have important implications for the mechanisms of memory formation. We hypothesise that dopamine, as a reward signal, changes the synaptic weights towards potentiation making the animal more likely to seek rewarded locations, and that cholinergic activity, which is strong during explorative behaviour, enables animals to learn from unrewarded events. A PhD project in this area would combine a basic mechanistic study of neuromodulation of plasticity with elucidating the behavioural consequences of this neuromodulation. Techniques would include electrophysiological recording, optogenetics, and behavioural memory testing. The research should lead to new insights into the mechanisms and functions of synaptic plasticity in the brain.

Jasper Poort

The role of GABAergic inhibitory interneurons in visual learning

Contacts

jp816@cam.ac.uk

https://www.pdn.cam.ac.uk/svl

References

Poort, J., Wilmes, K., Chadwick, A., Blot, A., Sahani, M. Clopath, C., Mrsic-Flogel, T., Hofer, S., Khan, A., (2021). Learning and attention increase neuronal response selectivity in mouse primary visual cortex through different mechanisms. BioRxiv, https://doi.org/10.1101/2021.01.31.429053 .

Khan, A.G, Poort, J., Chadwick, A., Blot, A., Sahani, M., Mrsic-Flogel, T., Hofer, S. (2018). Distinct learning-induced changes in stimulus selectivity and interactions of GABAergic interneuron classes in visual cortex. Nature Neuroscience, 21, 851-859. https://doi.org/10.1038/s41593-018-0143-z

Poort, J., Khan, A.G., Pachitariu, M., Nemri, A., Orsolic, I., Krupic, J., Bauza, M., Sahani, M., Keller, G., Mrsic-Flogel, T.D., and Hofer, S.B. (2015). Learning Enhances Sensory and Multiple Non-sensory Representations in Primary Visual Cortex. Neuron 86, 1478–1490. https://doi.org/10.1016/j.neuron.2015.05.037

 

The brain is continuously bombarded with visual input but has limited processing capacity. Learning to selectively process visual features relevant for behaviour is therefore crucial for optimal decision-making and thought to rely on activity of GABAergic inhibitory interneurons. Altered inhibition is linked to perceptual and learning impairments and associated with neurodevelopmental disorders including schizophrenia and autism.

The aim of this project is to understand the precise role of different types of GABAergic inhibitory interneurons in visual learning. Mice have a similarly organized visual cortex and show complex decision-making behaviours. Mouse brain circuits can be measured and manipulated during behaviour in ways not possible in humans.

Our approach is to train head-fixed mice, including pharmacological and genetic mouse models of neurodevelopmental disorders and healthy controls, in visual decision-making tasks. We measure activity in visual cortex in specific cell types using 2-photon imaging and electrophysiology and use optogenetics to activate or inactivate activity of specific interneuron cell types. We will also apply two new innovative methods to optically measure the inhibitory neurotransmitter GABA (developed in the Looger lab, UCSD) and to locally pharmacologically manipulate GABA levels in the brain (Malliaras and Proctor labs, Dept of Engineering, Cambridge) during visual learning.

The PhD project is associated with a Wellcome Trust funded Collaborative programme with a cross-disciplinary international research team to investigate the role of GABAergic inhibition in mice and humans at different scales, from local circuits to global brain networks.

Jasper Poort

Population coding in visual cortex during visually-guided decision-making

Contacts

jp816@cam.ac.uk

https://www.pdn.cam.ac.uk/svl

References

Poort, J., Wilmes, K., Chadwick, A., Blot, A., Sahani, M. Clopath, C., Mrsic-Flogel, T., Hofer, S., Khan, A., (2021). Learning and attention increase neuronal response selectivity in mouse primary visual cortex through different mechanisms. BioRxiv, https://doi.org/10.1101/2021.01.31.429053

Meyer, A.F., Poort, J., O'Keefe, J., Sahani, M., Linden, J.F. (2018) A head-mounted camera system integrates detailed behavioral monitoring with multichannel electrophysiology in freely moving mice. Neuron, 10, 46-60. https://doi.org/10.1016/j.neuron.2018.09.020

Poort, J., Khan, A.G., Pachitariu, M., Nemri, A., Orsolic, I., Krupic, J., Bauza, M., Sahani, M., Keller, G., Mrsic-Flogel, T.D., and Hofer, S.B. (2015). Learning Enhances Sensory and Multiple Non-sensory Representations in Primary Visual Cortex. Neuron 86, 1478–1490. https://doi.org/10.1016/j.neuron.2015.05.037

Visual stimuli drive activity in populations of neurons in the visual cortex. These populations jointly represent reliable information about the visual features, but also about the animal’s behaviour, and task-specific variables to guide decision-making. However, the precise encoding scheme is not yet well understood.

The first aim of this project is to determine how visual, behavioural and task-specific variables are represented in neuronal populations during visually-guided decision-making. The second aim is to understand how learning and visual attention modify visual population coding to improve decision-making.

We will analyse 2-photon imaging and electrophysiological recordings in head-fixed mice trained on visual discrimination tasks. Additionally, we will investigate recordings in freely moving mice performing visual discriminations, with a lightweight head-mounted camera system integrating detailed behavioural monitoring of eye and head movements with multichannel electrophysiology in freely moving mice (Meyer et al., 2018).

With generalized linear models (see Poort et al., 2015) we can quantify how different behavioural and task-specific predictor variables (e.g. licking of the reward spout, running speed, pupil size, visual stimulus onset, reward onset) contribute to activity in individual neurons before and after learning, and with and without attention. We can incorporate different predictors into a multivariate autoregressive model (MVAR) that predicts activity of neurons from simultaneously recorded neighbour cells and reveals changes in interactions between cells (Poort et al., 2021). Neural circuit modelling can provide further demonstration of computational mechanisms that can explain the experimental data (see Poort et al., 2021).

Eleanor Raffan

Genetics of obesity and body composition - from animals to man

Contacts

er311@cam.ac.uk

https://www.pdn.cam.ac.uk/directory/eleanor-raffan

We study spontaneously occurring phenotypes in veterinary species (dogs, farm animals and currently whales too) to learn how genes affect metabolism, body composition and weight gain.  We use a genome-wide association studies and other population genetics approaches, epidemiology, molecular biology and physiological studies to find relevant genetic variants and test their effects.  I can offer a projects to match students’ interest (bioinformatics or laboratory based, species, CNS vs peripheral metabolism).  These might include:

•  Identifying the genetic determinants of intramuscular fat (‘marbling’) in pigs, chickens, cattle and sheep and performing comparative analyses in human genomic data to identify whether genes involved are important in human obesity too.

•  Generating polygenic risk scores for obesity in dogs (from existing pilot GWAS data) and testing their predictive value for across different dog populations.

•  Comparing methodological approaches for genome wide association studies in mixed breed dog populations.

•  Investigating whether genes associated with fat mass in humans are under positive selection in whales – which we hypothesise may a survival advantage during migration (Collaboration with Susan Bengston-Nash).

•  Functional studies of mutations which are candidates for causing obesity in dogs, using a combination of bioinformatics and ‘wet lab’ cellular models to investigate their effect on protein function or gene expression.

•  Epidemiological modelling of environmental causes of obesity in a large multi-breed dog cohort.

If you are interested, please get in touch. We are a small friendly group with 3 PhD students and lots of support as you learn the approaches and techniques required for the project.

Angela Roberts and Kevin Mulvihill The contribution of subcallosal cingulate cortex to negative biases in decision making of relevance to mood and anxiety disorders

Contacts

acr4@cam.ac.uk

https://www.pdn.cam.ac.uk/directory/angela-roberts

References

1. Alexander L., Wood C.M., Gaskin, P.R.L., Sawiak S.J., Fryer T.D., Hong, Y.T., McIver L., Clarke, H.F., Roberts A.C. Over-activation of primate subgenual cingulate cortex enhances the cardiovascular, behavioural and neural responses to threat reactivity (2020) Nature Communications 11, 5386. https://doi.org/10.1038/s41467-020-19167-0

2. Alexander, A., Clarke, H.F., Roberts, AC. A Focus on the functions of area 25. (2019) Brain Sciences 9:129.

3. Hales CA, Robinson ES, Houghton CJ.  Diffusion Modelling Reveals the Decision Making Processes Underlying Negative Judgement Bias in Rats. (2016) PLoS One 29;11(3):e0152592. doi: 10.1371/journal.pone.0152592.

Using chemogenetics (DREADDs) technology this project will explore the differential contribution of distinct regions within the subcallosal cingulate cortex of the common marmoset to negative biases in decision making using an ambiguous cue paradigm. Skills will be acquired in primate cognitive testing, computational behavioural analysis, chemogenetics, immunohistochemistry and neuroanatomy.
Angela Roberts and Kevin Mulvihill Behavioural and brain development across adolescence in the common marmoset

Contacts

acr4@cam.ac.uk

https://www.pdn.cam.ac.uk/directory/angela-roberts

References

1. Sawiak S.J., Shiba Y., Oikonomidis L., Windle C.P., Santangelo A.M., Grydeland H., Cockcroft G., Bullmore E.T*., Roberts A.C. *  Trajectories and milestones of cortical and subcortical development of the marmoset brain from infancy to adulthood.  (2018) Cerebral Cortex 28:4440-4453.  

2. Foulkes L Blakemore S-J Is their heightened sensitivity to social stimuli in adolescence? Currrent Opinion in Neurobiology (2016) 40:81-85.

3. Adolescence and Mental Health The Lancet (2019) 393:2030-2031.

Adolescence is a vulnerable time in a human’s life and is a period of increased risk for developing anxiety and mood disorders. It is also a period of increased risk taking and increased sensitivity to social stimuli. To determine whether adolescent marmosets also show this heightened sensitivity to social reward their sensitivity to videos of social stimuli will be assessed at various stages of adolescence. Any behavioural changes in sensitivity will be correlated with changes in brain structure and function as measured by multi-modal neuroimaging, including structural, myelination and resting state. Skills will be acquired in primate cognitive testing, neuroanatomy and neuroimaging analysis .

Bénédicte Sanson

Role of the adhesion molecule Sidekick in tricellular junction remodeling during developmental morphogenesis

Contacts

bs251@cam.ac.uk

https://www.pdn.cam.ac.uk/directory/benedicte-sanson

References

1)  Lye CM, Naylor HW, Sanson B. (2014) Subcellular localisations of the CPTI collection of YFP-tagged proteins in Drosophila embryos. Development. doi: 10.1242/dev.111310 PMID: 25294944

2) Finegan TM, Hervieux N, Nestor-Bergmann A, Fletcher AG, Blanchard GB, Sanson B. (2019) The tricellular vertex-specific adhesion molecule Sidekick facilitates polarised cell intercalation during Drosophila axis extension. PLoS Biology. doi: 10.1371/journal.pbio.3000522 PMID: 31805038

3) Higashi T, Chiba H. Molecular organization, regulation and function of tricellular junctions. (2020) Biochim Biophys Acta Biomembr. doi: 10.1016/j.bbamem.2019.183143 PMID: 31812626.

 

Tricellular junctions form at cell corners, where three cells meet in epithelial sheets. These special sites harbor components that control passage during normal physiology and also maintain adhesion between cells when epithelia change shape and grow. Recent evidence indicates that tricellular vertices are important for tension sensing and force transmission during epithelial morphogenesis. These sites can also be exploited by invading pathogens and cancer cells, causing disease.

We have discovered that Sidekick, an Ig-family adhesion molecule known previously for its role in the visual system, has an unexpected localization and role at cell corners in Drosophila embryos during developmental morphogenesis (1, 2). In particular, Sidekick is required for polarized cell intercalation in several Drosophila tissues, which is important for tissue extension. Sidekick is unique because it localizes at the level of adherens junctions, while known tricellular junction proteins are found at the level of occluding junctions, both in vertebrates (angulins and tricellulins) and in invertebrates (Gliotactin, Anakonda, M6) (3). This novel role for Sidekick gives an entry point to study the role of tricellular adherens junctions. Aims for this PhD project are to investigate i) how Sidekick localizes at tricellular junctions ii) how this localization facilitates cell rearrangements and other morphogenetic cell behaviours iii) whether this role in Drosophila epithelia is conserved in vertebrates (In zebrafish, in collaboration with Dr. Clare Buckley, PDN). Approaches will include molecular and cellular studies, microscopy (including photo-activation and super-resolution, the latter in collaboration with Prof. Daniel St Johnston, Gurdon Institute) and image analysis.

Elena Scarpa
co-Supervisor Prof Ewa Paluch

Mechanical forces, cell shapes and sizes: asymmetric cell divisions and fate determination in vivo

Contacts

es697@cam.ac.uk

https://www.pdn.cam.ac.uk/staff/scarpa

References

Scarpa E., Finet C., Blanchard G.B., Sanson B. (2018). Actomyosin-Driven Tension at Compartmental Boundaries Orients Cell Division Independently of Cell Geometry In Vivo., Developmental Cell 17:47(6):727-740. PMCID: PMC6302072.

Scarpa E., Szabo A., Bibonne A., Theveneau E.,   Parsons M., Mayor R. (2015). Cadherin switch during EMT in neural crest cells leads to contact inhibition of locomotion via repolarization of forces, Developmental Cell 34(4):421-34. PMCID: PMC4552721.

Serres, M.P., Samwer, M., Truong Quang, B.A., Lavoie, G., Perera, U., Gorlich, D., Charras, G., Petronczki, M., Roux, P.P., and Paluch, E.K. (2020). F-Actin Interactome Reveals Vimentin as a Key Regulator of Actin Organization and Cell Mechanics in Mitosis. Dev Cell 52, 210-222 e217. 10.1016/j.devcel.2019.12.011.

Cells dividing in tissues must push against other cells around them.  In vitro studies have shown that physical confinement causes mechanical stress during mitosis, leading to asymmetric cell divisions and mitotic errors. However, the effects of mechanical forces on mitosis have not yet been studied in vivo.

Using translucent Zebrafish embryos we will investigate the role of mechanical stress on asymmetric divisions and cell fate decisions in vivo.

We will study a population of embryonic stem cells essential to vertebrate development, neural crest cells (NCCs). In the trunk of the Zebrafish, they migrate through narrow spaces in between other tissues, before differentiating into glia or neurons. We observed that Zebrafish NCCs divide asymmetrically, giving rise to a large daughter cell and a smaller one. Moreover, our preliminary long term movies show that daughter cells adopt different fates: the larger daughter becomes a neuron; the smaller one differentiates as glia.

We will use advanced imaging techniques to manipulate forces in live embryos. For example, to make more space for these cells to migrate, we will use laser surgery to target the tissues around them.  We will ask whether neural crest cells divide asymmetrically in vivo because of compression, and if this affects their differentiation. By live imaging chromosomes and cell membranes we will investigate daughter cells shapes and sizes in response to mechanical perturbations, and we will use a photoconvertible transgenic line to track cell fates.

To complement our in vivo experiments, we will set up primary Zebrafish NCCs cultures. In collaboration with Prof. Ewa Paluch, we will identify the compression threshold that leads to asymmetric divisions by confining cells under gels of different stiffness or size.

Erica Watson

Inherited oocyte factors involved in transgenerational epigenetic inheritance

Contacts

edw23@cam.ac.uk

https://www.pdn.cam.ac.uk/directory/watson-erica

References

Padmanabhan et al, 2013. Mutation in folate metabolism causes epigenetic instability and transgenerational effects on development. Cell 155(1): 81-93

Blake et al, 2021. Defective folate metabolism causes germline epigenetic inheritance and distinguishes Hira as a phenotype inheritance biomarker. Nature Communications 12(1): 3714.

Blake and Watson, 2016. Unravelling the complex mechanisms of transgenerational epigenetic inheritance. Current Opinion in Chemical Biology 33:1-7.

Despite decades of research, little is known about the molecular mechanism of folate metabolism during development. The Watson lab analyses mice with the Mtrr^gt mutation, which is hypomorphic and sufficiently disrupts folate metabolism to address this question. Mtrr^gt/gt mice display features of dietary folate deficiency in humans including altered DNA methylation, and a wide spectrum of developmental defects (e.g., growth defects, congenital malformations). Importantly, one Mtrr^gt allele can cause transgenerational inheritance of developmental phenotypes. Indeed, an Mtrr^gt allele in the F0 generation disrupts development of the F2-F4 generations, even when the F1 generation onwards is wildtype. The mode of TEI in the Mtrr^gt model is complex since phenotypes are inherited via the maternal grandparental lineage meaning that an Mtrr+/gt male or female can initiate the effect in their wildtype daughters. We hypothesize that the inducing factor of TEI differs when initiated by an oocyte v. sperm. Hira gene, which encodes for a histone chaperone, acts as a transcriptional biomarker of maternal phenotype inheritance. The project goal is to determine if HIRA is an oocyte factor that mediates TEI by influencing nucleosome spacing in the early embryo. Dysfunction might enable the reconstruction of epigenetic instability and phenotypes. The project will assess: i) Hira-/- knockout mice for evidence of TEI of developmental phenotypes and molecular markers similar to Mtrrgt model, and ii) HIRA expression (confocal imaging) and function (ChIP-seq of H3.3 and/or nucleosome spacing via ATAC-seq) in wildtype F1-F3 early embryos from an F0 Mtrr+/gt female or male. Results will help us understand whether HIRA dysregulation in oocytes induces TEI caused abnormal folate metabolism.

Fengzhu Xiong

Tissue mechanics in neural tube morphogenesis

Contacts

fx220@cam.ac.uk

https://www.gurdon.cam.ac.uk/research/xiong

The neural tube is a developmental precursor of the vertebrate central nervous system, including the brain and the spinal cord. It begins as a specified flat epithelial tissue called the neural plate, which then drastically deforms by bilaterally folding towards the midline. As the folds move to meet dorsally, they fuse to close the neural tube with an internal lumen. The folding process happens in conjunction with the morphogenesis of neighboring tissues including the paraxial mesoderm and the surface ectoderm. The failure of this folding and closure process underlies the neural tube defects (NTDs) in human development. From a physics perspective, tissue folding must require driving forces generated by the neural cells and/or the neighboring tissues. In addition, the mechanical properties of the neural tissue may also be regulated to ensure correct tissue deformation progress under the driving forces. The origins and magnitudes of forces from different sources are not well understood, nor are the regulatory mechanisms of tissue rheological properties. Consequently, an integrated picture of biomechanical model of neural tube morphogenesis is missing. To address this challenge, our project will involve (but not limit to) the following approaches using the avian embryonic neural folds as a model system: 1. use imaging to quantify the shape dynamics of the neural plate as it folds towards a tube; 2. use soft gels, magnetic droplets and cantilevers to dissect the contributions from tissue intrinsic (e.g., apical constrictions, intercalations) and extrinsic (e.g., paraxial mesoderm, endoderm) forces to neural tube folding; 3. use molecular and genetic methods to test relevant factors (e.g., cell polarity, adhesion, folate pathway) in regulating tissue biomechanics.

Fengzhu Xiong

Morphogenetic robustness under fluctuations of tissue tension

Contacts

fx220@cam.ac.uk

https://www.gurdon.cam.ac.uk/research/xiong

Early amniote embryos are mechanically supported by extraembryonic and maternal tissues. These structures contribute tissue forces that may be essential for normal development of the embryo. In avian embryos, for example, the gastrula is under tension sustained by the vitelline membrane that encloses the yolk. Recently, we found that this tension decreases over developmental time to facilitate stage-specific morphogenesis of the embryo body. Such changes appear to only take effect over long periods at persisted high or low levels. On shorter time scales, the tension fluctuates naturally as the eggs are moved and also in ex ovo experimental culture conditions. Interestingly, in contrast to persistent tension, the morphogenetic dynamics of embryonic tissues remain largely unaffected by these fluctuations. This project aims to identify the mechanism ensuring this robustness against mechanical stochasticity on the embryo. Using a combination of theory, imaging, and mechanical and molecular perturbations, we will profile the tension fluctuation and tissue soft matter properties and test the hypothesis that tissues distinguish true morphogenetic forces from random fluctuations by responding preferentially to stresses at long time scales.