Faculty of Science
Faculty of Science
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Cell and Developmental Biology (Mango)

Projects & Collaborations

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Defining the role of subnuclear concentration of heterochromatinassociated proteins in genome organization and silencing

Research Project  | 2 Project Members

During eukaryotic development, the genome organizes itself non-randomly into regions of active euchromatin and inactive heterochromatin that differ in their nuclear positioning and preference for long-distance genomic interactions. Post-translational modifications of histones are associated with genome activation or repression in these distinct compartments. Specifically, methylation of histone H3 lysine 9 (H3K9me) marks regions of constitutive heterochromatin, and conserved H3K9 histone methyltransferases (HMTs) have been shown to be able to drive perinuclear anchoring of heterochromatin and to reinforce topologically associated domains (TADs) between repressed regions of the genome. However, it remains unclear whether these HMTs promote TAD formation through mechanisms over and above their enzymatic activity, which is important because amplification or mistargeting of H3K9 HMTs is associated with tumor progression in many cancers. We recently demonstrated in C. elegans that the H3K9 HMT SETDB1 homolog MET-2 associates with an unstructured cofactor LIN-65/ATF7IP to become enriched at dynamic subnuclear foci that behave similarly to phase-separated condensates. Competence to make foci via interaction with LIN-65 was critical for H3K9 methylation, transcriptional repression, and germline viability. Surprisingly, MET-2 can make foci and preserve germline viability independent of its catalytic activity, leading us to hypothesize MET-2 foci themselves contribute physically to heterochromatin stability. To better understand how MET-2 contributes to silencing, we propose to: 1) map the genomic loci with which MET-2 foci stably interact and measure the transcriptional consequences of that interaction; and 2) demonstrate how MET-2 foci influence chromosome shape, long-range contacts, and epigenetic signatures across developmental time. These findings will demonstrate the importance of nuclear membrane-less compartments in higher order chromatin structure in an intact, multicellular organism as well as suggest new avenues for targeting H3K9 HMTs in disease.

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Dynamic organization of chromosomes during development

Research Project  | 1 Project Members

Dynamic Organization of Chromosomes during Development.The goal of this proposal is to map genome architecture during embryogenesis with single-cell resolution, to elucidate the role of chromosome structure in essential cellular processes. To understand the function of the genome, it is not sufficient to know its sequence and local epigenetic features - we must also consider the large-scale physical architecture of entire chromosomes and their positioning within the nucleus. The 3D organization of chromosomes is a key contributor of essential genomic functions, including replication, repair, recombination, and the regulation of gene expression. Recently developed methods to measure chromosome architecture have shown that the genome folds into structures of increasing sizes during interphase: loops, domains, compartments, and territories1,2. However, previous studies have mainly used genome-wide biochemical methods on cultured cell populations to indirectly infer structure and inform computational models of how chromosomes fold in the nucleus. As a result, many gaps remain in our knowledge regarding the role of chromosome structure in vivo. Specifically, how does chromosome organization change in the cells and tissues of an animal? Second, do chromosome structure changes reflect cellular replication, growth and division, or are they a feature of cell fate specification - or a combination of both? The first question will be addressed in Aim 1, and the second in Aim 2. To address these aims, I will exploit the stereotyped cell lineage of C. elegans, where each cell has been identified, and I will track chromosome conformation for specific cell types and their progenitors during embryonic development. I will directly map chromosome structure using a novel high-throughput imaging approach that preserves the cellular, tissue, and organismal contexts of chromosomes.In Aim 1, I will compare the interphase chromosome architecture of different cell lineages in the embryo to define cell-type specific architectures. The germline lineage is the most likely to have a unique architecture, and I will focus on comparing germline and somatic cells to address the relationship between structure and cell identity. I will determine if specific conformations precede, coincide, or follow functional transcriptional output and lineage restriction, which will help to elucidate the causal relationships between these events. In Aim 2, I will map chromosome architecture through the stages of the cell cycle to model the structural transitions that occur as embryonic cells grow and divide. This aim will identify inherited conformational signatures and identify the conformational rearrangements that occur as embryonic cells replicate, divide, and re-establish interphase architecture. I will collaborate with polymer physicists to incorporate structural measurements into computational models of chromosome transitions in the cell cycle. The results of my work will be a conformational atlas of chromosomes in embryogenesis, with a focus on cell type and cell cycle dynamics. I will determine how the specific history of a cell informs its current and future molecular state. This study will conform to the goals of SPARK by bringing new technologies to Switzerland, and through the unique integration of high-throughput imaging with unsupervised clustering and polymer modeling. My work will provide an unconventional analysis of 3D genome architecture in animals at the single-cell level, elucidating the links between chromosome organization and cellular lineage for the first time.

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Analysis of chromosome conformation during dosage compensation and pha-4 regulation

Research Project  | 2 Project Members

Chromosomes occupy territories within nuclei, where they fold into looped structures, including large-scale compartments and smaller topologically associated domains. An open question is whether and how higher-order chromosome conformation impacts transcriptional processes. We recently developed iterative DNA FISH methodology ("chromosome tracing") to define the conformation of chromosomes, or regions of chromosomes, in single cells of C. elegans (Sawh et al., Molecular Cell, 2020). We will use this method to examine the relationship between chromosome morphology and gene expression in two biological contexts. In Aim 1 we will explore the interconnection between chromosome conformation and genes that undergo intermittent transcriptional activation (bursting). In Aim 2, we will examine dosage compensation and its dependence on X chromosome morphology, in collaboration with Barbara Meyer (Univ of California - Berkeley, USA) and Peter Meister (Univ of Bern, CH). These studies will lay a foundation for understanding the role of chromosome organization in C. elegans transcription at the single cell level.

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C. elegans Embryogenesis

Research Project  | 6 Project Members

The formation and physiology of organs is one of the fundamental mysteries of biology: how are cells specified to be part of an organ? How is the development of organ precursors coordinated in space and time? My lab has a longstanding interest in organogenesis of the foregut, or pharynx, with a particular focus on its transcriptional regulatory processes. We use C. elegans because its simple, transparent anatomy and its rapid genetics, genomics and molecular biology provide a powerful system for analysis. In this application we explore how the foregut develops in response to changing environmental conditions. The central regulator of the C. elegans pharynx is pha-4, which encodes a FoxA transcription factor (Horner et al. 1998; Kalb et al. 1998). FoxA factors are critical regulators of digestive tract development in all animals studied to date. They function as selector genes that specify foregut fate during early embryogenesis, and they drive gut differentiation and morphogenesis at later stages (Mango 2009; Lalmansingh et al. 2012; Zaret and Mango 2016). To accomplish these diverse tasks, FoxA factors like PHA-4 control a broad spectrum of target genes expressed at different times or in different cells within the foregut (Gaudet and Mango 2002; Gaudet et al. 2004; Zhong et al. 2010; Zaret and Mango 2016; Von Stetina et al. 2017b). While studies in multiple organisms have identified conserved components required to establish the foregut (Mango 2009; Grapin-Botton 2008), it is less clear what mechanisms modulate these components to allow the digestive tract to respond to varying environmental conditions. C. elegans worms are acutely aware of their surroundings and can sense a wide array of chemicals through 32 chemosensory neurons. These chemicals elicit both short-term responses (feeding or movement) and longer-term effects that alter longevity or development (Bargmann 2006; Hsieh et al. 2017). My lab has found a surprising link between mutations that alter chemosensation in the mother and pha-4 silencing in progeny. We will test the hypothesis that mothers detect chemicals in the environment and impart this information to their progeny. In Aim 1 we will examine the genetic requirements for this cross-generational interaction. This will help elucidate the mechanism. In Aim 2 we will explore the anatomical site of action of factors involved in this process, to delineate how signaling in neurons communicates to embryos. In Aim 3, we will sequence RNAs inherited in very early embryos to identify candidate regulatory molecules. We will test these candidates for modulatory behavior.