Spatiotemporal control of gene expression in Caenorhabditis elegans

Cell-type specific regulation of transcription drives the production of the myriad of different cells generated during development. Profiling genome-wide gene expression landscapes in different tissues has improved our understanding of the physiological and pathological processes taking place during development. Yet, the mechanisms underlying cell-type specific transcription are not well understood. Promoters and enhancers are the key loci that orchestrate spatiotemporal patterns of gene expression. Their activities can range from ubiquitous to highly cell-type specific, and their composition and arrangement define the regulatory grammar directing gene transcription across development. More comprehensive in vivo studies of these regulatory grammars would improve our understanding of how different patterns of gene expression are obtained across tissues. Caenorhabditis elegans is an important model organism for studying develop- mental processes. At the beginning of my PhD, I helped characterize the dynamics of gene expression and chromatin activity across development and aging. Follow- ing this, I aimed to identify and characterize the regulatory elements involved in tissue-specific control of transcription in C. elegans. I jointly profiled chromatin accessibility and gene expression landscapes across the five main tissues of the adult nematode. To achieve this, I developed a method to sort fluorescently labelled nuclei from individual C. elegans tissues. Analyzing the datasets I generated, I first showed that around 80% of the regulatory elements in C. elegans are specifically active in subsets of tissues. I then revealed fundamental differences in the genetic structure and regulatory architecture of genes expressed ubiquitously or in individual tissues, and I defined two distinctive regulatory grammars associated with specific sets of genes. I also uncovered striking and unsuspected differences in nucleosome arrangement and sequence features of ubiquitous and germline-specific promoters compared to somatic promoters. Finally, I optimized a single nucleus method to analyze chromatin accessibility and gene expression during embryogenesis and did a pilot study of early embryo development. My work provides a comprehensive resource of chromatin accessibility and transcription patterns in the different tissues of C. elegans. It sheds light on fundamental differences between the mechanisms of transcription regulation of germline-active genes or somatic tissue-specific genes. The outcome of this work will greatly enable and push forward C. elegans transcription regulation research. The first datasets jointly profiling chromatin accessibility and nuclear transcription across the majority of tissues in a multicellular organism will also be of benefit for the broader community studying gene regulation in eukaryotes

Sox10 regulates enteric neural crest cell migration in the developing gut

Concurrent Sessions 1: 1.3 - Organs to organisms: Models of Human Diseases: abstract no. 1417th ISDB 2013 cum 72nd Annual Meeting of the Society for Developmental Biology, VII Latin American Society of Developmental Biology Meeting and XI Congreso de la Sociedad Mexicana de Biologia del Desarrollo. The Conference's web site is located at http://www.inb.unam.mx/isdb/Sox10 is a HMG-domain containing transcription factor which plays important roles in neural crest cell survival and differentiation. Mutations of Sox10 have been identified in patients with Waardenburg-Hirschsprung syndrome, who suffer from deafness, pigmentation defects and intestinal aganglionosis. Enteric neural crest cells (ENCCs) with Sox10 mutation undergo premature differentiation and fail to colonize the distal hindgut. It is unclear, however, whether Sox10 plays a role in the migration of ENCCs. To visualize the migration behaviour of mutant ENCCs, we generated a Sox10NGFP mouse model where EGFP is fused to the N-terminal domain of Sox10. Using time-lapse imaging, we found that ENCCs in Sox10NGFP/+ mutants displays lower migration speed and altered trajectories compared to normal controls. This behaviour was cell-autonomous, as shown by organotypic grafting of Sox10NGFP/+ gut segments onto control guts and vice versa. ENCCs encounter different extracellular matrix (ECM) molecules along the developing gut. We performed gut explant culture on various ECM and found that Sox10NGFP/+ ENCCs tend to form aggregates, particularly on fibronectin. Time-lapse imaging of single cells in gut explant culture indicated that the tightly-packed Sox10 mutant cells failed to exhibit contact inhibition of locomotion. We determined the expression of adhesion molecule families by qPCR analysis, and found integrin expression unaffected while L1-cam and selected cadherins were altered, suggesting that Sox10 mutation affects cell adhesion properties of ENCCs. Our findings identify a de novo role of Sox10 in regulating the migration behaviour of ENCCs, which has important implications for the treatment of Hirschsprung disease.postprin

From parasite genomes to one healthy world: Are we having fun yet?

In 1990, the Human Genome Sequencing Project was established. This laid the ground work for an explosion of sequence data that has since followed. As a result of this effort, the first complete genome of an animal, Caenorhabditis elegans was published in 1998. The sequence of Drosophila melanogaster was made available in March, 2000 and in the following year, working drafts of the human genome were generated with the completed sequence (92%) being released in 2003. Recent advancements and next-generation technologies have made sequencing common place and have infiltrated every aspect of biological research, including parasitology. To date, sequencing of 32 apicomplexa and 24 nematode genomes are either in progress or near completion, and over 600k nematode EST and 200k apicomplexa EST submissions fill the databases. However, the winds have shifted and efforts are now refocusing on how best to store, mine and apply these data to problem solving. Herein we tend not to summarize existing X-omics datasets or present new technological advances that promise future benefits. Rather, the information to follow condenses up-to-date-applications of existing technologies to problem solving as it relates to parasite research. Advancements in non-parasite systems are also presented with the proviso that applications to parasite research are in the making

90th Annual Meeting of the Virginia Academy of Science: Proceedings

Full proceedings of the 90th Annual Meeting of the Virginia Academy of Science, Norfolk State University, Norfolk Virginia, May 23-25, 201

A regulated response to mitochondrial dysfunction modulates longevity and rates of living

Aging is a long-standing biological question of tremendous social and cultural importance. Despite this, only in the last 15 years has biology started to make significant progress in understanding the underlying mechanisms that regulate aging. This progress stemmed mainly from the use of model organisms, which allowed the discovery of several genes directly modulating longevity. Interestingly, several of these longevity genes are necessary for normal mitochondrial function, and disruption of their activity delays the aging process. This is somewhat paradoxical, considering the importance of cellular respiration for energy production and viability of eukaryotic organisms. One possible rationalization for this is that by decreasing cellular respiration, reactive oxygen species (ROS) generation is also reduced, and in that way, cellular decay and aging are delayed.(...

Defining the Proximal Interactome of Death-Associated Protein Kinase 1

Death-associated protein kinase 1 (DAPK1) is a multidomain cell signalling macromolecule which has been implicated in a plethora of biological processes. The domain topology of this protein comprises a calcium/calmodulin dependent serine/threonine kinase, a Ras of complex proteins (ROC) GTPase and a number of further protein-protein interaction (PPI) interfaces. The precise role and regulation of DAPK1 is unclear although it appears to be complex. Due to the potential for targeting this protein for therapeutic intervention, most notably in relation to cancer and neurodegeneration, understanding the physiological function of DAPK1 is important. Developing our understanding of protein function in the wider cellular context can be achieved by defining its proximal interactome. In this research a PPI network analysis of the human ROCO proteins was performed utilising literature-derived PPI data and novel experimental data to shed light on the commonalities and distinctions within the interaction and functional profiles of these structurally related proteins. This was facilitated by the development of a PPI query resource, termed Protein Interaction Network Online Tool (PINOT), for extracting and processing PPI data from a number of major molecular interaction data repositories. The pursuit of defining the DAPK1 interactome was then translated into the context of the Caenorhabditis elegans proteome for predictive and evidence based mapping of the DAPK-1 interactome. This revealed intriguing novel DAPK-1 interactors, MEP-1, SYD-9 and UNC-14, for further investigation. In addition, a number of mutant dapk-1 C. elegans strains, FLAG-dapk-1, dapk-1 K57W and dapk-1 T715N, were engineered to further assess the role and regulation of DAPK-1 in vivo. Initial phenotypic analysis provided insight into novel DAPK-1 related functions for future examination. Collectively, the analyses performed and resources developed throughout the course of this research project contribute to our understanding of the DAPK1/DAPK-1 interactomes and will guide future investigation into the complex functionality of this protein