[
European Worm Neurobiology Meeting,
2009]
Authors acknowledge support from MRC. Numerous studies support a role for aggregated Ab as a mediator of the toxicity that underlies Alzheimer.s disease (AD) and other diseases such as Inclusion Body myositis (IBM), an acquired muscle disease affecting people over 50 years old. In this pathology, muscle weakness and degeneration is accompanied by chronic muscle inflammation. Remarkably, despite the inflammation component of the disease, IBM patients are only poorly responsive to anti-inflammatory drugs suggesting that inflammation per se may not be the primary cause of the pathology. We have used a C. elegans transgenic line over-expressing human amyloid 1-42 peptide in the muscles as a model with which to test the efficiency of new compounds on in vivo amyloid toxicity. This C. elegans line becomes paralyzed shortly after the induction of amyloid expression in the muscles, which makes it an easily-recordable phenotype useful for exploring candidate treatments to alleviate the paralysis. We report on the actions of two novel chemicals, which inhibit amyloid aggregation and partially rescue the amyloid-induced phenotype. References: Wu Y, W.Z., Butko P, Christen Y, Lambert MP, Klein WL, Link CD, Luo Y. (2006) J. Neurosci., 26, 13102-13. Link CD, T.A., Kapulkin V, Duke K, Kim S, Fei Q, Wood DE, Sahagan BG. (2003) Neurobiol Aging., 24, 397-413. Jones, A.K., Buckingham, S.D. and Sattelle, D.B. (2005). Nat Rev Drug Discov, 4, 321-30.
[
International Worm Meeting,
2021]
Despite containing identical genomes, developing cells differentiate into a plethora of diverse cell types. Distinct patterns of gene expression now form the basis for classifying different cell fates. How the combinatorial activity of transcription factors, chromatin regulators and histone modifications achieve the proper spatiotemporal patterns of gene expression is a major question in developmental biology. Biologists increasingly appreciate the need to investigate gene expression regulation at the single-cell level because much heterogeneity and complexity is lost when averaging across populations of cells. However, profiling chromatin at the single cell level is challenging due to limited input material. Chromatin immunocleavage with sequencing (ChIC-seq) is an efficient method to study chromatin modifications from low input samples. ChIC-seq utilises antibody targeted micrococcal nucleases, leading to controlled, binding-dependent enzymatic digestion of DNA. This releases short fragments which become preferentially incorporated during library preparation and enables high resolution mapping of genomic positions. Crucially, the absence of crosslinking and immunoprecipitation steps, required in less sensitive techniques such as ChIP-seq, leads to minimal material loss. Recently, ChIC-seq was used to profile histone modifications in single human cells [1]. Adapting ChIC-seq to profile histone modifications in C. elegans will provide a powerful tool for studying the epigenetic regulation of development. Here, we present progress in optimising ChIC-seq for profiling chromatin modifications at single-cell level across a developmental time-course in C. elegans. Specifically, we combine Cre/Lox lineage tracing with cell isolation and FACS procedures in order to isolate postembryonic mesoderm cells. Following prolonged quiescence, the mesoblast precursor resumes proliferation and produces fourteen muscle cells, two scavenger cells, and two migratory bipotent myoblasts over 24-hours. By profiling chromatin modifications at high temporal resolution, we aim to reveal regulatory processes controlling cellular proliferation and differentiation. This work will shed light on how epigenetic modifications contribute to cellular decision making in a living animal. [1] Ku WL, Nakamura K, Gao W, Cui K, Hu G, Tang Q, Ni B, Zhao K. (2019) Single-cell chromatin immunocleavage sequencing (scChIC-seq) to profile histone modification. Nature Methods, vol. 16, pages 323-325.