An updatedC. elegansnuclear body muscle transcriptome for studies in muscle formation and function

Abstract

The body muscle is an important tissue used in organisms for proper viability and locomotion. The contractile unit of the muscle is the sarcomere, which is ultimately responsible for the contraction reaction leading to movement. Although this tissue is generally well studied and characterized, and many pathways have been elucidated throughout the years, we still lack a comprehensive understanding of its transcriptome, and how it controls muscle development and function. Here, we have updated a nuclear FACS sorting-based approach to isolate and sequence a high-quality muscle transcriptome from C. elegans mixed stage animals. We have identified 2,848 muscle-specific protein-coding genes, including 78 transcription factors and 206 protein-coding genes containing an RNA binding domain. We studied their interaction network, performed a detailed promoter analysis, and identified novel muscle-specific cis-acting elements. We have also identified 16 high-quality muscle-specific miRNAs, studied their function in vivo using fluorochrome-based analyses, and developed the first high-quality miRNA Interactome in a living organism, incorporating other muscle-specific datasets produced by our lab and others. Our study expands our understanding of how muscle tissue functions in C. elegans and in turn, provide results that can in the future be applied to humans to study muscular-related diseases.

Tissue-specific profiling of age-dependent miRNAomic changes in Caenorhabditis elegans

“…Our results overlap with three previous miR-NAomic studies of worm tissues with more miRNAs identified (Supplementary Fig. 1, Supplementary Data 2) [25][26][27] . Of the miRNAs reported in miRbase, 113 are considered highly confident 28 , and we detected 83 of them.…”

“…One is to immunoprecipitate Argonaute (AGO) protein expressed tissuespecifically and profile the associated miRNAs 26 . The last one is through FACS of the nucleus which are labelled using tissue-specific fluorescent proteins 27 . None of the methods directly profile miRNAs in the worm cells.…”

“…Moreover, since these two methods sample the whole worm, miRNAs from larger tissues (e.g., intestine) could be much easier to be detected than those from smaller ones (e.g., coelomocytes). As for the last one, it discards cytoplasm, focusing on a fraction of total miRNAs 27 .…”

“…This could be due to their low expression in the examined tissues at D1 or D8. Besides, PmiR::GFP may fail to reflect their endogenous transcription because 4 of the 11 miRNA genes were not detected in previous miRNAomic analysis in worm tissues (Supplementary Data 2) 20,[25][26][27] . To avoid any potential ambiguity, these miRNAs were therefore excluded from our analysis.…”

A Comprehensive Analysis of 3-UTRs in Caenorhabditis elegans

“…Throughout the years, our lab has refined the 3'UTR collection in C. elegans in the whole animal and in several somatic tissues in various stages of development 10,13,14,24,26,27 (Main Figure 1B).…”