Cell-Specific mRNA Profiling of the Caenorhabditis elegans Somatic Gonadal Precursor Cells Identifies Suites of Sex-Biased and Gonad-Enriched Transcripts

Abstract

The Caenorhabditis elegans somatic gonad differs greatly between the two sexes in its pattern of cell divisions, migration, and differentiation. Despite decades of study, the genetic pathways directing early gonadal development and establishing sexual dimorphism in the gonad remain largely unknown. To help define the genetic networks that regulate gonadal development, we employed cell-specific RNA-seq. We identified transcripts present in the somatic gonadal precursor cells and their daughter cells of each sex at the onset of sexual differentiation. We identified several hundred gonad-enriched transcripts, including the majority of known regulators of early gonadal development, and transgenic reporter analysis confirmed the effectiveness of this approach. Before the division of the somatic gonad precursors, few sex-biased gonadal transcripts were detectable; less than 6 hr later, after their division, we identified more than 250 sex-biased transcripts, of which about a third were enriched in the somatic gonad compared to the whole animal. This indicates that a robust sex-biased developmental program, some of it gonad-specific, initiates in the somatic gonadal precursor cells around the time of their first division. About 10% of male-biased transcripts had orthologs with male-biased expression in the early mouse gonad, suggesting possible conservation of gonad sex differentiation. Cell-specific analysis also identified approximately 70 previously unannotated mRNA isoforms that are enriched in the somatic gonad. Our data illustrate the power of cell-specific transcriptome analysis and suggest that early sex differentiation in the gonad is controlled by a relatively small suite of differentially expressed genes, even after dimorphism has become apparent.

Cell Non-autonomous Function of daf-18/PTEN in the Somatic Gonad Coordinates Somatic Gonad and Germline Development in C. elegans Dauer Larvae

“…We used tissue-specific promoters to drive daf-18(+) as part of our analysis of the cellular focus for daf-18 activity. The promoters ckb-3p [22] and hlh-12p [71] have been described in continuous development; we corroborated that they have similar expression in dauer (see below).…”

“…We tested this inference by restoring DAF-18 to the somatic gonad primordium using the ckb-3 promoter [22]. In dauer larvae, ckb-3p::mCherry::H2B expression is seen in all cells of the somatic gonad primordium and in only three other cells, tentatively identified as MS-derived pharyngeal muscle segment pm6 of the pharynx (STAR Methods).…”

“…As in all other constructs with the ckb-3 promoter, a synthetic intron ''S'' was included to increase expression efficiency (see below and [56,57]). The ckb-3p [22] drives highest expression in Z1 and Z4 and becomes progressively weaker as the lineages progress.…”

Spatiotemporal Gene Expression Analysis of theCaenorhabditis elegansGermline Uncovers a Syncytial Expression Switch

“…Finally, our data set (Tables S1-S11) can be used to find new paths for the control of germline development, gametogenesis, and the different layers of gene regulation at the chromosomal and genome-wide levels. Recently, transcriptome profiling of the C. elegans somatic gonad precursor cells has been published (Kroetz and Zarkower 2015). As data sets for germline precursor cells, and the transcriptomes for different single-cell germline developmental stages become available, for C. elegans and other organisms, it will also be possible to assess the evolutionary conservation of these findings, and use these comparisons to find both common as well as unique strategies by which gene regulation is used to control germline proliferation and differentiation.…”

Phenotypic Screening in C. elegans as a Tool for the Discovery of New Geroprotective Drugs

“…However, it is possible to cultivate isolated cells from embryos or larval stages and induce their in vitro differentiation [155][156][157]. In addition, FACS sorting of cells obtained from dissociated larvae can be performed [158,159]. The lack of commercial antibodies against most C. elegans antigens combined with the fact that many proteins in C. elegans are divergent enough from their vertebrate counterparts to preclude the use of vertebrate antibodies is also a disadvantage of this organism [160].…”