Defining the gene expression cascade underlying cell fate determination is a key element in understanding development. The availability of whole genome microarrays, coupled with the isolation of nearly pure cell type populations, allows one to begin to define the transcriptome associated with specific cell fates. Muscle cells have been attractive targets for such studies in mature animals due to their ease of isolation andor culture and their importance in human pathologies. It has been more difficult, however, to determine early in vivo embryonic myogenic gene expression patterns that would give insights into the regulation of muscle development. To this end, we have applied Micro-Array Profiling of C. elegans (MAPCeL), a techinque developed by our collaborators in the Miller lab, to muscle cell populations extracted from developing nematode embryos that have been marked by the myosin heavy chain promoter from the myo-3 gene (myo-3::gfp). ? ? Fluorescence Activated Cell Sorting (FACS) was used to isolate myo-3::gfp-positive muscle cells, and their cultured derivatives, from dissociated early C. elegans embryos. Microarray analysis of gene expression in these cell populations identified 6,693 expressed genes, 700 of which are enriched in the myo-3::gfp positive cell population. The muscle-enriched gene set was validated by comparisons to known muscle markers, independently derived expression data, and GFP reporters in transgenic strains. The results confirm the validity and power of MAPCeL for cell type-specific expression profiling and reveal a set of 600 genes that are very likely to be specifically enriched in non-pharyngeal muscle cells during embryogenesis. This study demonstrated the power and utility of MAPCeL in defining the in vivo gene expression profile of developing C. elegans muscle cells. The results provide an initial characterization of the muscle cell transcriptome and its deployment that underlies myofiber assembly and function.? ? We have also begun to study the in vivo target genes activated by the myogenic regulator, HLH-1, a homolog of the vertebrate MyoD family. We have previously shown that ectopic expression of hlh-1 in early C. elegans embryos is sufficient to convert most blastomeres to a body wall muscle like fate. To define the transcriptional targets of HLH-1 that underlie muscle cell fate specification and differentiation, we have use chromatin immunoprecipitation (ChIP) followed by probing whole genome tilling microarrays (Chip). The results are beginning to reveal the in vivo binding sites for this transcription factor, allowing us to correlate DNA binding with gene expression in the early embryo. Our hope is that this approach will allow a more exhaustive description of the transcriptional cascade that play out during muscle cell development.
| Lei, Haiyan; Liu, Jun; Fukushige, Tetsunari et al. (2009) Caudal-like PAL-1 directly activates the bodywall muscle module regulator hlh-1 in C. elegans to initiate the embryonic muscle gene regulatory network. Development 136:1241-9 |
| Fox, Rebecca M; Watson, Joseph D; Von Stetina, Stephen E et al. (2007) The embryonic muscle transcriptome of Caenorhabditis elegans. Genome Biol 8:R188 |
