Identification of lineage-specific zygotic transcripts in early Caenorhabditis elegans embryos.
During Caenorhabditis elegans embryogenesis, a maternally supplied transcription factor, SKN-1, is required for the specification of the mesendodermal precursor, EMS, in the 4-cell stage embryo. When EMS divides, it gives rise to a mesoderm-restricted precursor, MS, and an endoderm-restricted precursor, E. To systematically identify genes that function as key regulators of MS and/or E-derived tissues, we identified, by microarray analyses, genes that are newly transcribed within a short developmental window (approximately 30 min) encompassing the generation and fate specification of the MS and E blastomeres. By comparing total cDNAs generated from individual, carefully staged embryos, we identified 275 genes up-regulated in 12-cell embryos compared to 4-cell embryos. Fifty of these 275 genes are down-regulated in 12-cell skn-1 mutant embryos and are designated skn-1-dependent zygotic (sdz) genes. The spatial and temporal expression patterns in C. elegans embryos of 10 randomly selected sdz genes were analyzed by a nuclear GFP reporter driven by the endogenous 5' regulatory sequence of each gene. GFP expression, although absent at the 4-cell stage, was detected at the 12- to 16-cell stage for all 10 genes and was restricted to EMS-derived lineages for 7 of the 10. Among the seven lineage-specific genes, three genes are expressed equally in both MS and E lineages, two are expressed exclusively or predominantly in the MS lineage, and two are expressed exclusively in the E lineage. Depletion of skn-1 by RNAi abolishes the expression of all seven reporter transgenes in vivo, confirming that these genes are indeed skn-1 dependent. These results demonstrate the successful combination of single-staged embryo cDNAs, genetic mutants, and whole transcriptome microarray analysis to identify stage- and lineage-specific transcripts in early C. elegans embryos.
Robertson
Scott M
SM
Department of Molecular Biology, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX 75390, USA.
Shetty
Premnath
P
Lin
Rueyling
R
D000818:Animals; D030801:Animals, Genetically Modified; D017173:Caenorhabditis elegans / Q000166:cytology / Q000196:embryology* / Q000235:genetics*; D029742:Caenorhabditis elegans Proteins / Q000235:genetics / Q000378:metabolism; D019070:Cell Lineage; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental; D017930:Genes, Reporter; D020411:Oligonucleotide Array Sequence Analysis; D011897:Random Allocation; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D015053:Zygote / Q000166:cytology / Q000502:physiology
D029742:Caenorhabditis elegans Proteins; D004268:DNA-Binding Proteins; D014157:Transcription Factors; C073702:skn-1 protein, C elegans