Inhibition of transcription by the Caenorhabditis elegans germline protein PIE-1: genetic evidence for distinct mechanisms targeting initiation and elongation.
In Caenorhabditis elegans embryos, specification of the germ lineage depends on PIE-1, a maternal protein that blocks mRNA transcription in germline blastomeres. Studies in mammalian cell culture have suggested that PIE-1 inhibits P-TEFb, a kinase that phosphorylates serine 2 in the carboxyl-terminal domain (CTD) repeats of RNA polymerase II during transcriptional elongation. We have tested this hypothesis using an in vivo complementation assay for PIE-1 function. Our results support the view that PIE-1 inhibits P-TEFb using the CTD-like motif YAPMAPT. This activity is required to block serine 2 phosphorylation in germline blastomeres, but unexpectedly is not essential for transcriptional repression or specification of the germline. We find that sequences outside of the YAPMAPT are required to inhibit serine 5 phosphorylation, and that this second inhibitory mechanism is essential for transcriptional repression and specification of the germ lineage. Our results suggest that PIE-1 uses partially redundant mechanisms to block transcription by targeting both the initiation and elongation phases of the transcription cycle.
Ghosh
Dolan
D
Department of Molecular Biology and Genetics and Howard Hughes Medical Institute, Center for Cell Dynamics, Johns Hopkins School of Medicine, Baltimore, Maryland 21205-2185, USA.
Seydoux
Geraldine
G
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D001757:Blastomeres / Q000166:cytology / Q000378:metabolism; D017173:Caenorhabditis elegans / Q000166:cytology / Q000196:embryology / Q000235:genetics*; D029742:Caenorhabditis elegans Proteins / Q000737:chemistry / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism; D016213:Cyclins / Q000378:metabolism; D004625:Embryo, Nonmammalian / Q000166:cytology / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D005854:Germ Cells / Q000378:metabolism*; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism*; D010768:Phosphoserine / Q000378:metabolism; D011485:Protein Binding; D021381:Protein Transport; D012333:RNA, Messenger / Q000378:metabolism; D014158:Transcription, Genetic; D015053:Zygote / Q000166:cytology / Q000378:metabolism
D029742:Caenorhabditis elegans Proteins; D016213:Cyclins; D009687:Nuclear Proteins; D012333:RNA, Messenger; C100950:pie-1 protein, C elegans; D010768:Phosphoserine