Transcriptional modulation of the pre-implantation embryo-specific Rnf35 gene by the Y-box protein NF-Y/CBF.
Maternal-to-zygotic transition of a fertilized egg and the subsequent pre-implantation development of the embryo involve zygotic genome activation and reprogramming of gene expression. The goal of the present study is to establish a model suitable for the characterization of transcriptional modulation of mammalian pre-implantation development. Rnf35 is a mouse RING-finger protein gene that is temporally transcribed in the early embryo, but is permanently silenced before the blastocyst stage of development. We first show that the Chinese-hamster ovary-K1 cells are unique in supporting Rnf35 promoter activities in transient transfection assays. Using the permissive Chinese-hamster ovary-K1 cell line, we show that Rnf35 transcription is driven by an Inr (initiator) core promoter element in the absence of a TATA box; the Inr promoter function is confirmed by direct microinjection of mouse one-cell embryos. This is the first demonstration of the involvement of an Inr core promoter element in transcription in pre-implantation development. We show that the Rnf35 promoter is regulated by three obligatory Y-box (CCAAT-box) elements: two Y boxes (Y(I) and Y(II)) located at -81 are coupled in a palindrome and act synergistically in contributing to Rnf35 transcription; the third Y box (Y(III)) is situated at -13, just upstream of the Inr element, and may be an integral part of the Inr function. Electrophoretic mobility-shift assays and competition experiments further reveal that the Y(I) box is bound by the ubiquitous NF-Y (nuclear factor-Y)/CBF (CCAAT-binding factor) and that Y(II) is targeted by an unidentified protein(s) that acts synergistically with the NF-Y. We suggest that the NF-Y, targeting at a Y-box sequence, may function as an important activator in transcriptional regulation of the Rnf35 gene in the pre-implantation embryo.
Huang
Chiu-Jung
CJ
Department of Animal Science, College of Agriculture, Chinese Culture University, Taipei 11192, Taiwan.
Wu
Shinn-Chih
SC
Choo
Kong-Bung
KB
D000818:Animals; D001483:Base Sequence; D001755:Blastocyst / Q000378:metabolism*; D023081:CCAAT-Binding Factor / Q000502:physiology*; D016466:CHO Cells; D017124:Conserved Sequence; D006224:Cricetinae; D003412:Cricetulus; D047108:Embryonic Development; D018507:Gene Expression Regulation, Developmental; D005801:Genes, Homeobox; D051379:Mice; D008813:Mice, Inbred ICR; D011401:Promoter Regions, Genetic; D014157:Transcription Factors / Q000096:biosynthesis / Q000235:genetics*; D014158:Transcription, Genetic
D023081:CCAAT-Binding Factor; C468760:Rnf35 protein, mouse; D014157:Transcription Factors