title: The CCAAT element in the CIWI promoter regulates transcriptional initiation in chicken primordial germ cells.
abstract: The P-element-induced wimpy testis (PIWI) protein, which associates with small non-coding RNAs, is responsible for maintaining the integrity of germ cells and stem cells. Thus, transcriptional regulation of PIWI is critical for its effective functional modulation. In this study, we identified the promoter region of the PIWI homolog in chicken (CIWI), and investigated the transcriptional regulatory elements that control expression of CIWI in chicken primordial germ cells (PGCs). We constructed a vector that included the enhanced green fluorescent protein (eGFP) gene controlled by the 4-kb CIWI promoter. The vector was expressed in chicken PGCs, but not in chicken embryonic fibroblasts. Based on promoter deletion and fragmentation assays, we found that a 252-bp fragment of the CIWI promoter (-577 to -326 bp) was crucial for CIWI expression in PGCs. A CCAAT transcriptional regulatory element (-498 to -494 bp) was detected in the proximal region from the transcription initiation site of CIWI, and mutational analysis confirmed that this element regulates transcriptional initiation in chicken PGCs. Interestingly, the regions flanking the CCAAT element, which are positioned differently in HIWI (human), Miwi (mouse), and CIWI orthologs, were highly conserved. In addition, we predicted that specificity protein 1 (SP1) motifs modulate the transcriptional initiation of CIWI by binding to the 5'-flanking regions of the CCAAT box. Overall, 252 bp of the CIWI promoter possessing the transcriptional regulatory element CCAAT is crucial for regulating CIWI gene expression in chicken PGCs. This promoter may be applicable for the regulation of CIWI expression during germ-cell development.
authors.affiliation: Department of Agricultural Biotechnology, College of Agriculture and Life Sciences, Seoul National University, Seoul, Korea.
authors.forename: Yoon Ah
authors.initials: YA
authors.lastname: Sohn
authors.forename: Sang In
authors.initials: SI
authors.lastname: Lee
authors.forename: Hee Jung
authors.initials: HJ
authors.lastname: Choi
authors.forename: Hyun Jeong
authors.initials: HJ
authors.lastname: Kim
authors.forename: Ki Hyun
authors.initials: KH
authors.lastname: Kim
authors.forename: Tae Sub
authors.initials: TS
authors.lastname: Park
authors.forename: Jae Yong
authors.initials: JY
authors.lastname: Han
mesh_terms: D000818:Animals; D060565:Argonaute Proteins / Q000235:genetics*; D001665:Binding Sites; D023081:CCAAT-Binding Factor / Q000235:genetics*; D002478:Cells, Cultured; D002642:Chick Embryo; D002645:Chickens; D005854:Germ Cells / Q000502:physiology*; D017354:Point Mutation; D050436:Regulatory Elements, Transcriptional; D016415:Sequence Alignment; D014157:Transcription Factors / Q000235:genetics*
chemical_list: D060565:Argonaute Proteins; D023081:CCAAT-Binding Factor; D014157:Transcription Factors