短通讯:人核糖体转录因子UBF 5′区的分子分析

S. Dühr, A. Torres-Montaner, A. Astola, F. García-Cózar, C. Pendón, J. Bolívar, M. Valdivia
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摘要

上游结合因子UBF是参与核糖体DNA基因转录的核仁自身抗原。此前,人类基因组克隆证明了单个基因的替代前mrna剪接可用于形成UBF1和UBF2。为了完整表征该核仁转录因子的5 '端基因组组织,我们使用仓鼠UBF cDNA作为探针,从人胎盘基因组文库中分离出含有人UBF基因的lambda克隆。从HeLa基因组DNA中分离出一个额外的PCR产物,覆盖了含有ATG起始密码子的基因的第一个翻译的60nt。我们还通过引物延伸分析确定了该基因在起始ATG密码子上游nt 188的转录起始位点。它首先用于鉴定覆盖人类UBF cDNA首121 nt的UBF基因上的一个未翻译的初始外显子,并建立近端启动子序列。人类UBF启动子缺乏TATA和CAAT盒子,但包含SP1、API、AP2、TFIID、NF-1的多个结合位点和NFAT-1的单个结合位点。因此,我们确定了人类UBF基因的前五个外显子,覆盖7.5 kb。完整的基因现在由20个外显子和中间序列组成,长度约为15kb。
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Short Communication: Molecular Analysis of the 5′ Region of Human Ribosomal Transcription Factor UBF
Upstream binding factor, UBF, is a nucleolar autoantigen involved in the transcription of ribosomal DNA genes. Previously, human genomic clones served to demonstrate that an alternative pre-mRNA splicing of a single gene is used to form UBF1 and UBF2. Here, to complete characterizing the 5′end genomic organization of this nucleolar transcription factor, lambda clones containing the human UBF gene were isolated from a human placenta genomic library using a hamster UBF cDNA as a probe. An additional PCR product was isolated from HeLa genomic DNA to cover the first translated 60 nt of the gene containing the ATG initiation codon. We have also determined the transcription start site of the gene by primer extension analysis at nt 188 upstream from the start ATG codon. It served first, to identify an untranslated initial exon on the UBF gene covering the first 121 nt of human UBF cDNA, and also to establish the sequence of the proximal promoter. The human UBF promoter lacks a TATA and CAAT boxes but contains multiple binding sites for SP1, API, AP2, TFIID, NF-1 and a single site for NFAT-1. Consequently we have defined the first five exons of the human UBF gene covering 7.5 kb. The complete gene now consists of 20 exons with intervening sequences and spans approximately 15kb of DNA.
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