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首页> 外文期刊>Molecular and Cellular Biology >Transcription initiation sites and structural organization of the extreme 5' region of the rat neural cell adhesion molecule gene.
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Transcription initiation sites and structural organization of the extreme 5' region of the rat neural cell adhesion molecule gene.

机译：大鼠神经细胞粘附分子基因5'端的转录起始位点和结构组织。

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Through analysis of rat genomic cosmid clones, the 5'-most exon of the rat neural cell adhesion molecule (NCAM) gene was identified. This exon, here named exon 0, contained the entire 5' untranslated region and the N-terminal signal sequence of the polypeptide. Exon 0 was isolated from a 1.6-kilobase (kb) EcoRI-HindIII fragment of rat genomic cosmid clone 9 which was 35 kb in length. This fragment was sequenced and found to contain approximately 940 base pairs (bp) of 5'-flanking sequence, exon 0, which was approximately 245 bp in length, and approximately 400 bp of the following intron 0. By using information derived from this fragment and the pR18 rat NCAM cDNA, the transcription initiation sites were determined with two assays. Both primer extensions and nuclease S1 protection assays of postnatal day 7 rat brain RNA identified seven initiation sites within a single 10-bp region at positions -195 to -186 relative to the translation start site. An additional minor site was found at position -329. In the immediate 5' region, no consensus TATA or CCAAT sequences were found. Potential regulatory elements within this region include Sp1 consensus binding sites and also a 178-bp homopurine-homopyrimidine sequence containing several mirror repeats. NCAM has multiple transcripts which are regulated in a developmental and tissue-specific fashion. To determine whether these transcripts are initiated at the same sites, transcription initiation sites were analyzed in postnatal day 7 and adult rat brain and also in cultured cell lines of neuronal, glial, and muscle phenotypes. These tissues and cells exhibited distinct NCAM transcript populations in Northern (RNA) dot blot analysis. In all cases similar transcription start sites were found, suggesting that all major NCAM transcripts have similar or identical initiation sites. These results provide essential information to begin analysis of NCAM regulation in different tissues and during development.
机译：通过对大鼠基因组粘粒克隆的分析，鉴定了大鼠神经细胞粘附分子（NCAM）基因的5'-最外显子。该外显子，这里称为外显子0，包含多肽的整个5'非翻译区和N端信号序列。从长度为35kb的大鼠基因组粘粒克隆9的1.6-千碱基（kb）的EcoRI-HindIII片段中分离出外显子0。对该片段进行测序，发现其包含约940个碱基对（bp）的5'侧翼序列，外显子0（其长度约为245 bp）和约400 bp的以下内含子0。和pR18大鼠NCAM cDNA，转录起始位点用两种测定法确定。出生后第7天大鼠脑RNA的引物延伸和核酸酶S1保护试验均在相对于翻译起始位点-195至-186的单个10 bp区域内确定了七个起始位点。在位置-329处发现了另一个未成年人站点。在紧邻的5'区域中，未发现共有的TATA或CCAAT序列。该区域内的潜在调控元件包括Sp1共有结合位点，以及包含几个镜像重复的178bp高嘌呤-高嘧啶序列。 NCAM具有多种转录本，这些转录本以发育和组织特异性的方式受到调控。为了确定这些转录本是否在相同位点启动，在出生后第7天和成年大鼠脑中以及神经元，神经胶质和肌肉表型的培养细胞系中分析了转录起始位点。这些组织和细胞在Northern（RNA）点印迹分析中表现出不同的NCAM转录物种群。在所有情况下，都发现了相似的转录起始位点，这表明所有主要的NCAM转录本都具有相似或相同的起始位点。这些结果为开始分析不同组织和发育过程中的NCAM调控提供了重要信息。

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