al-Ubaidi M R, Pittler S J, Champagne M S, Triantafyllos J T, McGinnis J F, Baehr W
Department of Ophthalmology, Cullen Eye Institute, Baylor College of Medicine, Houston, Texas 77030.
J Biol Chem. 1990 Nov 25;265(33):20563-9.
The single copy mouse opsin gene produces five major transcripts, varying in size from 1.7 to 5.1 kilobases. The mRNAs are present at levels that vary over 2 orders of magnitude and can be detected as early as postnatal day 1. Each of the transcripts is polyadenylated and can be identified in polysome-bound RNA, suggesting that each is translated in vivo. To elucidate the molecular basis of this complex transcription pattern, we have characterized genomic fragments covering the entire mouse opsin gene, including several kilobases of 5'- and 3'-flanking regions. Transcription initiates at a single site 97 base pairs upstream of the translation start codon. Northern hybridization with exon- and intron-specific probes demonstrated that the various transcripts are not generated by partial or alternative splicing. Sequence analysis of the 3' end of the gene showed the presence of multiple polyadenylation signals. Analysis by polymerase chain reaction of the 3' end of opsin cDNA demonstrated that the complex transcription pattern originated from the selective use of these polyadenylation sites, generating transcripts that differ only in the length of the 3'-untranslated region. Transcript heterogeneity similar to that observed in mouse was also found in rat and, to a lesser degree, in human and frog opsin mRNAs.
单拷贝的小鼠视蛋白基因产生五种主要转录本，大小从1.7到5.1千碱基不等。这些mRNA的水平变化超过两个数量级，早在出生后第1天就能检测到。每个转录本都进行了多聚腺苷酸化，并且可以在多核糖体结合的RNA中鉴定出来，这表明每个转录本都在体内进行翻译。为了阐明这种复杂转录模式的分子基础，我们对覆盖整个小鼠视蛋白基因的基因组片段进行了表征，包括5'和3'侧翼区域的几千碱基。转录起始于翻译起始密码子上游97个碱基对处的单个位点。用外显子和内含子特异性探针进行的Northern杂交表明，各种转录本不是由部分或可变剪接产生的。对该基因3'端的序列分析表明存在多个多聚腺苷酸化信号。对视蛋白cDNA 3'端进行的聚合酶链反应分析表明，这种复杂的转录模式源于对这些多聚腺苷酸化位点的选择性使用，产生的转录本仅在3'非翻译区的长度上有所不同。在大鼠中也发现了与小鼠中观察到的类似的转录本异质性，在人类和青蛙视蛋白mRNA中程度较小。
