两个加工假基因以及小鼠U1 snRNP特异性蛋白C的cDNA的克隆与特性分析

Nelissen R L, Gunnewiek J M, Lambermon M H, Van Venrooij W J

Department of Biochemistry, University of Nijmegen, The Netherlands.

Gene. 1997 Jan 15;184(2):273-8. doi: 10.1016/s0378-1119(96)00612-9.

Genes for the snRNP proteins U1-70K, U1-A, Sm-B'/B, Sm-D1 and Sm-E have been isolated from various metazoan species. The genes for Sm-D1 and Sm-E, which were isolated from a murine and human source respectively, appear to belong to a multigene family. It has been suggested that also for the mammalian U1-C protein such a multigene family exists. With the human U1-C cDNA as a probe, two genes containing sequences homologous to the probe sequence were isolated from a mouse genomic library. Simultaneously, a murine U1-C cDNA was isolated from a mouse cDNA library. This 0.74 kb cDNA contains an open reading frame (ORF) of 477 bp encoding a polypeptide of 159 amino acids (aa) which differs at only one position (position 65) from the human U1-C protein. One of the isolated U1-C genes contains an ORF as well and shares 92% nucleotide sequence identity with the mouse U1-C cDNA. The features of this gene, in particular the absence of introns, the acquisition of a 3' poly(A) tail and flanking direct repeats, indicate that it represents a processed pseudogene. At the predicted aa sequence level, substitutions of conserved residues at functionally important positions are observed, strongly suggesting that expression of this gene would not lead to a functional polypeptide. The second U1-C gene appeared to be a pseudogene as well because it is also intronless and contains a frameshift mutation compared to the ORF in the mouse U1-C cDNA. The characterization of these two pseudogenes points to the existence of a U1-C multigene family in mice. Furthermore, comparison of aa sequences of the murine, human and Xenopus U1-C shows that the protein is highly conserved through evolution. Since the Xenopus U1-C differs from the two mammalian counterparts solely at a number of positions in the C-terminal region, it can be concluded that aa changes are less well tolerated in the N-terminal region of U1-C than in the rest of the protein.

已从多种后生动物物种中分离出小核核糖核蛋白（snRNP）蛋白U1 - 70K、U1 - A、Sm - B'/B、Sm - D1和Sm - E的基因。分别从小鼠和人类来源分离出的Sm - D1和Sm - E基因似乎属于一个多基因家族。有人提出，哺乳动物的U1 - C蛋白也存在这样一个多基因家族。以人U1 - C cDNA为探针，从小鼠基因组文库中分离出两个含有与探针序列同源序列的基因。同时，从小鼠cDNA文库中分离出一个小鼠U1 - C cDNA。这个0.74 kb的cDNA包含一个477 bp的开放阅读框（ORF），编码一个159个氨基酸（aa）的多肽，该多肽与人类U1 - C蛋白仅在一个位置（第65位）不同。分离出的一个U1 - C基因也包含一个ORF，与小鼠U1 - C cDNA的核苷酸序列同一性为92%。该基因的特征，特别是没有内含子、获得3'聚（A）尾和侧翼直接重复序列，表明它代表一个加工假基因。在预测的氨基酸序列水平上，观察到功能重要位置的保守残基被取代，强烈表明该基因的表达不会产生功能性多肽。第二个U1 - C基因似乎也是一个假基因，因为它也没有内含子，并且与小鼠U1 - C cDNA中的ORF相比含有一个移码突变。这两个假基因的特征表明小鼠中存在U1 - C多基因家族。此外，对小鼠、人类和非洲爪蟾U1 - C的氨基酸序列比较表明，该蛋白在进化过程中高度保守。由于非洲爪蟾U1 - C与两种哺乳动物对应物仅在C末端区域的一些位置不同，可以得出结论，U1 - C的N末端区域比蛋白质的其他部分对氨基酸变化的耐受性更低。