Youn B S, Liu C C, Kim K K, Young J D, Kwon M H, Kwon B S
Department of Microbiology and Immunology, Indiana University School of Medicine, Indianapolis 46202.
J Exp Med. 1991 Apr 1;173(4):813-22. doi: 10.1084/jem.173.4.813.
We studied the 5' untranslated regions (UTRs) of the mouse lymphocyte pore-forming protein (PFP, perforin, and cytolysin). 5' UTRs were determined by primer extension analysis, sequencing PFP cDNA clone PFP-7, ribonuclease protection assays, and amplification of poly(A)+ RNA of cytolytic T lymphocyte using polymerase chain reaction (PCR). Two alternatively spliced 5' UTRs, designated type I and type II, of 222 and 115 bp, respectively, were found associated with PFP. Type II is identical to type I, except for being 107 bp shorter in the second exon. This deletion was generated by the use of alternative acceptor splice sites. The mouse PFP gene (Pfp) encodes three exons, is separated by two small introns, and spans a chromosomal region of approximately 7 kb. The first exon contains 79 bp of 5' UTR, the second exon contains 143 or 36 bp of 5' UTR (type I or type II UTR, respectively) plus the NH2-terminal region of the mouse PFP, and the third exon contains the rest of the COOH-terminal mouse PFP. The organization of the mouse Pfp is similar to that of the human gene. Moreover, the 5' flanking sequence of the mouse Pfp is highly homologous to that of the human Pfp. In contrast to the human sequence, the more immediate 5' flanking sequence of mouse Pfp contains two tandem "TATA" box-related elements and a GC box, but lacks a typical CAAT box-related sequence. Several other enhancer elements were found further upstream, including cAMP-, phorbol ester-, interferon-gamma-, and UV-responsive elements, and PU box-like and NFkB binding site-like elements. In addition, we found a nuclear inhibitory protein-like element, a transcriptional silencer, and a pair of purine-rich sequence motifs that were found in other T cell-specific genes, and three repeats of GGCCTG that may be a variation of a highly repetitious GCCCTG consensus sequence found in human Pfp.
我们研究了小鼠淋巴细胞孔形成蛋白（PFP，穿孔素和细胞溶素）的5'非翻译区（UTR）。通过引物延伸分析、对PFP cDNA克隆PFP-7进行测序、核糖核酸酶保护分析以及使用聚合酶链反应（PCR）扩增细胞毒性T淋巴细胞的聚腺苷酸加尾RNA（poly(A)+ RNA）来确定5'UTR。发现与PFP相关的两种选择性剪接的5'UTR，分别命名为I型和II型，长度分别为222和115 bp。II型与I型相同，只是第二个外显子短107 bp。这种缺失是由于使用了替代的剪接受体位点产生的。小鼠PFP基因（Pfp）编码三个外显子，由两个小内含子隔开，跨越约7 kb的染色体区域。第一个外显子包含79 bp的5'UTR，第二个外显子包含143或36 bp的5'UTR（分别为I型或II型UTR）加上小鼠PFP的NH2末端区域，第三个外显子包含小鼠PFP其余的COOH末端。小鼠Pfp的结构与人基因的结构相似。此外，小鼠Pfp的5'侧翼序列与人Pfp的高度同源。与人类序列不同，小鼠Pfp更直接的5'侧翼序列包含两个串联的“TATA”盒相关元件和一个GC盒，但缺乏典型的CAAT盒相关序列。在更上游还发现了其他几个增强子元件，包括cAMP、佛波酯、干扰素-γ和紫外线反应元件，以及PU盒样和NFkB结合位点样元件。此外，我们发现了一个核抑制蛋白样元件、一个转录沉默子以及在其他T细胞特异性基因中发现的一对富含嘌呤的序列基序，还有三个GGCCTG重复序列，它们可能是人类Pfp中发现的高度重复的GCCCTG共有序列的变体。
