Structure of the Mouse Pore-forming Protein (Perform) Gene: Analysis of Transcription Initiation Site, 5' Flanking Sequence, and Alternative Splicing of 5' Untranslated Regions

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 PFR7, 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 1, except for being 107 by 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 N7 kb. The first exon contains 79 by of 5' UTR, the second exon contains 143 or 36 by of 5' UTR (type I or type II UTR, respectively) plus the NHz-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 mousePfp 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-y-, and UVresponsive 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 .