Genomic organization of the mouse pore-forming protein (perforin) gene and localization to chromosome 10. Similarities to and differences from C9.

Abstract

The molecular mechanisms involved in the delivery of cellular injury or death by CTL and NK cells have not been elucidated (1, 2). The granule-exocytosis model of cellular cytotoxicity, the most closely investigated mechanism to date, postulates that an obligate facet of the delivery of the "lethal hit" is the directional release of cytotoxic polypeptides and other molecules (the granule contents) towards the target cell (3-5). Key molecules in this process are a cytolytic protein termed perforin (5, 6), cytolysin (3), pore-forming protein (PFP)t (4) or C9-related polypeptide (C9RP) (7), and a family of killer cell-specific, highly homologous serine esterases (SE) (8,9). PFP is postulated to intercalate the target cell membrane and compromise its osmotic stability; however, the function of SEs in the cytotoxic process is less clear. Intriguing parallels have been drawn between this mechanism and the complement cascade, including the close similarities between the immunological and physicochemical properties ofPFP and C9, the ninth component ofcomplement (7,(10)(11)(12) .Many ofthe granule polypeptides have been purified to homogeneity, and cDNA clones encoding PFP (13-16) and several of the SEs (17-25) have recently been characterized. To further analyze the role of PFP in the cytolytic process and the basis for its immunological and evolutionary relationship to C9, we have investigated the This work was supported in part by U. S.

Target cell apoptosis induced by cytotoxic T cells and natural killer cells involves synergy between the pore‐forming protein, perforin, and the serine protease, granzyme B

“…Unlike C9, however, monomers of perforin alone can produce a homopolymeric transmembrane array, while C9 has an obligate requirement for prior insertion of C5b, C6, C7 and C8 into the mernb~ane.~ In addition, the functional similarities between perforin and C9 do not extend to their gene structures, which are surprisingly disparate. 8 The observations relating to perforin's actions, together with its purification and c10ning~~'~ seemed to have settled the long-standing question of cytotoxic mechanisms. However, the morphological changes induced by perforin did not account for nuclear changes such as chromatin condensation and margination, and ultimately DNA fragmentation (apoptosis) observed in cytolytic models both in vivo and in vitro.".…”

Novel putative promoter/enhancer sequences are shared by the mouse and human perforin (Pfp) genes

“…The EMBL 3 bacteriophage clone, lambda PFP-64, encoding mouse Pfp, and the PFP cDNA clones, PFP-6 and PFP-15 have been described previously (13).…”

“…In order to define the exact boundaries of the intron at the 5' end of mouse Pfp, genomic libraries were rescreened with 5' probes from cDNA clones PFP-6 and PFP-15. PFP-6 contains most of the PFP cDNA coding sequence but lacks the 5' end, while PFP-15 is an overlapping clone which extends into the 5'UT (13). Despite repeated screening of several libraries, no clone iinking Exons I and I1 could be isolated.…”

“…Despite repeated screening of several libraries, no clone iinking Exons I and I1 could be isolated. Although the validity of the 5'-most sequences of PFP-15 had been checked by RNA blot analysis (13), this analysis was repeated using a 5' fragment of PFP-15 which had been cloned separately into an unrelated plasmid vector and repurified. No signal was detected with this probe in RNA from the mouse CTL cell line, CTLL-R8 (1 8), following autoradiography for 5 days (Fig.…”

Perfil de expressão gênica de linfócitos T CD4+ na infecção pelo HTLV-1

“…Foi demonstrado que a perforina apresenta homologia na sequência e estrutura com a molécula C9 do sistema complemento (TSCHOPP et al, 1986). A partir dessa informação, pressupõe-se que a perforina promova a formação de poros na membrana que facilitam a entrada das granzimas nas células (TSCHOPP et al, 1986;PODACK et al, 1985;TRAPANI et al, 1990).…”