The candidate tumour suppressor, XIAP associated factor 1 (XAF1), directly inhibits XIAP activity and induces G1 phase cell cycle arrest.

摘要

X&barbelow;IAP a&barbelow;ssociated f&barbelow;actor 1&barbelow; (XAF1) was initially isolated as novel 34 kDa protein which bound XIAP in a two-hybrid screening. The XAF1A protein consists of 301 a.a. and contains seven potential zinc finger domains. Two alternatively splice variants of XAF1 were later isolated. One isoform (XAF1B) was formed by the removal of a 57 bp exon, which leads to an in-frame deletion of the third zinc finger and the creation of a shorter 32.5 kDa protein. The other splice variant (XAF1C) contains a 154 bp exon insertion, which truncates the sixth and seventh zinc fingers to produce an 18.7 kDa protein. XAF1A and XAF1B, but not XAF1C, bound XIAP in in vitro pull down assays. Northern blot analysis showed at least four distinct sizes of xaf1 mRNA ranging between 3.9 and 7.0 kb, which may indicate other XAF1 isoforms yet to be discovered.; Though the possible role of these zinc fingers on the XAF1/XIAP interaction has yet to be determined, recent experiments indicate that XAF1A can block the ability of XIAP to inhibit caspase-3 in vitro. Furthermore, overexpression of XAF1A in HEL299 cells triggered a G1 cell cycle arrest. This G1 arrest coincides with an increase in p21, but not p53. The ability of XAF1 to block XIAP function and induce cell cycle arrest suggests a role for XAF1 in the control of both apoptosis and cell growth.; The coding regions of XAF1A, B and C are encoded on a total of 9 exons within a span of 20 kb. The single copy xaf1 gene has been mapped, using FISH analysis, distal to the TP53 gene on 17p13.2. Southern blot analysis of YACS within this region further localizes the xaf1 gene on YAC 746 C 10, which contains the markers D17S1831, D17S796, and D17S1881. These markers are located approximately 3 cM telomeric to the TP53 gene. Since the xaf1 gene is located in a region commonly deleted in numerous types of cancers, this may suggest a tumour suppressor role for XAF1 in cancer. To test this theory, a 60 cell line panel from the NCl was analyzed for xaf1 RNA expression by Taqman and heterozygosity status of markers proximal to xaf1. Taqman analysis indicated that the majority of cell lines expressed little or no xaf1 RNA while xiap levels were relatively high. A PCR study of markers near xaf1 showed significant loss of heterozygosity (LOH) in this region. The loss of xaf1 expression and significant LOH near the xaf1 gene indicate that the down-regulation of XAF1 may be important in the development of the transformed phenotype.