Little is known about the nature of recombination hotspots in the human genome and the relationship between crossover activity and patterns of linkage disequilibrium. We have therefore used both haplotype analysis and direct detection of crossovers in sperm to characterize a putative recombination hotspot in the TAP2 gene within the class II region of the MHC. Haplotype diversity provided evidence for a localized hotspot within intron 2 of this gene. Sperm DNA typing using allele-specific PCR primers to selectively amplify recombinant TAP2 molecules revealed a highly localized meiotic crossover hotspot approximately 1.2 kb long, unusually abundant in sequence polymorphisms and flanked by DNA much less active in recombination. Sperm crossover appeared to be fully reciprocal, and almost all crossover products were simple, involving a single exchange between adjacent heterozygous markers. This hotspot appears to be much more active in female than male meiosis. No primary sequence similarities could be found between any of the very few well defined crossover hotspots in the human genome, all of which show recombinationally active domains 1-2 kb long. Direct comparison of recombination frequency and haplotype diversity in TAP2 showed that linkage disequilibrium measures were a poor predictor of crossover frequency in this region, with non-recombining markers sometimes in free association and with examples of pairs of markers spanning the recombination hotspot showing substantial or even absolute linkage disequilibrium.

PAR bZIP (cells knockout for PAR bZIP transcription factors) proteins, thyrotroph embryonic factor (TEF), albumin D-site-binding protein (DBP), and hepatic leukemia factor (HLF), are a family of transcription factors that have been shown to contribute to the expression of genes involved in detoxification and drug metabolism. Recently, we showed that PAR bZIP proteins were able to regulate the BH3-only gene bcl-gS in tumor cells. Here, we have extended the role of these transcription factors in the control of apoptosis executors by analyzing the expression of BH3-only genes in PAR bZIP triple knockout mouse fibroblasts. We found that bik was the only BH3-only gene downregulated in knockout cells. Consistently, transfection of TEF or DBP induces the expression of endogenous bik, regardless of the presence of active p53. Moreover, both promoter-reporter and chromatin immunoprecipitation assays indicate that PAR bZIP proteins activate the bik promoter directly. Treatment with different stress stimuli reveals a higher survival of knockout fibroblasts compared with that of wild-type cells, especially after incubation with H 2 O 2 , which suggest that PAR bZIP proteins participate in oxidative stress-induced apoptosis. Furthermore, the apoptotic cell death promoted by treatment with H 2 O 2 can be impaired by reducing the expression of Bik in wild-type fibroblasts or enhanced by the overexpression of Bik in knockout cells. These findings reveal a novel transcriptional pathway relevant in transducing the apoptotic response to oxidative stress.

Proline-and acid-rich (PAR) basic region leucine zipper (bZIP) proteins thyrotroph embryonic factor (TEF), D-sitebinding protein (DBP), and hepatic leukemia factor have been involved in neurotransmitter homeostasis and amino acid metabolism. Here we demonstrate a novel role for these proteins in the transcriptional control of a BH3-only gene. PAR bZIP proteins are able to transactivate the promoter of bcl-gS. This promoter is particularly responsive to TEF activation and is silenced by NFIL3, a repressor that shares the consensus binding site with PAR bZIP proteins. Consistently, transfection of TEF induces the expression of endogenous bcl-gS in cancer cells, and this induction is independent of p53. A naturally occurring variant of DBP (tDBP), lacking the transactivation domain, has been identified and shown to impede the formation of active TEF dimers in a competitive manner and to reduce the TEF-dependent induction of bcl-gS. Of note, treatment of cancer cells with etoposide induces TEF activation and promotes the expression of bcl-gS. Furthermore, blockade of bcl-gS or TEF expression by a small interfering RNA strategy or transfection with tDBP significantly reduces the etoposide-mediated apoptotic cell death. These findings represent the first described role for PAR bZIP proteins in the regulation of a gene involved in the execution of apoptosis.Two genes, cell death specification protein 1 (ces-1) and ces-2, control the decisions of neuro-secretory motor sister cells in Caenorhabditis elegans to undergo apoptosis. A genetic approach showed that these factors regulate the genes required for apoptosis and that a gain of ces-1 function or a reduction of ces-2 function prevents these cells from dying (1). The pro-apoptotic CES-2 protein negatively regulates CES-1, which prevents the death of the neuro-secretory motor sister cells by transcriptional silencing of Egl-1, a BH3-only protein required for apoptosis in C. elegans (2, 3). CES-2 is similar to members of the proline-and acid-rich (PAR) 4 subfamily of basic region leucine zipper (bZIP) transcription factors, and both share the DNA binding specificity (4).Mammalian homologs of CES-2 include thyrotroph embryonic factor (TEF), albumin D-site-binding protein (DBP), and hepatic leukemia factor (HLF) (5-7). These PAR bZIP proteins have recently been shown to be involved in amino acid and neurotransmitter metabolism through transcriptome profiling analyses in both liver and brain (8). Thus, despite the pro-apoptotic activity described for CES-2 promoting the induction of Egl-1 through an indirect pathway, none of its human homologs have been associated with the transcriptional regulation of BH3-only genes or other executors of apoptosis.BH3-only proteins are a pro-apoptotic subgroup of the Bcl-2 family of apoptosis regulators, which share only the short BH3 region with the rest of the family (9). Genetic experiments have shown that these proteins are essential initiators of programmed cell death in species as distantly related as mice and C. elegans. They are r...