Fos family proteins form stable heterodimers with Jun family proteins, and each heterodimer shows distinctive transactivating potential for regulating cellular growth, differentiation, and development via AP-1 binding sites. However, the molecular mechanism underlying dimer specificity and the molecules that facilitate transactivation remain undefined. Here, we show that BAF60a, a subunit of the SWI⅐SNF chromatin remodeling complex, is a determinant of the transactivation potential of Fos/Jun dimers. BAF60a binds to a specific subset of Fos/Jun heterodimers using two different interfaces for c-Fos and c-Jun, respectively. Only when the functional SWI⅐SNF complex is present, can c-Fos/c-Jun (high affinity to BAF60a) but not Fra-2/JunD (no affinity to BAF60a) induce the endogenous AP-1-regulated genes such as collagenase and c-met. These results indicate that a specific subset of Fos/Jun dimers recruits SWI⅐SNF complex via BAF60a to initiate transcription.Transcription factor AP-1, which plays pivotal roles in cell growth, differentiation, development, and tumor formation, is composed of Fos family proteins (Fos; c-Fos, Fra-1, Fra-2, and FosB) and Jun family proteins (Jun; c-Jun, JunB, and JunD). The members of both families form dimers through a leucine zipper structure; Jun family members can form low-affinity homodimers and high affinity heterodimers with the Fos family (1, 2). However, members of the Fos family do not form stable homodimers. Although these hetero-and homodimers bind to similar sites on DNA (TGAC/GTCA, AP-1 binding sites) through the basic domains of both proteins, each dimer has a distinct transcriptional regulatory function that can be modulated either positively or negatively (3). For example, transcriptional activity of the c-Fos/c-Jun dimer is much higher than the Fra-2/c-Jun dimer. Although functional interactions between some members of the Fos/Jun family of proteins and adaptor proteins such as CREB-binding protein (CBP) or TATA-binding protein (TBP) have been reported (4, 5), crucial proteins that recognize the dimer specificity and/or facilitate the induction of transcriptional initiation were largely unknown. Therefore, we proposed here to isolate such a crucial protein using a yeast two-hybrid system and have demonstrated that BAF60a (6), a component of the SWI⅐SNF chromatin remodeling complex, can select specific Fos/Jun dimers and function as a determinant of transcriptional activation via AP-1 binding sites. EXPERIMENTAL PROCEDURESPlasmid Construction-The Gal4DBD-c-Jun (25-187) fusion construct (pDBLeu-cJ(25-187)) was generated by inserting the 0.49 kilobase pair EheI fragment of the rat c-jun gene into the MscI restriction endonuclease cleavage site within the open reading frame of Gal4DBD. For the construction of template DNA for in vitro translation, we first generated a starter plasmid from pGEM2-475/Jun-D, the translation initiation site of which has a Kozak consensus sequence with a unique NcoI site that is preceded by a fragment 475 sequence for an efficient translational ...

We show here that murine leukemia virus-based retrovirus vector transgene expression is rapidly silenced in human tumor cell lines lacking expression of Brm, a catalytic subunit of the SWI/SNF chromatin remodeling complex, even though these vectors can successfully enter, integrate, and initiate transcription. We detected this gene silencing as a reduction in the ratio of cells expressing the exogenous gene rather than a reduction in the average expression levels, indicating that downregulation occurs in an all-or-none manner. Retroviral gene expression was protected from silencing and maintained in Brm-deficient host cells by exogenous expression of Brm but not BRG1, an alternative ATPase subunit in the SWI/SNF complex. Introduction of exogenous Brm to these cells suppressed recruitment of protein complexes containing YY1 and histone deacetylase (HDAC) 1 and 2 to the 5-long terminal repeat region of the integrated provirus, leading to the enhancement of acetylation of specific lysine residues in histone H4 located in this region. Consistent with these observations, treatment of Brm-deficient cells with HDAC inhibitors but not DNA methylation inhibitors suppressed retroviral gene silencing. These results suggest that the Brmcontaining SWI/SNF complex subfamily (trithorax-G) and a complex including YY1 and HDACs (Polycomb-G) counteract each other to maintain transcription of exogenously introduced genes.

Bloodstained threads (1 cm in length) were tested to identify human origin by a direct ELISA-ABC method using biotinylated antibody against human HbA0. By this method human bloodstains were clearly distinguishable from bloodstains of other species including Japanese monkey. The minimum detection limit of bloodstains prepared from undiluted human whole blood was 1:5, 120 (28 ng Hb) and that of bloodstains from diluted human whole blood was 1:640-1:1, 280. Human Hb was more easily detectable in bloodstains prepared from diluted human blood after extraction with 5% ammonia than after extraction with phosphate-buffered saline.