SPBP (stromelysin-1 platelet-derived growth factorresponsive element binding protein) was originally cloned from a cDNA expression library by virtue of its ability to bind to a platelet-derived growth factor-responsive element in the human stromelysin-1 promoter. A 937-amino acid-long protein was deduced from a 3995-nucleotide murine cDNA sequence. By analyses of both human and murine cDNAs, we now show that SPBP is twice as large as originally found. The human SPBP gene contains six exons and is located on chromosome 22q13.1-13.3. Two isoforms differing in their C termini are expressed due to alternative splicing. PCR analyses of multitissue cDNA panels showed that SPBP is expressed in most tissues except for ovary and prostate. Functional mapping revealed that SPBP is a nuclear, multidomain protein containing an N-terminal region with transactivating ability, a novel type of DNA-binding domain containing an AT hook motif, and a bipartite nuclear localization signal as well as a C-terminal zinc finger domain. This type of zinc finger domain is also found in the trithorax family of chromatin-based transcriptional regulator proteins. Using cotransfection experiments, we find that SPBP enhances the transcriptional activity of various transcription factors such as c-Jun, Ets1, Sp1, and Pax6. Hence, SPBP seems to act as a transcriptional coactivator.SPBP (stromelysin-1 PDGF 1 -responsive element-binding protein) was originally isolated from a murine gt11 cDNA library as a protein that bound to the stromelysin-1 PDGFresponsive element (SPRE) of the human stromelysin-1 promoter (1). The published cDNA sequence of SPBP is 3995 base pairs (bp) long with an open reading frame (ORF) of 2822 bp encoding a protein of 937 amino acids (1). SPBP has also been called AR1 or TCF20 (2). Stromelysin-1, also known as MMP3, is an extracellular matrix-degrading metalloproteinase that is active against a broad range of substrates. These proteases are invariably up-regulated in epithelial cancers, recognized as targets of oncogenic signal transduction pathways and shown to contribute to tumor invasion and metastasis (3, 4). Recently, it was reported that stromelysin-1 actually also promoted mammary carcinogenesis in a mouse model system. Overexpression of stromelysin-1 in the mammary gland of transgenic mice induced all stages of tumor progression from hyperplasia to malignant carcinomas (5). In such experiments, stromelysin-1 seems to act as a natural tumor promoter enhancing cancer susceptibility (6).The transcriptional activity of the stromelysin-1 promoter is stimulated by PDGF, and SPBP is reported to contribute to this up-regulation (7,8). Interestingly, the expression of SPBP itself was also found to be induced by PDGF or serum (1). The stromelysin-1 promoter contains three elements that are important for induction by mitogenic stimuli. These are an AP-1 element binding the c-Fos/c-Jun transcription factors, two head-to-head PEA3 elements binding Ets family transcription factors, and the SPRE element that binds SPBP (see Ref. 7 ...

SPBP (Stromelysin-1 PDGF responsive element binding protein) is a ubiquitously expressed 220 kDa nuclear protein shown to enhance or repress the transcriptional activity of various transcription factors. A yeast two-hybrid screen, with the extended plant homeodomain (ePHD) of SPBP as bait, identified TopBP1 (topoisomerase II β-binding protein 1) as a candidate interaction partner of SPBP. TopBP1 has eight BRCA1 carboxy-terminal (BRCT) domains and is involved in DNA replication, DNA damage responses and in the regulation of gene expression. The interaction between SPBP and TopBP1 was confirmed in vitro and in vivo, and was found to be mediated by the ePHD domain of SPBP and the BRCT6 domain of TopBP1. Both SPBP and TopBP1 enhanced the transcriptional activity of Ets1 on the c-myc P1P2- and matrix metalloproteinase-3 (MMP3) promoters. Together they displayed a more than additive effect. Both proteins were associated with these promoters. The involvement of TopBP1 was dependent on the serine 1159 phosphorylation site, known to be important for transcriptional activation. Depletion of endogenous SPBP by siRNA treatment reduced MMP3 secretion by 50% in phorbol ester-stimulated human fibroblasts. Taken together, our results show that TopBP1 and SPBP interact physically and functionally to co-operate as co-activators of Ets1.

Our genome is assembled into and array of highly dynamic nucleosome structures allowing spatial and temporal access to DNA. The nucleosomes are subject to a wide array of post-translational modifications, altering the DNA-histone interaction and serving as docking sites for proteins exhibiting effector or “reader” modules. The nuclear proteins SPBP and RAI1 are composed of several putative “reader” modules which may have ability to recognise a set of histone modification marks. Here we have performed a phylogenetic study of their putative reader modules, the C-terminal ePHD/ADD like domain, a novel nucleosome binding region and an AT-hook motif. Interactions studies in vitro and in yeast cells suggested that despite the extraordinary long loop region in their ePHD/ADD-like chromatin binding domains, the C-terminal region of both proteins seem to adopt a cross-braced topology of zinc finger interactions similar to other structurally determined ePHD/ADD structures. Both their ePHD/ADD-like domain and their novel nucleosome binding domain are highly conserved in vertebrate evolution, and construction of a phylogenetic tree displayed two well supported clusters representing SPBP and RAI1, respectively. Their genome and domain organisation suggest that SPBP and RAI1 have occurred from a gene duplication event. The phylogenetic tree suggests that this duplication has happened early in vertebrate evolution, since only one gene was identified in insects and lancelet. Finally, experimental data confirm that the conserved novel nucleosome binding region of RAI1 has the ability to bind the nucleosome core and histones. However, an adjacent conserved AT-hook motif as identified in SPBP is not present in RAI1, and deletion of the novel nucleosome binding region of RAI1 did not significantly affect its nuclear localisation.