SAFB2, a New Scaffold Attachment Factor Homolog and Estrogen Receptor Corepressor

Abstract

We have characterized previously the nuclear matrix protein/scaffold attachment factor (SAFB) as an estrogen receptor corepressor and as a potential tumor suppressor gene in breast cancer. A search of the human genome for other potential SAFB family members revealed that KIAA00138 (now designated as SAFB2) has high homology to SAFB (now designated as SAFB1). SAFB1 and SAFB2 are mapped adjacent to each other on chromosome 19p13.3 and are arranged in a bidirectional divergent configuration (head to head), being separated by a short (<500 bp) GC-rich intergenic region that can function as a bidirectional promoter. SAFB1 and SAFB2 share common functions but also have unique properties. As shown previously for SAFB1, SAFB2 functions as an estrogen receptor corepressor, and its overexpression results in inhibition of proliferation. SAFB1 and SAFB2 interact directly through a C-terminal domain, resulting in additive repression activity. They are coexpressed in a number of tissues, but unlike SAFB1, which is exclusively nuclear, SAFB2 is found in the cytoplasm as well as the nucleus. Consistent with its cytoplasmic localization, we detected an interaction between SAFB2 and vinexin, a protein involved in linking signaling to the cytoskeleton. Our findings suggest that evolutionary duplication of the SAFB gene has allowed it to retain crucial functions, but also to gain novel functions in the cytoplasm and/or nucleus.Scaffold attachment factor B (SAFB, 1 also named HET and HAP in the literature) was originally identified based on its ability to bind to scaffold/matrix attachment regions (1) and as a protein binding to the small heat shock protein hsp27 gene promoter (2). SAFB is a large protein (130 kDa) with a number of putative functional domains. The function of many of these domains is still unclear but can be inferred from the roles of similar domains in other proteins. The N terminus contains an SAF box (3) (also called an SAP domain (4)), which is a homeodomain-like DNA binding motif. This motif is believed to play a role in chromatin organization and specifically in organizing the interaction between nuclear matrix proteins and scaffold/ matrix attachment regions. SAF boxes are found in proteins involved in very diverse processes such as transcription, RNA processing, apoptotic chromatin degradation, and DNA repair (3). Amino acids 409 -484 harbor an RNA recognition motif (RRM), which is often found in mRNA-processing proteins. SAFB can interact with a number of proteins from the RNAprocessing machinery, such as AUF1/hnRNP D, hnRNP A1, htra2-␤1, ASF/SF2, SRp30c, and CLK2 (5-7). Consistent with this, SAFB is able to alter the splice site selection of an E1A minigene (7). Because SAFB was also shown to interact with the C-terminal domain of RNA polymerase II, it has been suggested to be part of a "transcriptosome" complex, coupling transcription and RNA processing (7). SAFB has a nuclear localization signal (NLS), and using biochemical fractionation experiments, we and others have shown that SAFB is a nuclear protei...

The role of nuclear organization in cancer

“…The SAFB1 and SAFB2 genes are located in chromosome band 19p13, where they are coordinately regulated by a bidirectional promoter. This is one of the most frequently deleted genomic regions in breast cancer, and poor prognosis tumours have low expression of SAFB1 and SAFB2 [70][71][72]. Experimental over-expression of SAFB1 and SAFB2 in breast cancer cells causes growth arrest and aneuploidy [71].…”

Novel role of the RET finger protein in estrogen receptor-mediated transcription in MCF-7 cells

“…The yeast two-hybrid screen and interaction tests were undertaken using the Matchmaker3 system (BD Biosciences) with the appropriate controls according to the manufacturer's instructions as previously described [8,9]. The bait and test constructs were cotransformed into yeast strain AH109 which was then plated onto SD-Leu/Trp and SD-Leu/Trp/His/Ade medium containing X-α-gal as a substrate for α-galactosidase.…”

“…Immunohistochemistry was performed according to Townson et al [9] using anti-RFP antibodies (IBL America).…”

“…GST pulldown experiments were performed as recently described [8,9]. MCF-7 cells stabley expressing HA-RFP were used to coimmunoprecipitate RFP, ESR1, and SAFB.…”

“…We have previously shown that the members of the scaffold attachment factor B family, termed SAFB1 and SAFB2, can function as potent ESR1 corepressors [8,9]. SAFB1/2 are nuclear matrix proteins characterized by an N-terminal DNA-binding domain (SAF-box) that binds to scaffold/matrix attachment regions (S/MARs), and a central RNA recognition motif (RRM) [10].…”