Human BAG-1/RAP46 protein is generated as four isoforms by alternative translation initiation and overexpressed in cancer cells

Abstract

Previously, a Bcl-2-interacting protein, BAG-1, was cloned from mouse cells and was shown to interact with several other proteins and to be important for inhibition of apoptosis. Human BAG-1 (hBAG-1) cDNA, recently isolated by us and two other groups, has been shown to be identical to a hormone receptor-binding protein, RAP46. However, di erent molecular masses of hBAG-1 protein products were noted by these three groups. Here we demonstrated that hBAG-1 protein was expressed as four isoforms, designated p50, p46, p33 and p29, with apparent molecular masses of 50 kDa, 46 kDa, 33 kDa and 29 kDa, respectively. Deletion, site-directed mutagenesis and in vitro transcription/translation analysis showed that the four protein products of hBAG-1 were expressed by alternative initiation from four di erent start codons through a leaky scanning mechanism. Furthermore, we demonstrated that the distinct forms of hBAG-1 have di erent subcellular localizations, suggesting that they may have distinct functions in the cells. Characterization of hBAG-1 RNA and protein also showed that hBAG-1 was overexpressed in human cervical, breast and lung cancer cell lines. Taken together, these data clarify the con¯icting observations reported in the literature and suggest that hBAG-1 is expressed as four forms of protein products, which may play a di erential role in apoptosis and oncogenesis of human cells.

Differentially expressed genes associated with mouse lung tumor progression

“…The decrease in Bax expression in adenocarcinomas is more signi®cant since it is expected to promote apoptosis through heterodimerization with two potent antiapoptotic genes Bcl- (Oltvai et al, 1993). High levels of BAG-1 and Bcl-2 are associated with lung cancers (Yang et al, 1998). In highly metastatic subpopulations of a human pulmonary adenocarcinoma cell line, FasL was downregulated (Gemma et al, 2001).…”

BAG‐1, An Anti‐Apoptotic Tumour Marker

“…We discovered one new isoform of BAG-1 protein (37). We demonstrated that BAG-1 protein was expressed as three major isoforms, designated p50 (50 kDa, BAG-IL), p46 (46 kDa, BAG-1M), and p33 (33 kDa, BAG-1S), and one minor isoform, p29 (29 kDa).…”

BAG-1 expression and function in human cancer

“…We initially identified a potential nuclear localisation signal (NLS) within the unique N-terminal domain of BAG-1L, consistent with the predominantly nuclear localisation of this isoform (Packham et al, 1997;Takayama et al, 1998;Yang et al, 1998;Brimmell et al, 1999). By contrast, BAG-1S and BAG-1M lack this sequence and BAG-1S is largely located in the cytoplasm whilst BAG-1M partitions between the nucleus and cytoplasm (Packham et al, 1997;Takayama et al, 1998;Yang et al, 1998). The nuclear/cytoplasmic distribution of BAG-1 isoforms is regulated under some conditions and it is important to recognise that nuclear BAG-1 immunostaining in cancer cells may not be a reliable measure of BAG-1L expression.…”

“…BAG-1 proteins are expressed as multiple isoforms generated by alternate translation initiation from a single mRNA (Packham et al, 1997;Takayama et al, 1998;Yang et al, 1998). The major human BAG-1 isoform, BAG-1S, initiates at an internal AUG codon, whereas translation of the larger BAG-1L and BAG-1M proteins initiate at upstream CUG and AUG codons, respectively.…”