Expression of CtBP family protein isoforms in breast cancer and their role in chemoresistance

Background information. CtBPs [C-terminal (of E1A) binding protein] have roles in the nucleus as transcriptional co-repressors, and in the cytoplasm in the maintenance of vesicular membranes. CtBPs are expressed from two genes, CTBP1 and CTBP2, mRNA products of which are alternatively spliced at their 5'-ends to generate distinct protein isoforms. Extensive molecular and cellular analyses have identified CtBPs as regulators of pathways critical for tumour initiation, progression and response to therapy. However, little is known of the expression or regulation of CtBP isoforms in human cancer, nor of the relative contributions of CTBP1 and CTBP2 to the tumour cell phenotype. Results. Expression of CtBP proteins and CTBP1 and CTBP2 mRNA splice forms in breast cancer cell lines and tumour tissue was examined. CtBP1 proteins are identifiable as a single band on Western blots and are ubiquitously detectable in breast tumour samples, by both Western blotting and immunohistochemistry. CtBP1 is present in six of six breast cancer cell lines, although it is barely detectable in SKBr3 cells due to reduced CTBP1 mRNA expression. In the cell lines, the predominant CTBP1 mRNA splice form encodes CtBP1-S protein; in tumours, both major CTBP1 mRNA splice forms are variably expressed. CtBP2 proteins are ubiquitously expressed in all lines and tumour samples. The predominant CTBP2 mRNA encodes CtBP2-L, although an alternatively spliced form that encodes CtBP2-S, previously unidentified in humans, is expressed at low abundance. Both CtBP2-L and CtBP2-S are readily detectable as two distinct bands on Western blots; here we show that the CTBP2-L mRNA is translated from two AUG codons to generate both CtBP2-L and CtBP2-S. We have also identified an autoregulatory feedback mechanism whereby CtBP protein abundance is maintained in proliferating breast cancer cells through the post-transcriptional regulation of CtBP2. This feedback is disrupted by UV-C radiation or exposure to cisplatin. Finally, we demonstrate that CtBP1 and CtBP2 both have p53-dependent and -independent roles in suppressing the sensitivity of breast cancer cells to mechanistically diverse cancer chemotherapeutic agents. Conclusions. These studies support recent evidence that CtBP family proteins represent potential targets for therapeutic strategies lior the treatment of cancer in general, and breast cancer in particular.