Splice variants of cytosolic polyadenylation element–binding protein 2 (CPEB2) differentially regulate pathways linked to cancer metastasis

Abstract

The translational regulator cytosolic polyadenylation element-binding protein 2 (CPEB2) has two isoforms, CPEB2A and CPEB2B, derived by alternative splicing of RNA into a mature form that either includes or excludes exon 4. Previously, we reported that this splicing event is highly dysregulated in aggressive forms of breast cancers, which overexpress CPEB2B. The loss of CPEB2A with a concomitant increase in CPEB2B was also required for breast cancer cells to resist cell death because of detachment (anoikis resistance) and metastasize To examine the mechanism by which CPEB2 isoforms mediate opposing effects on cancer-related phenotypes, we used next generation sequencing of triple negative breast cancer cells in which the isoforms were specifically down-regulated. Down-regulation of the CPEB2B isoform inhibited pathways driving the epithelial-to-mesenchymal transition and hypoxic response, whereas down-regulation of the CPEB2A isoform did not have this effect. Examining key nodes of these pathways showed that CPEB2B induced the expression of regulatory DNA-factors ( HIF1α and TWIST1). Specifically, CPEB2B functioned as a translational activator of TWIST1 and HIF1α. Functional studies showed that specific down-regulation of either HIF1α or TWIST1 inhibited the ability of CPEB2B to induce the acquisition of anoikis resistance and drive metastasis. Overall, this study demonstrates that CPEB2 alternative splicing is a major regulator of key cellular pathways linked to anoikis resistance and metastasis.

Tumor suppressor role of cytoplasmic polyadenylation element binding protein 2 (CPEB2) in human mammary epithelial cells

“…The ratio of A/B isoforms (ratios of mean of ΔCT) was higher in non-tumor tissues (Fig. 8a), supporting the reports that A is the tumor-suppressor isoform and B the tumor-promoter isoform [29, 30]. The information on ER, PR and HER2 status (n value) was available in 98 out of 105 samples as follows: 19 HER2+, 64 HER2-, 75 ER+, 18 ER-, 64 PR+, 29 PR-, 11 ER/PR/HER+ and 10 ER/PR/HER2-.…”

“…The roles of CPEB2 in human breast tumorigenesis have so far remained a paradox, until isoform-specific roles were identified [29, 30]. Selecting cells from TNBC cell lines for anioikosis-resistance, these authors observed that alternative splicing resulting in the loss of CPEB2A with concomitant increase in CPEB2B was responsible for their metastatic phenotype.…”

“…CPEB2 was shown to suppress the translation of two oncogenic transcription factors, TWIST1 [27] - an EMT inducer, and HIF1α [24] associated with many oncogenic functions [24, 25]. Deligio et al [30] reported that anoikosis-resistance and metastasis phenotype of TNBC cell lines resulted from CPEB2B isoform mediated translational activation of HIF1α and TWIST1. They suggested that CPEB2B plays an antagonistic role against CPEB2A by alleviating the translational inhibition of HIF1α and TWIST1 imparted by CPEB2A.…”

“…TWIST1 can mediate EMT [48], and also inhibit apoptosis through evasion of p53-induced cell death [49]. CPEB2 mediated translational repression of TWIST1, a function ascribed to the A isoform [30] could be a mechanism by which EMT is suppressed. In support, in CPEB2KO cells exhibited no change in TWIST1 mRNA expression, suggesting post- transcriptional regulation by CPEB2.…”

“…They suggested that isoform A which excludes exon 4 is a tumor- suppressor, whereas isoform B that includes exon 4, is a tumor-promoter. This suggestion was validated by next generation sequencing and isoform-specific down-regulation of CPEB2A and B in TNBC lines [30]. They concluded that CPEB2B plays an antagonistic role against CPEB2A by alleviating the translational inhibition of HIF-1α and Twist 1 imparted by CPEB2A.…”