Emetine regulates the alternative splicing of caspase 9 in tumor cells

Abstract

Abstract. Exons 3 to 6 in the caspase 9 gene undergo alternative splicing in which the larger caspase 9 splice variant promotes apoptosis, in contrast to the dominant negative anti-apoptotic splice variant, the smaller caspase 9b. In this study, the regulation of the alternative splicing of caspase 9 pre-mRNA was examined in response to Emetine. Treatment of C33A cells, breast cancer MCF-7 cells and MCF-7/Adr cells with Emetine dihydrochloride upregulated the level of smaller caspase 9b mRNA and concomitantly decreased the mRNA level of larger caspase 9 in a dose-and time-dependent manner, indicating that Emetine desensitizes C33A, MCF-7 and MCF-7/Adr to cell death. In contrast, treatment of PC3 cells, a prostate cancer cell line, manifested an opposite effect: a greater production of the larger caspase 9 mRNA with a concomitant decrease of caspase 9b mRNA. Pretreatment with calyculin A, an inhibitor of protein phosphatase 1 (PP1) and protein phosphatase 2A (PP2A) blocked Emetine-induced alternative splicing in cells, in contrast to okadaic acid, a specific inhibitor of PP2A, demonstrating a PP1-mediated mechanism. These results suggest that the various splicing patterns of the caspase 9 gene that are regulated by chemotherapy reagents may contribute to the resistance or sensitization of the tumors to other cell death inducers.

Alternative splicing in human cancer cells is modulated by the amiloride derivative 3,5‐diamino‐6‐chloro‐N‐(N‐(2,6‐dichlorobenzoyl)carbamimidoyl)pyrazine‐2‐carboxide

“…By contrast, pretreatment with the PP1 inhibitor OA could partially restore the effects of BS008 on the phosphorylation levels of SRSF3 as well as the AS of HIPK3 and SMAC transcripts. These results are similar to those of other reports that revealed that 4β‐hydroxywithanolide E (Lee et al ., ), ceramide (Chalfant et al ., ; Massiello et al ., ), emetine (Boon‐Unge et al ., ; Pan et al ., ), and epigallocatechin‐3‐gallate/ibuprofen can modulate (Kim, ) the AS of apoptotic gene transcripts through a PP1‐mediated splicing mechanism. These publications imply that PP1 plays a crucial role in regulating the AS of apoptotic gene transcripts.…”

Modulation of alternative splicing by anticancer drugs

“…In another study, Gemticabine was found to downregulate the expression of caspase‐9b in A549 cells . Emetine was found to regulate caspase‐9 splicing showing differential splicing pattern in different cell types …”

“…Calyculin A, an inhibitor of both PP1 and PP2A, was found to block the emitine effects on caspase‐9 splicing in C33A and PC3 cells, while okadaic acid which is a selective inhibitor of PP2A showed no effect. Hence, it was concluded that emitine exerts its affect through PP1‐dependent mechanism . However, emitine showed opposite effect on caspase‐9 splicing in C33A and PC3 cells .…”

“…PP1 affects the phosphorylation of SR proteins and hence control their activity (Figure ). Emitine, a protein synthesis inhibitor in eukaryotes, was found to regulate alternative splicing of Bcl‐x and caspase‐9 through phosphorylation . Calyculin A, an inhibitor of both PP1 and PP2A, was found to block the emitine effects on caspase‐9 splicing in C33A and PC3 cells, while okadaic acid which is a selective inhibitor of PP2A showed no effect.…”

Pharmacology of Modulators of Alternative Splicing

“…Whereas caspase 8 is normally proapoptotic, an intron retention forms Cas8L with antiapoptotic properties (Himeji et al, 2002). Skipping of exons 3, 4, 5, and 6 in caspase 9 results in a shorter protein that also has antiapoptotic properties—this event being regulated by SRSF1 and SRSF2 (Shultz et al, 2010; Pan et al, 2011). Yet another type of AS—5′ alternative splice sites—is responsible for a proapoptotic isoform of the well-known survival factor Bcl-xL, termed Bcl-xS (Cloutier et al, 2008).…”