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Caspase-2(S) is targeted by Nonsense-Mediated Decay

Benjamin O'Connor

Appointment Period: 2008-2010, Grant Years: [24,25]

Benjamin O'ConnorResearch has demonstrated that Caspase-2(L) can act as a tumor suppressor in a murine lymphoma model. The Caspase-2 gene encodes two distinct mRNA transcripts: Caspase-2(S) and Caspase-2(L). We demonstrated in multiple human/murine cancer cell lines and mouse tissues that Caspase-2(L) mRNA transcripts are present at much higher levels than those of the truncated, alternative Caspase-2 mRNA, Caspase-2(S). We hypothesized that the low levels of Caspase-2(S) transcript were the result of Caspase-2(S) being targeted by a cellular RNA disposal system known as Nonsense-Mediated Decay (NMD) due to the presence of a pre-mature termination codon in the transcript. We assessed the Caspase-2(S) mRNA expression levels after applying multiple chemicals and shRNA constructs known to knock down and inhibit the expression of molecular components of NMD. In the case of each shRNA knockdown construct or NMD-inhibiting chemical, Caspase-2(S) mRNA levels were significantly higher in treated samples than in control samples. We were able to confirm the NMD-targeting of Caspase-2(S) by replicating our experiments with an additional construct designed to express Green-Fluorescent-Protein (GFP) when NMD is inhibited. Every treatment condition that resulted in an increase of Caspase-2(S) also elicited a green signal, indicating that our experimental conditions were inhibiting NMD’s action. Identifying Caspase-2(S) as an NMD-regulated transcript is important due to Caspase-2(L)’s potential role in cancer. Inhibition of NMD’s surveillance and disposal of Caspase-2(S) transcript results in an increase in of Caspase-2(S) mRNA. Increasing transcript levels could potentially increase the Caspase-2(S) protein sufficiently enough to interfere in a dominant negative manner with Caspase-2(L)’s dimerization/cleavage process, and to potentially inhibit tumor suppression.

O’Connor BF. Regulation of Caspase-2S Isoform Production By Nonsense-Mediated Decay. (2012) PhD Thesis. In preparation.