Regulation of myc expression and apoptosis by the ubiquitin-proteasome system

C-myc has been implicated extensively in apoptosis. Numerous factors have been shown to regulate myc transcription. The ubiquitin (Ub)-proteasome system plays an important role in both Myc protein degradation and apoptosis. The data presented in this dissertation originated from the study of these three interactive players i.e. c-Myc, apoptosis, and the Ub-proteasome pathway. Two main topics are discussed in this dissertation: (1) the regulation of Myc protein level by protein kinase C (PKC) and the Ub-proteasome system and (2) activation of caspase-3 by the inhibition of proteasome.; Activation of PKC was known to decrease myc transcription. The decrease is due to transcription pausing near the first exon/intron border. We reported that PKC activation increased nuclear protein binding to MIE1, a c-myc intron 1 element that defines an RFX1-binding X box. Promoter analysis showed that the RFX1-binding X box was required for the down-regulation of reporter gene expression upon PKC activation. Increased MIE1 binding was independent of protein synthesis and was due to the nuclear translocation of RFX1. On the other hand, PKC activation does not affect the rate of Myc protein turnover. We showed in HL-60 cells the polyUb conjugates of Myc and demonstrated that Myc protein is degraded by Ub-proteasome pathway. We also showed that MycS, an endogenous and functional form of Myc that lacks the amino-terminal transactivation domain, also is degraded by the Ub-proteasome pathway.; Caspase-3 is a pivotal executioner protease in apoptosis. The active caspase-3 is a heterotetramer comprised of p12 and p17 (or p20) subunits. We demonstrated that the inhibition of proteasome caused the accumulation of p12, p17, and p20 subunits in transfected 911 cells. We also showed the polyUb conjugates of p12, p17, and p20 subunits. Cotransfection with X-linked inhibitor of apoptosis protein (XIAP) increased the accumulation of polyubiquitinated p17 and p20. Our findings suggest: (1) that the inhibition of proteasome activates caspase-3 by preserving p12, p17, or p20 subunits from degradation; and (2) that XIAP promotes the ubiquitination of active caspase-3 subunits (a novel function).