Programming cellular gene expression by IKK mediated NF-kappaB activation.

摘要

The NF-kappaB transcription factor family is a well-characterized group of proteins known to be essential for the initiation and maintenance of an immune response, as well as the anti-apoptotic/survival cascade. Deregulation of this pathway has been implicated in a number of inflammatory diseases and in cancer progression. My work has focused on the IKK signalsome, a cytoplasmic complex that is an essential player in the activation of the NF-kappaB pathway by virtually all known stimuli. My project has employed microarray analysis of mouse embryonic fibroblasts individually knocked-out for three IKK signalsome subunits, IKKalpha, IKKbeta and NEMO, as well as pre-B cells missing the NEMO subunit. By using microarrays, I have been able to determine the global effect of signalsome subunit knockouts on the transcription of NF-kappaB dependent targets. I have discovered that all three subunits are required for the activation of NF-kappaB, a finding which initially created controversy, although recent findings by other groups has lent support to my conclusions. I have been able to discover that the signalsome is required for both stimulus dependent induction of NF-kappaB dependent transcription as well as basal NF-kappaB dependent transcription; My recent experiments have focused on determining whether IKKalpha expression alone is sufficient to restore normal stimulus dependent activation of NF-kappaB target genes. I have infected IKKalpha knockout fibroblasts via retroviruses, and have used microarray analysis to view the genome-wide effects of the re-introduction of IKKalpha. I have found that this expression alone is sufficient to impart normal activation of NF-kappaB in response to a pro-inflammatory cytokine. Furthermore, I have been able to rescue the IKKalpha knockout fibroblasts with a kinase dead mutant of IKKalpha. Under these conditions, I have noted only a small percentage of IKKalpha/NF-kappaB dependent target genes are properly stimulated. This suggest that IKKalpha can activate NF-kappaB on a limited basis without binding to ATP, which indicates that the kinase domain allows IKKalpha to fine-tune the transcription of NF-kappaB dependent target genes. My findings have implicated IKKalpha as a new target for anti-inflammatory drug research, as well as a possible target for anticancer therapies.