Functional Genomic Approaches To Identify Modulators Of NF-kappaB Signaling
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Chronic inflammation is associated with increased cancer risk. Acute inflammation is also a very dangerous and serious condition. NF-kappaB is one of the more important regulators of inflammation and therefore plays a major role in both of these conditions. We used a functional genomic approach to identify activators or suppressors of NF-kappaB (NASPs and shRNAs respectively). Two strategies were used to accomplish this goal. The first was a newly established technology called BASP technology that could identify activators of NF-kappaBand the second was a standard shRNA loss of function screens to identify NF-kappaB suppressors. NF-kappaB activating elements were identified and employed to address the role of chronic inflammation in cellular transformation. Our investigations revealed that NF-kappaB activating selectable peptides (NASPs) cooperate with oncogenic-Ras to promote transformation through weakening of p53 activity. Our second approach with the shRNA screens revealed inhibitors of NF-kappaB most notably, shCCR4. Mechanistic studies revealed that shCCR4 transcriptionally suppresses NF-kappaB activity by causing a reduction in the total p65 protein levels. In view of constitutive activation of NF-kappaB in the majority of tumors we proposed that shCCR4 could be used to suppress NF-kappaB for cancer therapy, and we found that shCCR4 lead to growth suppression in a variety of NF-kappaB dependent tumor cell lines. Overall, this system is a useful, unbiased tool to screen for functional molecules in biological systems that may have yet to be revealed. It also provides the necessary groundwork for the identification of novel drug targets using functional assays, even if the operative mechanisms are not well known.