XXXX Mutation Directed shRNA Screen Uncovers Novel @entity150 Growth Suppressors.

@entity1 @entity39195 @entity2534 @entity19 @entity5 @entity73074 @entity21121 @entity150 @entity2533 @entity52043 @entity73073
Next-generation sequencing (NGS) of @entity1 @entity150 has revealed many gene alterations compared with normal tissue, with most being predicted to be "loss of function." However, given the high number of alterations, evaluating the functional impact of each is impractical. Here, we develop and use a high-throughput, in vivo strategy to determine which alterations are loss of function in @entity5 growth suppressors. Genes reported as altered by NGS in @entity150 @entity1 were bioinformatically processed by MutationTaster and MutationAssessor, with 283 predicted as loss of function. An shRNA lentiviral library targeting these genes was transduced into @entity1 cells, a nontumorigenic @entity1 @entity150 cell line, followed by injection into @entity19 . @entity5 that arose were sequenced and the dominant shRNA constructs were found to target @entity52043 , @entity39195 , @entity73073 , @entity21121 , and @entity73074 genes. In vitro validation experiments revealed that shRNA molecules directed at @entity52043 showed the most profound increase in anchorage-independent growth of @entity1 cells. The clinical relevance of @entity52043 as a @entity5 growth suppressor is supported by the finding that its expression is lower in @entity150 compared with benign @entity1 urothelium in multiple independent datasets. Lower @entity52043 protein expression associated with higher @entity5 grade and decreased @entity1 survival. Finally, depletion of @entity52043 leads to increased @entity2534 with @entity2533 signaling, explaining in part how reduced @entity52043 promotes @entity5 growth. These findings suggest @entity52043 is a @entity150 growth suppressor that works via modulating @entity2533 signaling and is a potentially clinically useful biomarker. IMPLICATIONS: This study used gene mutation information from @entity1 -derived @entity150 specimens to inform the development of a screen used to identify novel @entity5 growth suppressors. This included identification of the protein @entity52043 as a potent @entity150 growth suppressor. Mol Cancer Res; 13(9); 1306-15.   2015 AACR.
