No abstract

The core LATS kinases of the Hippo tumor suppressor pathway phosphorylate and inhibit the downstream transcriptional coactivators YAP and TAZ, which are implicated in various cancers. Recent studies have identified various E3 ubiquitin ligases that negatively regulate the Hippo pathway via ubiquitination, yet few deubiquitinating enzymes (DUB) have been implicated. In this study, we report the DUB USP9X is an important regulator of the core kinases of this pathway. USP9X interacted strongly with LATS kinase and to a lesser extent with WW45, KIBRA, and Angiomotin, and LATS co-migrated exclusively with USP9X during gel filtration chromatography analysis. Knockdown of USP9X significantly downregulated and destabilized LATS and resulted in enhanced nuclear translocation of YAP and TAZ, accompanied with activation of their target genes. In the absence of USP9X, cells exhibited an epithelial-to-mesenchymal transition phenotype, acquired anchorage-independent growth in soft agar, and led to enlarged, disorganized, three-dimensional acini. YAP/TAZ target gene activation in response to USP9X knockdown was suppressed by knockdown of YAP, TAZ, and TEAD2. Deletion of USP9X in mouse embryonic fibroblasts resulted in significant downregulation of LATS. Furthermore, USP9X protein expression correlated positively with LATS but negatively with YAP/TAZ in pancreatic cancer tissues as well as pancreatic and breast cancer cell lines. Overall, these results strongly indicate that USP9X potentiates LATS kinase to suppress tumor growth. Cancer Res; 77(18); 4921-33. Ó2017 AACR.

Zebrafish is a popular system for studying vertebrate development and disease that shows high genetic conservation with humans. Molecular level studies at different stages of development are essential for understanding the processes deployed during ontogeny. Here, we performed comparative analysis of the whole proteome and transcriptome of the early stage (24 h post-fertilization) zebrafish embryo. We identified 8363 proteins with their approximate cellular abundances (the largest number of zebrafish embryo proteins quantified thus far), through a combination of thorough deyolking and extensive fractionation procedures, before resolving the peptides by mass spectrometry. We performed deep sequencing of the transcripts and found that the expressed proteome and transcriptome displayed a moderate correlation for the majority of cellular processes. Integrative functional mapping of the quantified genes demonstrated that embryonic developmental systems differentially exploit transcriptional and post-transcriptional regulatory mechanisms to modulate protein abundance. Using network mapping of the low-abundance proteins, we identified various signal transduction pathways important in embryonic development and also revealed genes that may be regulated at the post-transcriptional level. Our data set represents a deep coverage of the functional proteome and transcriptome of the developing zebrafish, and our findings unveil molecular regulatory mechanisms that underlie embryonic development.