A kinase shRNA screen links LATS2 and the pRB tumor suppressor

Abstract

pRB-mediated inhibition of cell proliferation is a complex process that depends on the action of many proteins. However, little is known about the specific pathways that cooperate with the Retinoblastoma protein (pRB) and the variables that influence pRB's ability to arrest tumor cells. Here we describe two shRNA screens that identify kinases that are important for pRB to suppress cell proliferation and pRB-mediated induction of senescence markers. The results reveal an unexpected effect of LATS2, a component of the Hippo pathway, on pRB-induced phenotypes. Partial knockdown of LATS2 strongly suppresses some pRB-induced senescence markers. Further analysis shows that LATS2 cooperates with pRB to promote the silencing of E2F target genes, and that reduced levels of LATS2 lead to defects in the assembly of DREAM (DP, RB [retinoblastoma], E2F, and MuvB) repressor complexes at E2F-regulated promoters. Kinase assays show that LATS2 can phosphorylate DYRK1A, and that it enhances the ability of DYRK1A to phosphorylate the DREAM subunit LIN52. Intriguingly, the LATS2 locus is physically linked with RB1 on 13q, and this region frequently displays loss of heterozygosity in human cancers. Our results reveal a functional connection between the pRB and Hippo tumor suppressor pathways, and suggest that low levels of LATS2 may undermine the ability of pRB to induce a permanent cell cycle arrest in tumor cells.

The Hippo pathway kinase Lats2 prevents DNA damage-induced apoptosis through inhibition of the tyrosine kinase c-Abl

“…Recent studies have shown links between the Hippo pathway and the pRB (retinoblastoma) pathway in the context of cellular senescence. [44][45][46][47] The latter studies show a role for Lats2 in mediating repression of E2F target genes by pRB in senescence. These studies induced senescence through oncogene expression or serum withdrawal.…”

Autophagy promotes radiation-induced senescence but inhibits bystander effects in human breast cancer cells

“…Thus, it is considered as an important tumor suppressive mechanism. 2,3 However, senescent cells are still metabolically active and ultimately develop senescence-associated secretory phenotypes, which participate in tumor suppression, tumor promotion, aging, and tissue repair. 4 SASPs composed of soluble proteins (cytokines, chemokines, and growth factors), secreted proteases and secreted insoluble proteins/extracellular matrix components may influence the fate of unirradiated neighboring cells, which is known as the bystander effect.…”

Identifying players in the functional network around pRB

“…The recently published results of shRNA screens give a glimpse of the types of signaling pathways that impact pRB function. 4 By quantifying the changes in cell number and the expression of senescence-associated β-galactosidase, we identified 39 kinases that were required for efficient pRB-inhibition of cell proliferation and 41 kinases that were necessary for pRB to establish a senescence-like state in SaOS2 cells, an osteosarcoma cell line that is mutant for RB1. These results demonstrate that there are many ways to enhance or suppress the ability of pRB to impose a permanent cell cycle arrest.…”

“…the kinases listed promote prB-dependent G 1 arrest or senescence. 4 shown in red are the subset of kinases that were also found in shrNa-based screens to support cell viability in cells that express mutant K-ras. [5][6][7] asterisks indicate that related (but non-identical) kinases were identified in the different types of screens.…”

“…This idea is supported by an intriguing functional relationship between pRB and LATS2 that was uncovered by these screens. 4 Depletion of LATS2, a component of the HIPPO pathway, prevented pRB from inducing markers of senescence, a form of permanent cell cycle arrest. Mechanistic analysis revealed that pRB and LATS2 converge in the regulation of the E2F transcription factor.…”