CAMK2D recruits RNF8-MAD2 complex to RNA splicing proteins for mitotic checkpoint activation in GBM

ABSTRACT: Glioblastoma (GBM) is a lethal brain tumor with remarkable intra-tumoral heterogeneity and therapeutic resistance which is linked to glioma stem cell (GSC). GSC contains unstable genome and elevated genomic instability could impair GSC self-renewal and tumorigenicity. In addition, GSC displays great dependency on BubR1, which is a key regulator of spindle assembly checkpoint (SAC) for proper chromosomal segregation. SAC prevents precocious anaphase progression in the presence of unattached kinetochores to ensure genomic integrity. To date, the regulation of SAC in GBM is still poorly understood and whether SAC could be exploited as a therapeutic target for GBM remains unknown. In our study, we revealed a previously unrecognized role of RNF8 in promoting SAC activation in GBM. GBM preferentially downregulates RNF8 in part by promoter methylation and low RNF8 expression correlates with inferior patient prognosis. Overexpression (OE) of wild-type RNF8 but not FHA and RING mutants significantly reduces GSC self-renewal, proliferation and tumorigenicity, and increases genomic instability. Mechanistically, the RING domain associates with the closed conformer of Mad2 (c-Mad2) and c-Mad2 exists as a complex with p31comet. CAMK2D binds transiently to the FHA domain via threonine 287 and this leads to RNF8 phosphorylation at serine 157. Furthermore, CAMK2D functions to recruit RNF8 to RNA splicing proteins to promote SAC activation independently of its enzymatic activity. To indirectly target RNF8-associated SAC in GBM, we exploited Connectivity Map (CMAP) and found that PLK1 inhibitor (PLK1i) could mimic RNF8-OE gene signature. Combination of PLK1i and proteostatic stressor leads to synergistic killing in GSCs. Collectively, our study unveiled an unconventional role of RNF8 in SAC regulation, dissected its molecular mechanisms and highlights the biological complexity of E3 ubiqtuitin ligases.

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Sek Tong Derrick, Ong 

PROVIDER: PXD038123 | JPOST Repository | Fri Jul 21 00:00:00 BST 2023

REPOSITORIES: jPOST