RESUMEN
Among sequenced organisms, the genome of Dictyostelium discoideum is unique in that it encodes for a massive amount of repeat-rich sequences in the coding region of genes. This results in the Dictyostelium proteome encoding for thousands of repeat-rich proteins, with nearly 24% of the Dictyostelium proteome encoding Q/N-rich regions that are predicted to be prion like in nature. To begin investigating the role of prion-like proteins in Dictyostelium, we decided to investigate ERF3, the Dictyostelium ortholog of the well-characterized yeast prion protein Sup35. ERF3 lacks the Q/N-rich region required for prion formation in yeast, raising the question of whether this protein aggregates and has prion-like properties in Dictyostelium. Here, we found that ERF3 formed aggregates in response to acute cellular stress. However, unlike bona fide prions, we were unable to detect transmission of aggregates to progeny. We further found that aggregation of this protein is driven by the ordered C-terminal domain independently of the disordered N-terminal domain. Finally, we also observed aggregation of ERF3 under conditions that induce multicellular development, suggesting that this phenomenon may play a role in Dictyostelium development. Together, these findings suggest a role for regulated protein aggregation in Dictyostelium cells under stress and during development.IMPORTANCEPrion-like proteins have both beneficial and deleterious effects on cellular health, and many organisms have evolved distinct mechanisms to regulate the behaviors of these proteins. The social amoeba Dictyostelium discoideum contains the highest proportion of proteins predicted to be prion like and has mechanisms to suppress their aggregation. However, the potential roles and regulation of these proteins remain largely unknown. Here, we demonstrate that aggregation of the Dictyostelium translation termination factor ERF3 is induced by both acute cellular stress and by multicellular development. These findings imply that protein aggregation may have a regulated and functional role in the Dictyostelium stress response and during multicellular development.
RESUMEN
Platinum resistance accounts for much of the high mortality and morbidity associated with ovarian cancer. Identification of targets with significant clinical translational potential remains an unmet challenge. Through a high-throughput synthetical lethal screening for clinically relevant targets using 290 kinase inhibitors, we identify calcium/calmodulin-dependent protein kinase II gamma (CAMK2G) as a critical vulnerability in cisplatin-resistant ovarian cancer cells. Pharmacologic inhibition of CAMK2G significantly sensitizes ovarian cancer cells to cisplatin treatment in vitro and in vivo. Mechanistically, CAMK2G directly senses ROS, both basal and cisplatin-induced, to control the phosphorylation of ITPKB at serine 174, which directly regulates ITPKB activity to modulate cisplatin-induced ROS stress. Thereby, CAMK2G facilitates the adaptive redox homeostasis upon cisplatin treatment and drives cisplatin resistance. Clinically, upregulation of CAMK2G activity and ITPKB pS174 correlates with cisplatin resistance in human ovarian cancers. This study reveals a key kinase network consisting of CAMK2G and ITPKB for ROS sense and scavenging in ovarian cancer cells to maintain redox homeostasis, offering a potential strategy for cisplatin resistance treatment.
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CisplatinoRESUMEN
PURPOSE: Although platinum compounds are the first-line treatment for ovarian cancer, the majority of patients relapse and develop resistance to treatment. However, the mechanism underlying resistance is unclear. The goal of our study is to decipher the mechanism by which a metabolic kinase, diacylglycerol kinase alpha (DGKA), confers platinum resistance in ovarian cancer. EXPERIMENTAL DESIGN: Metabolic kinase RNAi synthetic lethal screening was used to identify a cisplatin resistance driver in ovarian cancer. DGKA variants were used to demonstrate the need for DGKA activity in cisplatin resistance. Phospho-proteomic and genomic screens were performed to identify downstream effectors of DGKA. Therapeutic efficacy of targeting DGKA was confirmed and clinical relevance of DGKA signaling was validated using ovarian cancer patient-derived tumors that had different responses to platinum-based therapy. RESULTS: We found that platinum resistance was mediated by DGKA and its product, phosphatidic acid (PA), in ovarian cancer. Proteomic and genomic screens revealed that DGKA activates the transcription factor c-JUN and consequently enhances expression of a cell-cycle regulator, WEE1. Mechanistically, PA facilitates c-JUN N-terminal kinase recruitment to c-JUN and its nuclear localization, leading to c-JUN activation upon cisplatin exposure. Pharmacologic inhibition of DGKA sensitized ovarian cancer cells to cisplatin treatment and DGKA-c-JUN-WEE1 signaling positively correlated with platinum resistance in tumors derived from patients with ovarian cancer. CONCLUSIONS: Our study demonstrates how the DGKA-derived lipid messenger, PA, contributes to cisplatin resistance by intertwining with kinase and transcription networks, and provides preclinical evidence for targeting DGKA as a new strategy in ovarian cancer treatment to battle cisplatin resistance.
