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Ras proteins are integral to the mediation of signaling cascades to downstream effectors, regulating a multitude of cellular processes. Mutations within Ras and its associated signaling pathways are implicated in various human pathologies, including inflammatory disorders and malignancies. The immune checkpoint proteins, programmed cell death protein 1 (PD-1) and its ligands PD-L1, along with Indoleamine 2,3-dioxygenase-1 (IDO1), are pivotal in facilitating tumor immune escape. While the influence of oncogenic Ras on PD-L1 expression is extensively documented, the regulatory role of KRas in IDO1 expression remains inadequately understood. In the current study, we demonstrate that IDO1 and PD-L1 expressions are differentially regulated in KRas-mutant cancers. Treatment with the KRasG12C-specific inhibitor, ARS-1620, significantly increased IDO1 expression, which inversely correlated with PD-L1 expression in the KRasG12C-mutant H358 cell line. Notably, IDO1 expression was slightly diminished in KRas-mutant patients with lung and pancreatic ductal adenocarcinomas. Experimental data revealed that IFN-γ induces IDO1 expression; however, this induction is attenuated in the presence of constitutively active KRas. These findings suggest that KRas signaling negatively regulates IDO1 expression while enhancing PD-L1 expression. Moreover, the induction of IDO1 expression following KRas inhibition appears to operate independently of the MAPK pathway. Our results propose that concurrent targeting of KRas and IDO1 could potentiate therapeutic efficacy in KRas-mutant cancers, overcoming resistance to immune checkpoint blockade.

Abstract While EZH2 enzymatic activity is well-known, emerging evidence suggests that EZH2 can exert functions in a methyltransferase-independent manner. In this study, we have uncovered a novel mechanism by which EZH2 positively regulates the expression of SKP2, a critical protein involved in cell cycle progression. We demonstrate that depletion of EZH2 significantly reduces SKP2 protein levels in several cell types, while treatment with EPZ-6438, an EZH2 enzymatic inhibitor, has no effect on SKP2 protein levels. Consistently, EZH2 depletion leads to cell cycle arrest, accompanied by elevated expression of CIP/KIP family proteins, including p21, p27, and p57, whereas EPZ-6438 treatment does not modulate their levels. We also provide evidence that EZH2 knockdown, but not enzymatic inhibition, suppresses SKP2 mRNA expression, underscoring the transcriptional regulation of SKP2 by EZH2 in a methyltransferase-independent manner. Supporting this, analysis of the Cancer Genome Atlas database reveals a close association between EZH2 and SKP2 expression in human malignancies. Moreover, EZH2 depletion but not enzymatic inhibition positively regulates the expression of major epithelial-mesenchymal transition (EMT) regulators, such as ZEB1 and SNAIL1, in transformed cells. Our findings shed light on a novel mechanism by which EZH2 exerts regulatory effects on cell proliferation and differentiation through its methyltransferase-independent function, specifically by modulating SKP2 expression.

Cyanobacterial blooms and the associated release of cyanotoxins pose problems for many conventional water treatment plants due to their limited removal by typical unit operations. In this study, a conventional water treatment process consisting of coagulation, flocculation, sedimentation, filtration, and sludge dewatering was assessed in lab-scale experiments to measure the removal of microcystin-LR and Microcystis aeruginosa cells using liquid chromatography with mass spectrometer (LC-MS) and a hemacytometer, respectively. The overall goal was to determine the effect of recycling cyanotoxin-laden dewatered sludge supernatant on treated water quality. The lab-scale experimental system was able to maintain the effluent water quality below relevant the United States Environmental Protection Agency (US EPA) and World Health Organisation (WHO) standards for every parameter analyzed at influent concentrations of M. aeruginosa above 106 cells/mL. However, substantial increases of 0.171 NTU (Nephelometric Turbidity Unit), 7 × 104 cells/L, and 0.26 µg/L in turbidity, cyanobacteria cell counts, and microcystin-LR concentration were observed at the time of dewatered supernatant injection. Microcystin-LR concentrations of 1.55 µg/L and 0.25 µg/L were still observed in the dewatering process over 24 and 48 h, respectively, after the initial addition of M.aeruginosa cells, suggesting the possibility that a single cyanobacterial bloom may affect the filtered water quality long after the bloom has dissipated when sludge supernatant recycling is practiced.