Inactivation of mitochondrial pyruvate carrier promotes NLRP3 inflammasome activation and gout development via metabolic reprogramming.

The nucleotide-binding and oligomerization domain, leucine-rich repeats, and pyrin domain-containing protein 3 (NLRP3) inflammasome plays a crucial role in innate immunity and is involved in the pathogenesis of autoinflammatory diseases. Glycolysis regulates NLRP3 inflammasome activation in macrophages. However, how lactic acid fermentation and pyruvate oxidation controlled by the mitochondrial pyruvate carrier (MPC) affect NLRP3 inflammasome activation and autoinflammatory disease remains elusive. We found that the inactivation of MPC with genetic depletion or pharmacological inhibitors, MSDC-0160 or pioglitazone, increased NLRP3 inflammasome activation and IL-1ß secretion in macrophages. Glycolytic reprogramming induced by MPC inhibition skewed mitochondrial ATP-associated oxygen consumption into cytosolic lactate production, which enhanced NLRP3 inflammasome activation in response to monosodium urate (MSU) crystals. As pioglitazone is an insulin sens MSDC-itizer used for diabetes, its MPC inhibitory effect in diabetic individuals was investigated. The results showed that MPC inhibition exacerbated MSU-induced peritonitis in diabetic mice and increased the risk of gout in patients with diabetes. Altogether, we found that glycolysis controlled by MPC regulated NLRP3 inflammasome activation and gout development. Accordingly, prescriptions for medications targeting MPC should consider the increased risk of NLRP3-related autoinflammatory diseases.

ATP8B1: A prognostic prostate cancer biomarker identified via genetic analysis.

BACKGROUND: Controlling the asymmetric distribution of phospholipids across biological membranes plays a pivotal role in the life cycle of cells; one of the most important contributors that maintain this lipid asymmetry are phospholipid-transporting adenosine triphosphatases (ATPases). Although sufficient information regarding their association with cancer exists, there is limited evidence linking the genetic variants of phospholipid-transporting ATPase family genes to prostate cancer in humans. METHODS: In this study, we investigated the association of 222 haplotype-tagging single-nucleotide polymorphisms (SNPs) in eight phospholipid-transporting ATPase genes with cancer-specific survival (CSS) and overall survival (OS) of 630 patients treated with androgen-deprivation therapy (ADT) for prostate cancer. RESULTS: After multivariate Cox regression analysis and multiple testing correction, we found that ATP8B1 rs7239484 was remarkably associated with CSS and OS after ADT. A pooled analysis of multiple independent gene-expression datasets demonstrated that ATP8B1 was under-expressed in tumor tissues and that a higher ATP8B1 expression was associated with a better patient prognosis. Moreover, we established highly invasive sublines using two human prostate cancer cell lines to mimic cancer progression traits in vitro. The expression of ATP8B1 was consistently downregulated in both highly invasive sublines. CONCLUSION: Our study indicates that rs7239484 is a prognostic factor for patients treated with ADT and that ATP8B1 can potentially attenuate prostate cancer progression.

ADAR1 promotes robust hypoxia signaling via distinct regulation of multiple HIF-1α-inhibiting factors.

Adenosine deaminase acting on RNA (ADAR)-catalyzed adenosine-to-inosine RNA editing is potentially dysregulated in neoplastic progression. However, how this transcriptome recoding process is functionally correlated with tumorigenesis remains largely elusive. Our analyses of RNA editome datasets identify hypoxia-related genes as A-to-I editing targets. In particular, two negative regulators of HIF-1A-the natural antisense transcript HIF1A-AS2 and the ubiquitin ligase scaffold LIMD1-are directly but differentially modulated by ADAR1. We show that HIF1A-AS2 antagonizes the expression of HIF-1A in the immediate-early phase of hypoxic challenge, likely through a convergent transcription competition in cis ADAR1 in turn suppresses transcriptional progression of the antisense gene. In contrast, ADAR1 affects LIMD1 expression post-transcriptionally, by interfering with the cytoplasmic translocation of LIMD1 mRNA and thus protein translation. This multi-tier regulation coordinated by ADAR1 promotes robust and timely accumulation of HIF-1α upon oxygen depletion and reinforces target gene induction and downstream angiogenesis. Our results pinpoint ADAR1-HIF-1α axis as a hitherto unrecognized key regulator in hypoxia.

Tumor-associated intronic editing of HNRPLL generates a novel splicing variant linked to cell proliferation.

Processing of the eukaryotic transcriptome is a dynamic regulatory mechanism that confers genetic diversity, and splicing and adenosine to inosine (A-to-I) RNA editing are well-characterized examples of such processing. Growing evidence reveals the cross-talk between the splicing and RNA editing, but there is a paucity of substantial evidence for its mechanistic details and contribution in a physiological context. Here, our findings demonstrate that tumor-associated differential RNA editing, in conjunction with splicing machinery, regulates the expression of variants of HNRPLL, a gene encoding splicing factor. We discovered an HNRPLL transcript variant containing an additional exon 12A (E12A), which is a substrate of ADAR1 and ADAR2. Adenosine deaminases acting on RNA (ADAR) direct deaminase-dependent expression of the E12A transcript, and ADAR-mediated regulation of E12A is largely splicing-based, and does not affect the stability or nucleocytoplasmic distribution of the transcript. Furthermore, ADAR-mediated modification of exon 12A generates an enhancer for the oncogenic splicing factor SRSF1 and consequently promotes the frequency of alternative splicing. Gene expression profiling by RNA-seq revealed that E12A acts distinctly from HNRPLL and regulates a set of growth-related genes, such as cyclin CCND1 and growth factor receptor TGFBR1 Accordingly, silencing E12A expression leads to impaired clonogenic ability and enhanced sensitivity to doxorubicin, thus highlighting the significance of this alternative isoform in tumor cell survival. In summary, we present the interplay of RNA editing and splicing as a regulatory mechanism of gene expression and also its physiological relevance. These findings extend our understanding of transcriptional dynamics and provide a mechanistic explanation to the link of RNA editors to tumorigenesis.

The PPARγ-SETD8 axis constitutes an epigenetic, p53-independent checkpoint on p21-mediated cellular senescence.

Cellular senescence is a permanent proliferative arrest triggered by genome instability or aberrant growth stresses, acting as a protective or even tumor-suppressive mechanism. While several key aspects of gene regulation have been known to program this cessation of cell growth, the involvement of the epigenetic regulation has just emerged but remains largely unresolved. Using a systems approach that is based on targeted gene profiling, we uncovered known and novel chromatin modifiers with putative link to the senescent state of the cells. Among these, we identified SETD8 as a new target as well as a key regulator of the cellular senescence signaling. Knockdown of SETD8 triggered senescence induction in proliferative culture, irrespectively of the p53 status of the cells; ectopic expression of this epigenetic writer alleviated the extent doxorubicin-induced cellular senescence. This repressive effect of SETD8 in senescence was mediated by directly maintaining the silencing mark H4K20me1 at the locus of the senescence switch gene p21. Further in support of this regulatory link, depletion of p21 reversed this SETD8-mediated cellular senescence. Additionally, we found that PPARγ acts upstream and regulates SETD8 expression in proliferating cells. Downregulation of PPARγ coincided with the senescence induction, while its activation inhibited the progression of this process. Viewed together, our findings delineated a new epigenetic pathway through which the PPARγ-SETD8 axis directly silences p21 expression and consequently impinges on its senescence-inducing function. This implies that SETD8 may be part of a cell proliferation checkpoint mechanism and has important implications in antitumor therapeutics.

ADAR1-mediated 3' UTR editing and expression control of antiapoptosis genes fine-tunes cellular apoptosis response.

Adenosine-to-inosine RNA editing constitutes a crucial component of the cellular transcriptome and critically underpins organism survival and development. While recent high-throughput approaches have provided comprehensive documentation of the RNA editome, its functional output remains mostly unresolved, particularly for events in the non-coding regions. Gene ontology analysis of the known RNA editing targets unveiled a preponderance of genes related to apoptosis regulation, among which proto-oncogenes XIAP and MDM2 encode two the most abundantly edited transcripts. To further decode this potential functional connection, here we showed that the main RNA editor ADAR1 directly targets this 3' UTR editing of XIAP and MDM2, and further exerts a negative regulation on the expression of their protein products. This post-transcriptional silencing role was mediated via the inverted Alu elements in the 3' UTR but independent of alteration in transcript stability or miRNA targeting. Rather, we discovered that ADAR1 competes transcript occupancy with the RNA shuttling factor STAU1 to facilitate nuclear retention of the XIAP and MDM2 mRNAs. As a consequence, ADAR1 may acquire functionality in part by conferring spatial distribution and translation efficiency of the target transcripts. Finally, abrogation of ADAR1 expression or catalytic activity elicited a XIAP-dependent suppression of apoptotic response, whereas ectopic expression reversed this protective effect on cell death. Together, our results extended the known functions of ADAR1 and RNA editing to the critical fine-tuning of the intracellular apoptotic signaling and also provided mechanistic explanation for ADAR1's roles in development and tumorigenesis.