Characterization of Mitoribosomal Small Subunit unit genes related immune and pharmacogenomic landscapes in renal cell carcinoma.

Mitoribosomes are essential for the production of biological energy. The Human Mitoribosomal Small Subunit unit (MRPS) family, responsible for encoding mitochondrial ribosomal small subunits, is actively engaged in protein synthesis within the mitochondria. Intriguingly, MRPS family genes appear to play a role in cancer. A multistep process was employed to establish a risk model associated with MRPS genes, aiming to delineate the immune and pharmacogenomic landscapes in clear cell renal cell carcinoma (ccRCC). MRPScores were computed for individual patients to assess their responsiveness to various treatment modalities and their susceptibility to different therapeutic targets and drugs. While MRPS family genes have been implicated in various cancers as oncogenes, our findings reveal a contrasting tumor suppressor role for MRPS genes in ccRCC. Utilizing an MRPS-related risk model, we observed its excellent prognostic capability in predicting survival outcomes for ccRCC patients. Remarkably, the subgroup with high MRPS-related scores (MRPScore) displayed poorer prognosis but exhibited a more robust response to immunotherapy. Through in silico screening of 2183 drug targets and 1646 compounds, we identified two targets (RRM2 and OPRD1) and eight agents (AZ960, carmustine, lasalocid, SGI-1776, AZD8055_1059, BPD.00008900_1998, MK.8776_2046, and XAV939_1268) with potential therapeutic implications for high-MRPScore patients. Our study represents the pioneering effort in proposing that molecular classification, diagnosis, and treatment strategies can be formulated based on MRPScores. Indeed, a high MRPScore profile appears to elevate the risk of tumor progression and mortality, potentially through its influence on immune regulation. This suggests that the MRPS-related risk model holds promise as a prognostic predictor and may offer novel insights into personalized therapeutic strategies.

Effect of Rhodotorula mucilaginosa inoculation on the aroma development of a fermented vegetables simulated system.

Fermented vegetables are known for their unique flavors and aromas, which are influenced by the complex microbial processes that occur during fermentation. Rhodotorula mucilaginosa is a red yeast strain that is frequently isolated from fermented vegetables. However, the specific mechanisms underlying their effects on aroma production remain unclear. In this study, a simulated system of vegetables fermented using vegetable juices was used to investigate the effects of R. mucilaginosa inoculation on aroma development. The results demonstrated that this red yeast strain could utilize the nutrients present in the vegetable juices to support its growth and reproduction. Moreover, the inoculation of fermented vegetable juices with this yeast strain led to an increase in the levels of umami amino acids and sweet amino acids. Furthermore, this yeast strain was found able to significantly reduce the content of sulfur-containing compounds, which may decrease the unpleasant odor of fermented vegetables. Additionally, the yeast strain was capable of producing high concentrations of aromatic compounds such as phenylethyl alcohol, methyl 2-methylbutyrate, methyl butyrate, and nonanoic acid in a minimum medium. However, only phenylethyl alcohol has been identified as a core aromatic compound in fermented vegetable juice. The three fermented vegetable juices exhibited significantly different flavor profiles according to comparative analysis. Therefore, the core flavor compounds found in fermented vegetables are primarily derived from the release and modification of endogenous flavors naturally present in the vegetables, facilitated by the yeast during fermentation.

Increased chromatin accessibility drives transition to androgen receptor splice variant dependence in castration-resistant prostate cancer.

Androgen receptor (AR) splice variants, of which ARv7 is the most common, are increased in prostate cancer (PC) that develops resistance to androgen signaling inhibitor drugs, but the extent to which these variants drive AR activity, and whether they have novel functions or dependencies, remain to be determined. We generated a subline of VCaP PC cells (VCaP16) that is resistant to the AR inhibitor enzalutamide (ENZ) and found that AR activity was independent of the full-length AR (ARfl), despite its continued high-level expression, and was instead driven by ARv7. The ARv7 cistrome and transcriptome in VCaP16 cells mirrored that of the ARfl in VCaP cells, although ARv7 chromatin binding was weaker, and strong ARv7 binding sites correlated with higher affinity ARfl binding sites across multiple models and clinical samples. Notably, although ARv7 expression in VCaP cells increased rapidly in response to ENZ, there was a long lag before it gained chromatin binding and transcriptional activity. This lag was associated with an increase in chromatin accessibility, with the AR and nuclear factor I (NFI) motifs being most enriched at these more accessible sites. Moreover, the transcriptional effects of combined NFIB and NFIX knockdown versus ARv7 knockdown were highly correlated. These findings indicate that ARv7 can drive the AR program, but that its activity is dependent on adaptations that increase chromatin accessibility to enhance its intrinsically weak chromatin binding.

Dissecting transcription of the 8q24-MYC locus in prostate cancer recognizes the equilibration between androgen receptor direct and indirect dual-functions.

BACKGROUND: Androgen receptor (AR) activation and repression dual-functionality only became known recently and still remains intriguing in prostate cancer (PCa). MYC is a prominent oncogene that functionally entangles with AR signaling in PCa. Further exploration of AR regulatory mechanisms on MYC gene transcription bears clinical and translation significance. METHODS: Bioinformatics analysis of PCa cell line and clinical RNA-Seq and ChIP-Seq (chromatin immunoprecipitation-sequencing) datasets to anchor interactions of AR and MYC transcriptional networks. ChIP-qPCR and 3C (chromosome conformation capture) analyses to probe MYC distal regulation by AR binding sites (ABSs). CRISPR/Cas9-mediated genome-editing to specify functions of ABS within the 8q24-MYC locus on androgen-mediated MYC transcription. Global FoxA1 and HoxB13 distribution profiling to advance AR transcriptional mechanisms. RESULTS: Here we recognize AR bi-directional transcription mechanisms by exploiting the prominent 8q24-MYC locus conferring androgen hyper-sensitivity. At ~ 25 Kb downstream of the MYC gene, we identified an undefined ABS, P10. By chromatin analyses, we validated androgen-dependent spatial interaction between P10 and MYC-Promoter (MYC-Pro) and temporal epigenetic repression of these MYC-proximal elements. We next designed a CRISPR/Cas9-mediated double genomic knock-out (KO) strategy to show that P10-KO slightly lessened androgen-elicited MYC transrepression in LNCaP-AR cells. In similar genomic editing assays, androgen-mediated MYC repression became slightly deepened upon KO of P11, an ABS in the PVT1 gene locus highly enriched in AR-binding motifs and peaks. We also investigated multiple ABSs in the established PCAT1 super-enhancer that distally interacts with MYC-Pro for transactivation, with each KO pool consistently shown to relieve androgen-elicited MYC repression. In the end, we systemically assessed androgen effects in the 8q24-MYC locus and along PCa genome to generalize H3K27ac and BRD4 re-distribution from pioneer factors (FoxA1 and HoxB13) to AR sites. CONCLUSION: Together, we reconciled these observations by unifying AR dual-functions that are mechanistically coupled to and equilibrated by co-factor redistribution.

Androgen receptor and MYC transcriptomes are equilibrated in multilayer regulatory circuitries in prostate cancer.

BACKGROUND: The discovery of androgen receptor (AR) having transrepression effects completes the circle of its functionalities as a typical transcription factor, which intrinsically bears dual functions of activation and repression linked to co-factor competition and redistribution. Indeed, AR dual functions are exemplified by locus-wide regulation of the oncogenic 8q24-MYC region. METHODS: RT-qPCR assay and public RNA-profiling datasets were used to assess MYC transcription in androgen-sensitive cell lines. Public ChIP-seq and RNA-Seq datasets were computed to evaluate AR-MYC direct and indirect signatures. Gene sets in typical MYC and AR pathways were monitored to validate their cross-talks. Bio-informatics and chromosome conformation capture (3C) assay were performed in the AR gene locus to examine androgen-elicited distal regulation. Finally, co-factor re-distribution were globally tracked between AR and MYC binding sites. RESULTS: In this report, we found MYC responded negatively to androgen with hypersensitivity, rivaling AR natural functions as an innate androgen effector. Furthermore, both direct and indirect AR and MYC transcriptional programs were actively in equilibration. With established androgen-mediated versus MYC-mediated gene subsets, we validated AR and MYC pathways were both bidirectional and extensively entangled. In addition, we determined that the AR gene locus resembled the MYC gene region and both loci were androgen-repressed via epigenetics and chromatin architectural alterations. Significantly, transcriptional factor profiling along the prostate cancer (PCa) genome exposed that PCa transcriptomes were dynamically equilibrated between AR-binding site and MYC-binding site. CONCLUSION: Together, our findings stratified AR-MYC interactions that are extensively wired and intricately organized to compensate for essential PCa transcriptional programs and neutralize excessive signaling.

Deficiency of mindin reduces renal injury after ischemia reperfusion.

BACKGROUND: Acute renal injury (AKI) secondary to ischemia reperfusion (IR) injury continues to be a significant perioperative problem and there is no effective treatment. Mindin belongs to the mindin/F-spondin family and involves in inflammation, proliferation, and cell apoptosis. Previous studies have explored the biological functions of mindin in liver and brain ischemic injury, but its role in AKI is unknown. METHOD: To investigate whether mindin has a pathogenic role, mindin knockout (KO) and wild-type (WT) mice were used to establish renal IR model. After 30 min of ischemia and 24 h of reperfusion, renal histology, serum creatinine, and inflammatory response were examined to assess kidney injury. In vitro, proinflammatory factors and inflammatory signaling pathways were measured in mindin overexpression or knockdown and vector cells after hypoxia/reoxygenation (HR). RESULTS: Following IR, the kidney mindin level was increased in WT mice and deletion of mindin provided significant protection for mice against IR-induced renal injury as manifested by attenuated the elevation of serum creatinine and blood urea nitrogen along with less severity for histological alterations. Mindin deficiency significantly suppressed inflammatory cell infiltration, TNF-α and MCP-1 production following renal IR injury. Mechanistic studies revealed that mindin deficiency inhibits TLR4/JNK/NF-κB signaling activation. In vitro, the expression levels of TNF-α and MCP-1 were increased in mindin overexpression cells compared with vector cells following HR. Moreover, TLR4/JNK/NF-κB signaling activation was elevated in the mindin overexpression cells in response to HR stimulation while mindin knockdown inhibited the activation of TLR4/JNK/ NF-κB signaling after HR in vitro. Further study showed that mindin protein interacted directly with TLR4 protein. And more, mindin protein was confirmed to be expressed massively in renal tubule tissues of human hydronephrosis patients. CONCLUSION: These data demonstrate that mindin is a critical modulator of renal IR injury through regulating inflammatory responses. TLR4/JNK/NF-κB signaling most likely mediates the biological function of mindin in this model of renal ischemia.

The Alternatively Spliced Isoforms of Key Molecules in the cGAS-STING Signaling Pathway.

Alternative splicing of pre-mRNA increases transcriptome and proteome diversity by generating distinct isoforms that encode functionally diverse proteins, thus affecting many biological processes, including innate immunity. cGAS-STING signaling pathway, whose key molecules also undergo alternative splicing, plays a crucial role in regulating innate immunity. Protein isoforms of key components in the cGAS-STING-TBK1-IRF3 axis have been detected in a variety of species. A chain of evidence showed that these protein isoforms exhibit distinct functions compared to their normal counterparts. The mentioned isoforms act as positive or negative modulators in interferon response via distinct mechanisms. Particularly, we highlight that alternative splicing serves a vital function for the host to avoid the overactivation of the cGAS-STING signaling pathway and that viruses can utilize alternative splicing to resist antiviral response by the host. These findings could provide insights for potential alternative splicing-targeting therapeutic applications.

Androgen receptor splice variant 7 functions independently of the full length receptor in prostate cancer cells.

One mechanism for reactivation of androgen receptor (AR) activity after androgen deprivation therapy in castration-resistant prostate cancer (CRPC) is expression of splice variants such as ARv7 that delete the ligand binding domain and have constitutive activity. Exogenous overexpressed ARv7 can function as a homodimer or heterodimer with full length AR (ARfl), which is highly expressed with ARv7 in CRPC. However, the extent to which endogenous ARv7 function is dependent on heterodimerization with ARfl remains to be determined. We used double-crosslinking to stabilize AR complexes on chromatin in a CRPC cell line expressing endogenous ARfl and ARv7 (LN95 cells), and established that only trace levels of ARfl were associated with ARv7 on chromatin. Consistent with this result, depletion of ARfl with an AR degrader targeting the AR ligand binding domain did not decrease ARv7 binding to chromatin or its association with HOXB13, but did decrease overall AR transcriptional activity. Comparable results were obtained in CWR22RV1 cells, another CRPC cell line expressing ARfl and ARv7. These results indicate that ARv7 function in CRPC is not dependent on ARfl, and that both contribute independently to overall AR activity.

Phosphorylation of the androgen receptor at Ser81 is co-sustained by CDK1 and CDK9 and leads to AR-mediated transactivation in prostate cancer.

Androgen receptor (AR) is the principal molecule in prostate cancer (PCa) etiology and therapy. AR re-activation still remains a major challenge during treatment of castration-resistant prostate cancer (CRPC) tumors that relapse after castration therapies. Recent reports have indicated the enrichment of Ser81-phosphorylated AR (pS81) in the nucleus of CRPC cells, and CDK1 and CDK9 as the kinases phosphorylating AR at S81. In the current study we showed that pS81 is preferentially localized in the nucleus in both rapid biopsy metastatic CRPC samples and PCa xenografts, and nuclear pS81 localization is correlated with AR transactivation in tumor xenografts. Chromatin immunoprecipitation (ChIP) analysis demonstrated an alignment of S81 phosphorylation and AR-mediated transactivation with the chromatin locus openness. Moreover, pS81-specific ChIP-Seq showed a disproportional occupancy of pS81 on AR-activated promoters, while 3C-ChIP assays further indicated an enrichment of pS81 at the PSA enhancer-promoter loop, a known AR activating hub. In the latter, CDK9 was shown to modulate the transactivation of the AR and RNA Pol II. Indeed, ChIP and re-ChIP assays also confirmed that AR-dependent activation of the PSA enhancer and promoter mediated by pS81 was coupled with activation of Pol II and the pTEFb complex. Mechanistically, we determined that CDK1 and CDK9 sustained the pS81 AR modification in the soluble and chromatin-bound fractions of PCa cells, respectively. Finally, we demonstrated that CDK1 activity was maintained throughout the cell cycle, and that CDK1 inhibitors restored androgen sensitivity in CRPC tumor cells. Based on these findings, CDK1 and CDK9 could be targeted as pS81 kinases in patients with CRPC, either alone or in conjunction with direct AR antagonists.

NSUN5 Facilitates Viral RNA Recognition by RIG-I Receptor.

The RIG-I receptor induces the innate antiviral responses upon sensing RNA viruses. The mechanisms through which RIG-I optimizes the strength of the downstream signaling remain incompletely understood. In this study, we identified that NSUN5 could potentiate the RIG-I innate signaling pathway. Deficiency of NSUN5 enhanced RNA virus proliferation and inhibited the induction of the downstream antiviral genes. Consistently, NSUN5-deficient mice were more susceptible to RNA virus infection than their wild-type littermates. Mechanistically, NSUN5 bound directly to both viral RNA and RIG-I, synergizing the recognition of dsRNA by RIG-I. Collectively, to our knowledge, this study characterized NSUN5 as a novel RIG-I coreceptor, playing a vital role in restricting RNA virus infection.

The predictive value for excellent response to initial therapy in differentiated thyroid cancer: preablation-stimulated thyroglobulin better than the TNM stage.

PURPOSE: This study aimed to identify the predictive value of the low preablation-stimulated thyroglobulin (ps-Tg, <2 ng/ml) for excellent response to radioiodine remnant ablation in differentiated thyroid carcinoma. PATIENTS AND METHODS: A total of 398 consecutive patients who underwent total thyroidectomy and radioactive iodine remnant ablation therapy were reviewed retrospectively. Each patient was risk-stratified using the American Joint Cancer Committee and risk staging systems and using response to the initial therapy reclassification system. ps-Tg was defined as less than 2 ng/ml with negative thyroglobulin antibody under thyroid-stimulating hormone stimulation. A multivariate analysis was carried out for ps-Tg, TNM stage, and other potential clinical and pathologic factors. RESULTS: We followed the patients for a median of 32.7 months. Overall, an excellent rate of response was achieved in 367 (92.2%) of the 398 patients. The only variable found to be associated with excellent response was ps-Tg (odds ratio=2.530, P=0.009) by multivariate analysis. The subgroups with 0

LEF1 targeting EMT in prostate cancer invasion is mediated by miR-181a.

Lymphoid enhancer-binding factor-1 (LEF1) is a key transcription factor mediating Wnt signaling pathway. Our previous studies indicate that LEF1 is highly expressed in androgen-independent prostate cancer (PCa) and enhances invasion ability in androgen-independent PCa cells. However, the molecular mechanism of LEF1 effect on invasion remains largely unknown. Using microRNA profiling analysis comparing androgen-independent LNCaP-AI PCa cells with high levels of endogenous LEF1 to LNCaP-AI cells with LEF1 knockdown by LEF1shRNA, we found miR-181a to be increased 12.3-fold in LNCaP-AI cells. We confirmed a positive correlation between LEF1 and miR-181a expression across multiple PCa cell lines. Additionally, we showed that in PCa cells, overexpression of LEF1 increased miR-181a expression and subsequently induced EMT associated migration and invasion, whereas LEF1 knockdown decreased miR-181a expression and subsequently resulted in inhibition of EMT, migration and invasion. Mechanistically, we demonstrated by chromatin immunoprecipitation assays that LEF1 could enhance miR-181a expression via its binding to the promoter regions of hsa-miR-181a. Overall, this study identified a novel LEF1-miR-181a-EMT axis in regulation of PCa migration and invasion.

LEF1 Targeting EMT in Prostate Cancer Invasion Is Regulated by miR-34a.

UNLABELLED: The microRNA-34a (miR-34a), a tumor-suppressive microRNA (miRNA), is implicated in epithelial-mesenchymal transition (EMT) and cancer stem cells. Lymphoid enhancer-binding factor-1 (LEF1) is a key transcription factor in the Wnt signaling pathway, and has been suggested to be involved in regulation of cell proliferation and invasion. Here, the molecular mechanism of miR-34a and LEF1 in cooperatively regulating prostate cancer cell invasion is described. Molecular profiling analysis of miRNA levels in prostate cancer cells revealed a negative correlation between miR-34a and LEF1 expression, and the downregulation of LEF1 by miR-34a was confirmed by luciferase assays. Furthermore, miR-34a specifically repressed LEF1 expression through direct binding to its 3'-untranslated regions (3'-UTR). miR-34a modulated the levels of LEF1 to regulate EMT in prostate cancer cells. Functionally, miR-34a negatively correlated with the migration and invasion of prostate cancer cells through LEF1. An analysis of miR-34a expression levels in matched human tumor and benign tissues demonstrated consistent and statistically significant downregulation of miR-34a in primary prostate cancer specimens. These data strongly suggest that miR-34a/LEF1 regulation of EMT plays an important role in prostate cancer migration and invasion. IMPLICATIONS: The miR-34a-LEF1 axis represents a potential molecular target for novel therapeutic strategies in prostate cancer.

Epithelial and stromal expression of miRNAs during prostate cancer progression.

Global microRNA (miRNA) profile may predict prostate cancer (PCa) behaviors. In this study, we examined global miRNA expression by miRNA profiling as well as specific miRNA expression levels in PCa epithelium and stroma by in situ hybridization (ISH) and correlated with various clinicopathological features. We first performed comprehensive miRNA profiling on 27 macrodissected cases of PCa by miRNA microarray. A total of 299 miRNAs were significantly dysregulated in high grade and advanced stage PCa. We demonstrated that PCa can be readily classified into high grade/stage and low-grade/stage groups by its global miRNA expression profile. Next, we examined the expression of several selected dysregulated miRNAs, including let-7c, miR-21, miR-27a, miR-30c, and miR-219, in PCa by ISH. The levels of miRNA expression in epithelial and stromal cells were scored semiquantitatively and compared with clinicopathological features, including age, race, Gleason score, stage, PSA recurrence, metastasis, hormone resistance and survival. We found that the expression of miR-30c and miR-219 were significantly down-regulated in PCa. miR-21 and miR-30c were significantly down-regulated in PCa in African Americans compared to Caucasian Americans. In addition, down-regulation of let-7c, miR-21, miR-30c, and miR-219 are associated with metastatic disease. Furthermore, down-regulation of miR-30c and let-7c are significantly associated with androgen-dependent PCa. In PCa stromal cells, let-7c downregulation is significantly associated with extraprostatic extension. Our data suggest that selected miRNAs may serve as potential biomarkers to predict cancer progression.

MiR-138 induces renal carcinoma cell senescence by targeting EZH2 and is downregulated in human clear cell renal cell carcinoma.

MiR-138 has been shown to be downregulated in various cancers, including head and neck squamous cell carcinoma (HNSCC) and clear cell renal carcinoma (ccRCC). In the present study, we aimed to reveal the mechanism of miR-138 induction of senescence in renal carcinoma cells and identify its specific target genes. We used qRT-PCR to analyze miR-138 expression levels in renal carcinoma cell lines and ccRCC samples. The activity of ß-galactosidase was measured for functional analysis after miR-138 mimic transfection. To identify the targets of miR-138, we used three types of target prediction software to determine three candidate target genes. Furthermore, a 3'UTR luciferase assay was performed. Western blotting was used to detect the protein expression levels of candidate target genes. Additionally, knockdown of EZH2 by its siRNA was performed. The expression of miR-138 was downregulated in RCC cells lines and in tumor samples compared with their controls. Transfection of miR-138 mimic induced SN-12 cell senescence, decreased the protein expression of EZH2, and increased the protein expression of P16. Furthermore, miR-138 decreased the 3'UTR luciferase activity of EZH2. The knockdown of EZH2 by siRNA induced SN-12 cell senescence, decreased the protein expression level of EZH2, and increased the protein expression of P16. MiR-138 is a tumor-suppressor miRNA in ccRCC that induces SN-12 cell senescence by downregulating EZH2 expression and upregulating P16 expression.