NAT10-mediated ac4C-modified ANKZF1 promotes tumor progression and lymphangiogenesis in clear-cell renal cell carcinoma by attenuating YWHAE-driven cytoplasmic retention of YAP1.

BACKGROUND: Lymphatic metastasis is one of the most common metastatic routes and indicates a poor prognosis in clear-cell renal cell carcinoma (ccRCC). N-acetyltransferase 10 (NAT10) is known to catalyze N4-acetylcytidine (ac4C) modification of mRNA and participate in many cellular processes. However, its role in the lymphangiogenic process of ccRCC has not been reported. This study aimed to elucidate the role of NAT10 in ccRCC lymphangiogenesis, providing valuable insights into potential therapeutic targets for intervention. METHODS: ac4C modification and NAT10 expression levels in ccRCC were assessed using public databases and clinical samples. Functional investigations involved manipulating NAT10 expression in cellular and mouse models to study its role in ccRCC. Mechanistic insights were gained through a combination of RNA sequencing, mass spectrometry, co-immunoprecipitation, RNA immunoprecipitation, immunofluorescence, and site-specific mutation analyses. RESULTS: We found that ac4C modification and NAT10 expression levels increased in ccRCC. NAT10 promoted tumor progression and lymphangiogenesis of ccRCC by enhancing the nuclear import of Yes1-associated transcriptional regulator (YAP1). Subsequently, we identified ankyrin repeat and zinc finger peptidyl tRNA hydrolase 1 (ANKZF1) as the functional target of NAT10, and its upregulation in ccRCC was caused by NAT10-mediated ac4C modification. Mechanistic analyses demonstrated that ANKZF1 interacted with tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein epsilon (YWHAE) to competitively inhibit cytoplasmic retention of YAP1, leading to transcriptional activation of pro-lymphangiogenic factors. CONCLUSIONS: These results suggested a pro-cancer role of NAT10-mediated acetylation in ccRCC and identified the NAT10/ANKZF1/YAP1 axis as an under-reported pathway involving tumor progression and lymphangiogenesis in ccRCC.

Erratum: FAM72A promotes glioma progression by regulating mitophagy through the Pink1/Parkin signaling pathway: Erratum.

[This corrects the article DOI: 10.7150/jca.82949.].

HIF2α Promotes Cancer Metastasis through TCF7L2-Dependent Fatty Acid Synthesis in ccRCC.

Recent studies have highlighted the notable involvement of the crosstalk between hypoxia-inducible factor 2 alpha (HIF2α) and Wnt signaling components in tumorigenesis. However, the cellular function and precise regulatory mechanisms of HIF2α and Wnt signaling interactions in clear cell renal cell carcinoma (ccRCC) remain elusive. To analyze the correlation between HIF2α and Wnt signaling, we utilized the Cancer Genome Atlas - Kidney Renal Clear Cell Carcinoma (TCGA-KIRC) public database, HIF2α RNA sequencing data, and conducted luciferase reporter assays. A Wnt-related gene set was employed to identify key regulators of Wnt signaling controlled by HIF2α in ccRCC. Furthermore, we assessed the biological effects of TCF7L2 on ccRCC metastasis and lipid metabolism in both in vivo and in vitro settings. Our outcomes confirm TCF7L2 as a key gene involved in HIF2α-mediated regulation of the canonical Wnt pathway. Functional studies demonstrate that TCF7L2 promotes metastasis in ccRCC. Mechanistic investigations reveal that HIF2α stabilizes TCF7L2 mRNA in a method based on m6A by transcriptionally regulating METTL3. Up-regulation of TCF7L2 enhances cellular fatty acid oxidation, which promotes histone acetylation. This facilitates the transcription of genes connected to epithelial-mesenchymal transition and ultimately enhances metastasis of ccRCC. These outcomes offer a novel understanding into the involvement of lipid metabolism in the signaling pathway regulation, offering valuable implications for targeted treatment in ccRCC.

TBC1D5 reverses the capability of HIF-2α in tumor progression and lipid metabolism in clear cell renal cell carcinoma by regulating the autophagy.

BACKGROUND: Clear cell renal cell carcinoma (ccRCC) is known for abnormal lipid metabolism and widespread activation of HIF-2α. Recently, the importance of autophagy in ccRCC has been focused, and it has potential connections with HIF-2α and lipid metabolism. However, the specific regulatory mechanism between HIF-2α, autophagy, and lipid metabolism in ccRCC is still unclear. METHODS: In this study, Bioinformatics Analysis and Sequencing of the whole transcriptome were used to screen our target. The expression of TBC1D5 in renal clear cell carcinoma was confirmed by database analysis, immunohistochemistry, PCR and Western blot. The effects of TBC1D5 on tumor cell growth, migration, invasion and lipid metabolism were examined by CCK8, Transwell and oil red staining, and the mechanism of TBC1D5 on autophagy was investigated by Western blot, fluorescence microscopy and electron microscopy. Chloroquine and rapamycin were used to verified the key role of autophagy in effects of TBC1D5 on tumor cell. The regulatory mechanism of TBC1D5 in renal clear cell carcinoma (RCC) was investigated by shhif-2α, shTBC1D5, mimic, inhibitor, ChIP and Luciferase experiments. The animal model of ccRCC was used to evaluate the biological function of TBC1D5 in vivo. RESULTS: In this study, TBC1D5 was found to be an important bridge between autophagy and HIF-2α. Specifically, TBC1D5 is significantly underexpressed in ccRCC, serving as a tumor suppressor which inhibits tumor progression and lipid accumulation, and is negatively regulated by HIF-2α. Further research has found that TBC1D5 regulates the autophagy pathway to reverse the biological function of HIF-2α in ccRCC. Mechanism studies have shown that HIF-2α regulates TBC1D5 through hsa-miR-7-5p in ccRCC, thereby affecting tumor progression and lipid metabolism through autophagy. CONCLUSIONS: Our research reveals a completely new pathway, HIF-2α/hsa-miR-7-5p/TBC1D5 pathway affects ccRCC progression and lipid metabolism by regulating autophagy.

miR-1182-mediated ALDH3A2 inhibition affects lipid metabolism and progression in ccRCC by activating the PI3K-AKT pathway.

In clear cell renal cell carcinoma (ccRCC), dysregulated lipid metabolism plays a pivotal role in tumor initiation and progression. This study delves into the unexplored landscape of Dysregulated Aldehyde Dehydrogenase 3 Family Member A2 (ALDH3A2) in ccRCC. Using a combination of "fatty acid metabolism" dataset analysis and differentially expressed genes (DEGs) derived from Gene Expression Omnibus (GEO) database, potential metabolic regulators in ccRCC were identified. Subsequent investigations utilizing public databases, clinical samples, and in vitro experiments revealed that ALDH3A2 was down-regulated in ccRCC, mediated by miR-1182, highlighting its role as an independent prognostic factor for patient survival. Functionally, ALDH3A2 exhibited tumor-suppressive properties, impacting ccRCC cell phenotypes and influencing epithelial-mesenchymal transition. Mechanistically, silencing ALDH3A2 promoted lipid accumulation in ccRCC cells by activating the PI3K-AKT pathway, thereby promoting tumor progression. These findings shed light on the critical role of the miR-1182/ALDH3A2 axis in ccRCC tumorigenesis, emphasizing the potential for targeting lipid metabolism as a promising anti-cancer strategy.

Structural regulation of asphalt-based hard carbon microcrystals based on liquid-phase crosslinking to enhance sodium storage.

Air-oxidation is an effective strategy to obtain promising carbon materials from asphalt for sodium-ion batteries. However, this method would generate a vast amount of gaseous pollutant, which pose challenges for recycling. Herein, a simple, cost-effective and environmentally friendly liquid-phase oxidation method is proposed. The oxygen-containing functional groups (-NO2) are introduced into asphalt, which effectively prevents the melting of asphalt and rearrangement of carbon layers during subsequent carbonization process. As a result, a carbon material with notable disorder degree, large interlayer spacing and abundant closed pores, is prepared. The as-prepared product demonstrates an impressive initial Coulombic efficiency of 88.3 % and an enhanced specific capacity of 317.0 mA h g-1, which is 2.6 times that of the pristine product. Moreover, when assembled with a Na3.32Fe2.34(P2O7)2 cathode, the full-cell delivers a high reversible capacity of 271.7 mA h g-1 at 30 mA g-1 with superb cycle life. This study offers a novel oxidation strategy and provides a solution for producing highly disordered carbon anodes from soft carbon precursors.

Karyotyping of aneuploid and polyploid plants from low coverage whole-genome resequencing.

BACKGROUND: Karyotype, as a basic characteristic of species, provides valuable information for fundamental theoretical research and germplasm resource innovation. However, traditional karyotyping techniques, including fluorescence in situ hybridization (FISH), are challenging and low in efficiency, especially when karyotyping aneuploid and polyploid plants. The use of low coverage whole-genome resequencing (lcWGR) data for karyotyping was explored, but existing methods are complicated and require control samples. RESULTS: In this study, a new protocol for molecular karyotype analysis was provided, which proved to be a simpler, faster, and more accurate method, requiring no control. Notably, our method not only provided the copy number of each chromosome of an individual but also an accurate evaluation of the genomic contribution from its parents. Moreover, we verified the method through FISH and published resequencing data. CONCLUSIONS: This method is of great significance for species evolution analysis, chromosome engineering, crop improvement, and breeding.

Downregulation of the expression of subgenomic chromosome A7 genes promotes plant height in resynthesized allopolyploid Brassica napus.

KEY MESSAGE: Homoeolog expression bias and the gene dosage effect induce downregulation of genes on chromosome A7, causing a significant increase in the plant height of resynthesized allopolyploid Brassica napus. Gene expression levels in allopolyploid plants are not equivalent to the simple average of the expression levels in the parents and are associated with several non-additive expression phenomena, including homoeolog expression bias. However, hardly any information is available on the effect of homoeolog expression bias on traits. Here, we studied the effects of gene expression-related characteristics on agronomic traits using six isogenic resynthesized Brassica napus lines across the first ten generations. We found a group of genes located on chromosome A7 whose expression levels were significantly negatively correlated with plant height. They were expressed at significantly lower levels than their homoeologous genes, owing to allopolyploidy rather than inheritance from parents. Homoeolog expression bias resulted in resynthesized allopolyploids with a plant height similar to their female Brassica oleracea parent, but significantly higher than that of the male Brassica rapa parent. Notably, aneuploid lines carrying monosomic and trisomic chromosome A7 had the highest and lowest plant heights, respectively, due to changes in the expression bias of homoeologous genes because of alterations in the gene dosage. These findings suggest that the downregulation of the expression of homoeologous genes on a single chromosome can result in the partial improvement of traits to a significant extent in the nascent allopolyploid B. napus.

ENO2 as a Biomarker Regulating Energy Metabolism to Promote Tumor Progression in Clear Cell Renal Cell Carcinoma.

BACKGROUND: Clear cell renal cell carcinoma (ccRCC) is the most common and metastatic type of renal cell carcinoma. Despite significant advancements, the current diagnostic biomarkers for ccRCC lack the desired specificity and sensitivity, necessitating the identification of novel biomarkers and elucidation of their underlying mechanisms. METHODS: Three gene expression profile datasets were obtained from the GEO database, and differentially expressed genes (DEGs) were screened. Gene Ontology and KEGG pathway analysis were conducted in ccRCC. To clarify the diagnosis and prognostic role of ENO2, Kaplan-Meier analysis and Cox proportional hazards regression analysis were performed. Functional experiments were also carried out to verify the significant role of ENO2 in ccRCC. Finally, tumor mutational burden analysis was utilized to investigate the potential role of ENO2 in gene mutations in ccRCC. RESULTS: The study showed that ENO2 is a potential biomarker for the diagnosis of ccRCC and can independently predict the clinical prognosis of ccRCC. Furthermore, we found that ENO2 can promote the occurrence and progression of ccRCC by affecting the glycolysis level of cells through the "Warburg effect". CONCLUSIONS: These findings provide new theories for the occurrence and development of ccRCC and can help formulate new strategies for its diagnosis and treatment.

The m6A modification-mediated OGDHL exerts a tumor suppressor role in ccRCC by downregulating FASN to inhibit lipid synthesis and ERK signaling.

Metabolic reprogramming is a hallmark of cancer, and the impact of lipid metabolism as a crucial aspect of metabolic reprogramming on clear cell renal cell carcinoma (ccRCC) progression has been established. However, the regulatory mechanisms underlying the relationship between metabolic abnormalities and ccRCC progression remain unclear. Therefore, this study aimed to identify key regulatory factors of metabolic reprogramming in ccRCC and provide potential therapeutic targets for ccRCC patients. Potential metabolic regulatory factors in ccRCC were screened using bioinformatics analysis. Public databases and patient samples were used to investigate the aberrant expression of Oxoglutarate dehydrogenase-like (OGDHL) in ccRCC. The function of OGDHL in ccRCC growth and metastasis was evaluated through in vitro and in vivo functional experiments. Mechanistic insights were obtained through luciferase reporter assays, chromatin immunoprecipitation, RNA methylation immunoprecipitation, and mutagenesis studies. OGDHL mRNA and protein levels were significantly downregulated in ccRCC tissues. Upregulation of OGDHL expression effectively inhibited ccRCC growth and metastasis both in vitro and in vivo. Furthermore, FTO-mediated OGDHL m6A demethylation suppressed its expression in ccRCC. Mechanistically, low levels of OGDHL promoted TFAP2A expression by inhibiting ubiquitination levels, which then bound to the FASN promoter region and transcriptionally activated FASN expression, thereby promoting lipid accumulation and ERK pathway activation. Our findings demonstrate the impact of OGDHL on ccRCC progression and highlight the role of the FTO/OGDHL/TFAP2A/FASN axis in regulating ccRCC lipid metabolism and progression, providing new targets for ccRCC therapy.

UCP1 alleviates renal interstitial fibrosis progression through oxidative stress pathway mediated by SIRT3 protein stability.

BACKGROUND: Renal interstitial fibrosis is a common pathway for the progressive development of chronic renal diseases (CKD) with different etiology, and is the main pathological basis leading to end-stage renal disease. Although the current research on renal interstitial fibrosis is gradually deepening, the diagnosis and treatment methods are still very lacking. Uncoupling protein 1 (UCP1) is a nuclear encoded protein in mitochondria inner membrane and plays an important role in regulating energy metabolism and mitochondrial homeostasis. However, the biological significance of UCP1 and potential regulatory mechanisms in the development of CKD remain unclear. METHODS: Unilateral ureteral obstruction (UUO) model was used to construct the animal model of renal fibrosis, and TGF-ß1 stimulation of HK2 cells was used to construct the vitro model of renal fibrosis. UCP1 expression was detected by Western blot, immunoblot analysis and immunohistochemistry. UCP1 was upregulated by UCP1 overexpressing lentivirus and UCP1 agonist CL316243. Western blot and immunofluorescence were used to detect epithelial mesenchymal transition (EMT)-related markers, such as collagen I, fibronectin, antioxidant enzyme SOD2 and CAT. Reactive oxygen species (ROS) production was detected by ROS detection kit. SIRT3 knockdown was performed by siRNA. RESULTS: This study presents that UCP1 is significantly downregulated in patients with renal fibrosis and UUO model. Further studies discover that UCP1 overexpression and CL316243 treatments (UCP1 agonists) reversed EMT and extracellular matrix (ECM) accumulation in renal fibrosis models in vivo and in vitro. Simultaneously, UCP1 reduced the ROS production by increasing the stability of SIRT3. When SIRT3 was knocked down, the production of ROS decreased. CONCLUSIONS: Elevating the expression of UCP1 can inhibit the occurrence of oxidative stress by stabilizing SIRT3, thereby reducing EMT and ECM accumulation, and ultimately alleviating renal interstitial fibrosis. It will provide new instructions and targets for the treatment of CKD.

Microvascularized tumor assembloids model for drug delivery evaluation in colorectal cancer-derived peritoneal metastasis.

Peritoneal metastasis (PM) is a fatal state of colorectal cancer, and only a few patients may benefit from systemic chemotherapy. Although hyperthermic intraperitoneal chemotherapy (HIPEC) brings hope for affected patients, the drug development and preclinical evaluation of HIPEC are seriously lagging behind, mainly due to the lack of an ideal in vitro PM model that makes drug development over-reliant on expensive and inefficient animal experiments. This study developed an in vitro colorectal cancer PM model [microvascularized tumor assembloids (vTA)] based on an assembly strategy of endothelialized microvessels and tumor spheroids. Our data showed that the in vitro perfusion cultured vTA could maintain a similar gene expression pattern to their parental xenografts. Also, the drug penetration pattern of the in vitro HIPEC in vTA could mimic the drug delivery behavior in tumor nodules during in vivo HIPEC. More importantly, we further confirmed the feasibility of constructing a tumor burden-controlled PM animal model using vTA. In conclusion, we propose a simple and effective strategy to construct physiologically simulated PM models in vitro, thus providing a basis for PM-related drug development and preclinical evaluation of locoregional therapies. STATEMENT OF SIGNIFICANCE: This study developed an in vitro colorectal cancer peritoneal metastasis (PM) model based on microvascularized tumor assembloids (vTA) for drug evaluation. With perfusion culture, vTA could maintain a similar gene expression pattern and tumor heterogeneity to their parental xenografts. And the drug penetration pattern in vTA was similar to the drug delivery behavior in tumor nodules under in vivo treatment. Moreover, vTA was more conducive to construct PM animal models with controllable tumor burden. In conclusion, the construction of vTA could provide a new strategy for the PM-related drug development and preclinical evaluation of locoregional therapies.

Dosage-sensitivity shapes how genes transcriptionally respond to allopolyploidy and homoeologous exchange in resynthesized Brassica napus.

The gene balance hypothesis proposes that selection acts on the dosage (i.e. copy number) of genes within dosage-sensitive portions of networks, pathways, and protein complexes to maintain balanced stoichiometry of interacting proteins, because perturbations to stoichiometric balance can result in reduced fitness. This selection has been called dosage balance selection. Dosage balance selection is also hypothesized to constrain expression responses to dosage changes, making dosage-sensitive genes (those encoding members of interacting proteins) experience more similar expression changes. In allopolyploids, where whole-genome duplication involves hybridization of diverged lineages, organisms often experience homoeologous exchanges that recombine, duplicate, and delete homoeologous regions of the genome and alter the expression of homoeologous gene pairs. Although the gene balance hypothesis makes predictions about the expression response to homoeologous exchanges, they have not been empirically tested. We used genomic and transcriptomic data from 6 resynthesized, isogenic Brassica napus lines over 10 generations to identify homoeologous exchanges, analyzed expression responses, and tested for patterns of genomic imbalance. Groups of dosage-sensitive genes had less variable expression responses to homoeologous exchanges than dosage-insensitive genes, a sign that their relative dosage is constrained. This difference was absent for homoeologous pairs whose expression was biased toward the B. napus A subgenome. Finally, the expression response to homoeologous exchanges was more variable than the response to whole-genome duplication, suggesting homoeologous exchanges create genomic imbalance. These findings expand our knowledge of the impact of dosage balance selection on genome evolution and potentially connect patterns in polyploid genomes over time, from homoeolog expression bias to duplicate gene retention.

FAM72A promotes glioma progression by regulating mitophagy through the Pink1/Parkin signaling pathway.

Background: There is growing evidence that aberrant expression of FAM72A contributes to biological dysfunction, especially mitochondrial dysfunction. However, its role in most tumors remains unclear, especially in glioma. Methods: Herein, a high-throughput sequencing approach was used here to identify FAM72A as the target molecule. Next, we detected the protein and mRNA expression levels of FAM72A in normal brain tissue (NBT) as well as different grades of glioma tissue. CCK-8, colony formation, Transwell assays, and Western blotting, were all used to determine the molecular effects of FAM72A on glioma cells. Results: FAM72A was significantly upregulated in glioma, was significantly correlated with WHO grade and was associated with poor clinical outcomes. In functional assays, FAM72A was shown to promote glioma cell growth. Subsequent mechanistic studies indicated that FAM72A promoted glioma progression by regulating mitophagy through the Pink1/Parkin signaling pathway. In addition, FAM72A promoted mitophagy and maintained Pink1 stability through the Pink1/Parkin signaling pathway. Finally, FAM72A promoted tumor immune escape by upregulating PD-L1 expression. Conclusion: All of these data indicate that FAM72A confers an aggressive phenotype and poor prognosis on gliomas. Targeting FAM72A might represent a new therapeutic strategy for glioma.

Epigenetic differences in NCOA2: a novel biomarker that predicts prognosis in clear cell renal cell carcinoma.

OBJECTIVES: Kidney cancer is one of the top ten cancers worldwide, and clear cell renal cell carcinoma (ccRCC) is the most common pathohistological type of kidney cancer. This study aimed to decipher the diagnostic and prognostic value of NCOA2 for ccRCC survival based on its expression and methylation. METHODS: We explored the mRNA and protein expression, DNA methylation, prognosis, cell function, and relevant immune infiltration of NCOA2 in ccRCC using data from public databases. Furthermore, GSEA (Gene Set Enrichment Analysis) was used to dissect the cell functions and signal pathways associated with NCOA2 involved in ccRCC and evaluated the close correlation between NCOA2 expression and immune cells. Finally, RT-qPCR (quantitative reverse transcription PCR) and IHC (immunohistochemistry) were utilized to verify the expression of NCOA2 in ccRCC among the tumor and adjacent normal tissues collected from patients. RESULTS: NCOA2 was lowly expressed in ccRCC tissue, which resulted from its methylation. High NCOA2 expression and low beta value of one of the CpG sites predicted better prognosis in patients with ccRCC. GSEA results and analysis of immune infiltration revealed that NCOA2 was associated with PD-1/PD-L1 expression and infiltration of other immune cells in ccRCC. CONCLUSIONS: NCOA2 has great potential to serve as a novel biomarker that can predict prognosis in ccRCC and may become a new therapeutic target in patients with late-stage ccRCC.

Current advances in the molecular regulation of abiotic stress tolerance in sorghum via transcriptomic, proteomic, and metabolomic approaches.

Sorghum (Sorghum bicolor L. Moench), a monocot C4 crop, is an important staple crop for many countries in arid and semi-arid regions worldwide. Because sorghum has outstanding tolerance and adaptability to a variety of abiotic stresses, including drought, salt, and alkaline, and heavy metal stressors, it is valuable research material for better understanding the molecular mechanisms of stress tolerance in crops and for mining new genes for their genetic improvement of abiotic stress tolerance. Here, we compile recent progress achieved using physiological, transcriptome, proteome, and metabolome approaches; discuss the similarities and differences in how sorghum responds to differing stresses; and summarize the candidate genes involved in the process of responding to and regulating abiotic stresses. More importantly, we exemplify the differences between combined stresses and a single stress, emphasizing the necessity to strengthen future studies regarding the molecular responses and mechanisms of combined abiotic stresses, which has greater practical significance for food security. Our review lays a foundation for future functional studies of stress-tolerance-related genes and provides new insights into the molecular breeding of stress-tolerant sorghum genotypes, as well as listing a catalog of candidate genes for improving the stress tolerance for other key monocot crops, such as maize, rice, and sugarcane.

Tumor-Associated Macrophage-Derived Exosomal LINC01232 Induces the Immune Escape in Glioma by Decreasing Surface MHC-I Expression.

Tumor-associated macrophage (TAM) infiltration facilitates glioma malignancy, but the underlying mechanisms remain unclear. Herein, it is reported that TAMs secrete exosomal LINC01232 to induce tumor immune escape. Mechanistically, LINC01232 is found to directly bind E2F2 and promote E2F2 entry into the nucleus; the two synergistically promots the transcription of NBR1. The increase in binding between NBR1 binding and the ubiquitinating MHC-I protein through the ubiquitin domain causes an increase in the degradation of MHC-I in autophagolysosomes and a decrease in the expression of MHC-I on the surface of tumor cells, which in turn led to tumor cell escape from CD8+ CTL immune attack. Disruption of E2F2/NBR1/MHC-I signaling with shRNAs or blockade with the corresponding antibodies largely abolishes the tumor-supportive effects of LINC01232 and inhibits tumor growth driven by M2-type macrophages. Importantly, knockdown of LINC01232 enhances the expression of MHC-I on the surface of tumor cells and improves the response to reinfusion with CD8+ T cells. This study reveals the existence of critical molecular crosstalk between TAMs and glioma mediates through the LINC01232/E2F2/NBR1/MHC-I axis to support malignant tumor growth, indicating that targeting this axis may have therapeutic potential.

N6-methyladenosine-modified DBT alleviates lipid accumulation and inhibits tumor progression in clear cell renal cell carcinoma through the ANXA2/YAP axis-regulated Hippo pathway.

BACKGROUND: The mechanism of metabolism reprogramming is an unsolved problem in clear cell renal cell carcinoma (ccRCC). Recently, it was discovered that the Hippo pathway altered tumor metabolism and promoted tumor progression. Thus, this study aimed at identifying key regulators of metabolism reprogramming and the Hippo pathway in ccRCC and pinpointing potential therapeutic targets for ccRCC patients. METHODS: Hippo-related gene sets and metabolic gene sets were used to screen potential regulators of the Hippo pathway in ccRCC. Public databases and samples from patients were applied to investigate the association of dihydrolipoamide branched chain transacylase E2 (DBT) with ccRCC and Hippo signaling. The role of DBT was confirmed by gain or loss of function assays in vitro and in vivo. Mechanistic results were yielded by luciferase reporter assay, immunoprecipitation, mass spectroscopy, and mutational studies. RESULTS: DBT was confirmed as a Hippo-related marker with significant prognostic predictive value, and its downregulation was caused by methyltransferase-like-3 (METTL3)-mediated N6-methyladenosine (m6 A) modification in ccRCC. Functional studies specified DBT as a tumor suppressor for inhibiting tumor progression and correcting the lipid metabolism disorder in ccRCC. Mechanistic findings revealed that annexin A2 (ANXA2) interacted with the lipoyl-binding domain of DBT to activate Hippo signaling which led to decreased nuclear localization of yes1-associated transcriptional regulator (YAP) and transcriptional repression of lipogenic genes. CONCLUSIONS: This study demonstrated a tumor-suppressive role for the DBT/ANXA2/YAP axis-regulated Hippo signaling and suggested DBT as a potential target for pharmaceutical intervention in ccRCC.

Genome balance and dosage effect drive allopolyploid formation in Brassica.

Polyploidy is a major evolutionary force that has shaped plant diversity. However, the various pathways toward polyploid formation and interploidy gene flow remain poorly understood. Here, we demonstrated that the immediate progeny of allotriploid AAC Brassica (obtained by crossing allotetraploid Brassica napus and diploid Brassica rapa) was predominantly aneuploids with ploidal levels ranging from near-triploidy to near-hexaploidy, and their chromosome numbers deviated from the theoretical distribution toward increasing chromosome numbers, suggesting that they underwent selection. Karyotype and phenotype analyses showed that aneuploid individuals containing fewer imbalanced chromosomes had higher viability and fertility. Within three generations of self-fertilization, allotriploids mainly developed into near or complete allotetraploids similar to B. napus via gradually increasing chromosome numbers and fertility, suggesting that allotriploids could act as a bridge in polyploid formation, with aneuploids as intermediates. Self-fertilized interploidy hybrids ultimately generated new allopolyploids carrying different chromosome combinations, which may create a reproductive barrier preventing allotetraploidy back to diploidy and promote gene flow from diploids to allotetraploids. These results suggest that the maintenance of a proper genome balance and dosage drove the recurrent conversion of allotriploids to allotetraploids, which may contribute to the formation and evolution of polyploids.