Trigred motif 36 regulates neuroendocrine differentiation of prostate cancer via HK2 ubiquitination and GPx4 deficiency.

Neuroendocrine prostate cancer (NEPC), the most lethal subtype of castration-resistant prostate cancer (PCa), may evolve from the neuroendocrine differentiation (NED) of PCa cells. However, the molecular mechanism that triggers NED is unknown. Trigred motif 36 (TRIM36), a member of the TRIM protein family, exhibits oncogenic or anti-oncogenic roles in various cancers. We have previously reported that TRIM36 is highly expressed to inhibit the invasion and proliferation of PCa. In the present study, we first found that TRIM36 was lowly expressed in NEPC and its overexpression suppressed the NED of PCa. Next, based on proteomic analysis, we found that TRIM36 inhibited the glycolysis pathway through suppressing hexokinase 2 (HK2), a crucial glycolytic enzyme catalyzing the conversion of glucose to glucose-6-phosphate. TRIM36 specifically bound to HK2 through lysine 48 (lys48)-mediated ubiquitination of HK2. Moreover, TRIM36-mediated ubiquitination degradation of HK2 downregulated the level of glutathione peroxidase 4 (GPx4), a process that contributed to ferroptosis. In conclusion, TRIM36 can inhibit glycolysis via lys48-mediated HK2 ubiquitination to reduce GPX4 expression and activate ferroptosis, thereby inhibiting the NED in PCa. Targeting TRIM36 might be a promising approach to retard NED and treat NEPC.

TRIM36 enhances lung adenocarcinoma radiosensitivity and inhibits tumorigenesis through promoting RAD51 ubiquitination and antagonizing hsa-miR-376a-5p.

The tripartite motif (TRIM) family proteins exhibit oncogenic or anti-oncogenic roles in various cancers. As a TRIM family member, TRIM36 is expressed in lung adenocarcinoma (LUAD), but its roles remain unexplored. Here, we set to investigate the clinical relevance and the biological actions of TRIM36 in LUAD. mRNA levels of TRIM36 and the Kaplan-Meier survival analysis for patients with LUAD were analyzed from The Cancer Genome Atlas (TCGA) database. Real-time PCR and western blotting were utilized to measure TRIM36 levels both in vivo and in vitro. We demonstrated that TRIM36 levels were significantly decreased in LUAD patients. The low expression of TRIM36 was correlated with a poor prognosis. Overexpression and knock-down studies illustrated that TRIM36 inhibits cell proliferation and promotes apoptosis in LUAD cell lines. LUAD cells were irradiated with 60Co. In addition, TRIM36 enhanced radiosensitivity in LUAD cell lines. Moreover, we found that TRIM36 expression was negatively associated with RAD51. Co-overexpressing RAD51 blocked TRIM36's effects on proliferation and apoptosis. TRIM36 formed a complex with RAD51, promoting its ubiquitination. Inhibiting hsa-miR-376a-5p enhanced apoptosis and the effects were mediated by TRIM36 in response to radiation. In conclusion, our results indicated that TRIM36 is anti-oncogenic in LUAD by promoting RAD1 ubiquitination. Hsa-miR-376a-5p acts upstream of TRIM36. TRIM36 and RAD1 may serve as prognostic indicators for LUAD. The interactions between TRIM36, RAD1 and hsa-miR-376a-5p can be future therapeutic targets to increase radiation sensitivity in LUAD patients.

TRIM36 inhibits tumorigenesis through the Wnt/ß-catenin pathway and promotes caspase-dependent apoptosis in hepatocellular carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is the most common type of primary liver cancer and has an extremely poor prognosis. We aimed to determine the latent relationships between TRIM36 regulation of apoptosis and the Wnt/ß-catenin pathway in HCC. METHODS: Immunohistochemistry and western blotting were used to characterize the aberrant expression of TRIM36 in HCC and adjacent tissues. Clinical information was analyzed using Kaplan-Meier and Cox methods. RNA-seq of potential targets was conducted to detect the regulation of TRIM36. Apoptosis assays and cellular proliferation, invasion and migration were conducted in a loss- and gain-of-function manner in cultured cells to determine the biological functions of TRIM36. A rescue experiment was conducted to confirm the role of Wnt/ß-catenin signaling in TRIM36 regulation. Finally, in vivo experiments were conducted using cell line-derived xenografts in nude mice to validate the central role of TRIM36 in HCC. RESULTS: TRIM36 expression was significantly downregulated in HCC tissues compared to adjacent non-tumor tissues. TRIM36 repressed the proliferation, migration, and invasion of Huh7 and HCCLM3 cells, whereas it stimulated apoptosis. Wnt/ß-catenin signaling was inhibited by TRIM36, and rescue experiments highlighted its importance in HCC proliferation, migration, and invasion. In vivo experiments further confirmed the effects of sh-TRIM36 on HCC tumorigenesis, inhibition of apoptosis, and promotion of Wnt/ß-catenin signaling. CONCLUSION: Our study is the first to indicate that TRIM36 acts as a tumor suppressor in HCC. TRIM36 activates apoptosis and inhibits cellular proliferation, invasion, and migration via the Wnt/ß-catenin pathway, which may serve as an important biomarker and promising therapeutic target for HCC.

Haprin-deficient spermatozoa are incapable of in vitro fertilization.

Haprin (TRIM36) is a ubiquitin-protein ligase that mediates ubiquitination and subsequent proteasomal degradation of target proteins. It is expressed in the testes in both mice and humans and is thought to be involved in spermiogenesis, the acrosome reaction, and fertilization. However, the functional role of Haprin is poorly understood. The aim of this study was to investigate the physiological role of Haprin in fertility. Homozygous haprin-deficient mice were generated and these mice, and their spermatozoa, were analyzed to detect morphological and fertility-related abnormalities. In these models, normal spermatogenesis was observed but sperm quality was reduced with haprin-deficient mice having poorer sperm morphology and motility than wild-type mice. Interestingly, haprin-deficient mice showed normal in vivo fertility but could not fertilize oocytes under standard in vitro fertilization conditions. In conclusion, this study demonstrated that Haprin deficiency causes morphological abnormalities in spermatozoa, indicating that Haprin is involved in spermiogenesis.

The dynamics of plus end polarization and microtubule assembly during Xenopus cortical rotation.

The self-organization of dorsally-directed microtubules during cortical rotation in the Xenopus egg is essential for dorsal axis formation. The mechanisms controlling this process have been problematic to analyze, owing to difficulties in visualizing microtubules in living egg. Also, the order of events occurring at the onset of cortical rotation have not been satisfactorily visualized in vivo and have been inferred from staged fixed samples. To address these issues, we have characterized the dynamics of total microtubule and plus end behavior continuously throughout cortical rotation, as well as in oocytes and unfertilized eggs. Here, we show that the nascent microtubule network forms in the cortex but associates with the deep cytoplasm at the start of rotation. Importantly, plus ends remain cortical and become increasingly more numerous and active prior to rotation, with dorsal polarization occurring rapidly after the onset of rotation. Additionally, we show that vegetally localized Trim36 is required to attenuate dynamic plus end growth, suggesting that vegetal factors are needed to locally coordinate growth in the cortex.

Specific CpG sites methylation is associated with hematotoxicity in low-dose benzene-exposed workers.

Benzene is a broadly used industrial chemicals which causes various hematologic abnormalities in human. Altered DNA methylation has been proposed as epigenetic biomarkers in health risk evaluation of benzene exposure, yet the role of methylation at specific CpG sites in predicting hematological effects remains unclear. In this study, we recruited 120 low-level benzene-exposed and 101 control male workers from a petrochemical factory in Maoming City, Guangdong Province, China. Urinary S-phenylmercapturic acid (SPMA) in benzene-exposed workers was 3.40-fold higher than that in control workers (P < 0.001). Benzene-induced hematotoxicity was characterized by reduced white blood cells counts and nuclear division index (NDI), along with an increased DNA damage and urinary 8-hydroxy-2'-deoxyguanosine (all P < 0.05). Methylation levels of TRIM36, MGMT and RASSF1a genes in peripheral blood lymphocytes (PBLCs) were quantified by pyrosequencing. CpG site 6 of TRIM36, CpG site 2, 4, 6 of RASSF1a and CpG site 1, 3 of MGMT methylation were recognized as hot CpG sites due to a strong correlation with both internal exposure and hematological effects. Notably, integrating hot CpG sites methylation of multiple genes reveal a higher efficiency in prediction of integrative damage compared to individual genes at hot CpG sites. The negative dose-response relationship between the combined methylation of hot CpG sites in three genes and integrative damage enabled the classification of benzene-exposed individuals into high-risk or low-risk groups using the median cut-off value of the integrative index. Subsequently, a prediction model for integrative damage in benzene-exposed populations was built based on the methylation status of the identified hot CpG sites in the three genes. Taken together, these findings provide a novel insight into application prospect of specific CpG site methylation as epi-biomarkers for health risk assessment of environmental pollutants.

FOXA2 Suppression by TRIM36 Exerts Anti-Tumor Role in Colorectal Cancer Via Inducing NRF2/GPX4-Regulated Ferroptosis.

The forkhead box transcription factor A2 (FOXA2) is a transcription factor and plays a key role in embryonic development, metabolism homeostasis and tumor cell proliferation; however, its regulatory potential in CRC is not fully understood. Here, it is found that FOXA2 expression is markedly up-regulated in tumor samples of CRC patients as compared with the normal tissues, which is closely associated with the worse survival in patients with CRC. Notably, a positive correlation between FOXA2 and nuclear factor erythroid 2-related factor 2 (Nrf2)/glutathione peroxidase 4 (GPX4) gene expression is observed in CRC patients. Mechanistically, FOXA2 depletion weakens the activation of Nrf2 pathway and decreases GPX4 level in CRC cells, thereby leading to ferroptosis, which is further supported by bioinformatic analysis. More intriguingly, the E3 ubiquitin ligase tripartite motif containing 36 (TRIM36) is identified as a key suppressor of FOXA2, and it is observed that TRIM36 can directly interact with FOXA2 and induce its K48-linked polyubiquitination, resulting in FOXA2 protein degradation in vitro. Taken together, all the studies demonstrate that FOXA2 mediated by TRIM36 promotes CRC progression by inhibiting the Nrf2/GPX4 ferroptosis signaling pathway, thus providing a new therapeutic target for CRC treatment.

TRIM36 suppresses cell growth and promotes apoptosis in human esophageal squamous cell carcinoma cells by inhibiting Wnt/ß-catenin signaling pathway.

Our recent study has shown that TRIM36, a member of tripartite motif-containing (TRIM) family proteins and tumor suppressor and ß-catenin may serve as a prognostic biomarker for esophageal squamous cell carcinoma (ESCC). Here, we sought to examine functional roles of TRIM36 and ß-catenin in ESCC cells. TRIM36 was overexpressed or silenced by lentivirus transduction. Cell proliferation was examined by Cell Counting Kit (CCK)-8 assay, while cell cycle distribution and cell apoptosis was assessed via flow cytometry analysis. Xenograft mouse model was applied for in vivo analysis. Overexpression of TRIM36 inhibited cell proliferation in human ESCC cells, and silencing of TRIM36 led to opposite effects. We also found that ectopic expression of TRIM36 enhanced the ratio of G0/G1 phase cells and induced apoptosis in ESCC cells. Our data further revealed that TRIM36 stimulated the ubiquitination of ß-catenin, and in turn, its inactivation. Finally, we confirmed these in vitro results in a xenograft mouse model and clinical specimens post-operatively obtained from patients of ESCC. In summary, these data support that TRIM36 can effectively inhibit tumorigenesis of ESCC by repressing Wnt/ß-catenin signaling pathway, which suggest that selectively repressing this signaling pathway in ESCC may lead to development of a novel therapeutic approach for controlling this disease.

Microtubular TRIM36 E3 Ubiquitin Ligase in Embryonic Development and Spermatogenesis.

TRIM36 is a member of the tripartite motif (TRIM) family of RING-containing proteins, also known as Haprin, which was first discovered for its abundance in testis and found to be implicated in the spermatozoa acrosome reaction. TRIM36 is a microtubule-associated E3 ubiquitin ligase that plays a role in cytoskeletal organization, and according to data gathered in different species, coordinates growth speed and stability, acting on the microtubules' plus end, and impacting on cell cycle progression. TRIM36 is also crucial for early developmental processes, in Xenopus, where it is needed for dorso-ventral axis formation, but also in humans as bi-allelic mutations in the TRIM36 gene cause a form of severe neural tube closure defect, called anencephaly. Here, we review TRIM36-related mechanisms implicated in such composite physiological and pathological processes.

Extracellular Polysaccharide from Rhizopus nigricans Inhibits Hepatocellular Carcinoma via miR-494-3p/TRIM36 Axis and Cyclin E Ubiquitination.

Background and Aims: This study was designed to uncover the mechanism for extracellular polysaccharide (EPS1-1)-mediated effects on hepatocellular carcinoma (HCC) development. Methods: HCC cells were treated with EPS1-1, miR-494-3p mimic, sh-TRIM36, and pcDNA3.1-TRIM36. The levels of miR-494-3p and TRIM36 were measured in normal hepatocytes, THLE-2, and HepG2 and HuH7HCC cell lines, along with the protein expression of cyclin D/E and p21. The proliferation, cell cycle, and apoptosis of HCC cells were assayed. The interactions between miR-494-3p and TRIM36, and between TRIM36 and cyclin E were assessed. Finally, the expression and localization of TRIM36 and cyclin E were monitored, and tumor apoptosis was detected, in tumor xenograft model. Results: EPS1-1 suppressed HCC cell proliferation and cyclin D/E expression and promoted apoptosis and p21 expression. miR-494-3p was upregulated and TRIM36 was downregulated in HCC cells. Transfection with miR-494-3p mimic or sh-TRIM36 facilitated HCC cell proliferation and the expression of cyclin D/E protein but they inhibited apoptosis and p21 expression in the presence of EPS1-1. Overexpression of TRIM36 further consolidated EPS1-1-mediated inhibition of HCC proliferation, cyclin D/E, and the promotion of apoptosis and p21 expression. Those effects were reversed by miR-494-3p overexpression. TRIM36 was a target gene of miR-494-3p, and TRIM36 induced cyclin E ubiquitination. EPS1-1 suppressed cyclin E expression, promoted TRIM36 expression and tumor apoptosis, all of which were abrogated by increasing the expression of miR-494-3p in vivo. Conclusions: EPS1-1 protected against HCC by limiting its proliferation and survival through the miR-494-3p/TRIM36 axis and by inducing cyclin E ubiquitination.