MUL1 identified as mitochondria-linked biomarker promoting cisplatin resistance in OC cells.

Ovarian cancer (OC) ranks among the prevalent tumors affecting the female reproductive system. The aim of this study was to evaluate mitochondria-associated platinum resistance genes using organoid models. Univariate Cox regression, LASSO and multivariate Cox regression analyses were performed on The Cancer Genome Atlas (TCGA) database to construct 2-gene prognostic signature (MUL1 and SSBP1), and GSE26712 dataset was used for external validation. In addition, the relationship between MUL1 and platinum resistance was examined by organoid culture, lentiviral transduction, CCK8 assay, and Western blot. The results showed that patients in the high-risk group exhibited significantly worse OS (P = 0.002, P = 0.017). Drug sensitivity analysis revealed that platinum resistance increased with the upregulation of MUL1 expression (Cor = 0.5154, P = 0.02). Our experimental findings demonstrated that knockout of the MUL1 gene significantly increased apoptosis and enhanced the sensitivity of the OC cell line A2780 to cisplatin. Through this study, we have provided strong evidence for further research on prognostic risk factors and individualized treatment in OC patients, and provided new insights into addressing platinum resistance in OC.

The effect of TFAP2A/ANXA8 axis on ferroptosis of cervical squamous cell carcinoma (CESC) in vitro.

Potential role and associated mechanisms of Annexin A8 (ANXA8), a member of the Annexins family, in cervical squamous cell carcinoma (CESC) are still unclear, despite being upregulated in various malignant tumors. Here, we observed a notably elevated expression of ANXA8 in CESC cells. The inhibition of ANXA8 amplified the susceptibility of CESC cells to Erastin and sorafenib-induced ferroptosis, whereas it exerted minimal influence on DPI7 and DPI10-induced ferroptosis. The results from the Fe2+ concentration assay showed no significant correlation between ANXA8 gene knockdown and intracellular Fe2+ concentration induced by ferroptosis inducers. Western blot analysis demonstrated that the knockdown of ANXA8 did not alter ACSL4 and LPCAT levels under ferroptosis-inducing conditions, but it did result in a reduction in intracellular GSH levels induced by the ferroptosis inducer. Subsequently, we identified TFAP2A as an upstream transcription factor of ANXA8, which plays a role in regulating cell ferroptosis. The knockdown of TFAP2A significantly elevated MDA levels and depressed GSH levels in the presence of a ferroptosis inducer, thereby inhibiting cell ferroptosis. However, this inhibitory effect could be reversed by ANXA8 overexpression. Therefore, our research suggests that the TFAP2A/ANXA8 axis exerts regulatory control over ferroptosis in CESC cells by mediating GSH synthesis in System Xc.

Identifying Immune Cell Infiltration and Hub Genes Related to M2 Macrophages in Endometriosis by Bioinformatics Analysis.

Endometriosis (EM) is a chronic, estrogen-dependent inflammatory disease. Presently, the pathophysiology of EM is still unclear, and numerous studies have established that the immune system plays a major role in the pathophysiology of EM. Six microarray datasets were downloaded from the GEO public database. A total of 151 endometrial samples (72 ectopic endometria and 79 controls) were included in this study. CIBERSORT and ssGSEA were applied to calculate the immune infiltration of EM and control samples. Moreover, we validated four different correlation analyses to explore immune microenvironment of EM and finally identified M2 macrophage-related hub genes and further conducted the specific immunologic signaling pathway analysis by GSEA. The logistic regression model was investigated by ROC and further validated by two external datasets. From the results of the two immune infiltration assays, we concluded that M2 macrophages, regulatory T cells (Tregs), M1 macrophages, activated B cells, T follicular helper cells, activated dendritic cells, and resting NK cells have a significant difference between control and EM tissues. Through multidimensional correlation analysis, we found that macrophages play an important central role in cell-to-cell interactions, especially M2 macrophages. Four immune-related hub genes, namely FN1, CCL2, ESR1, and OCLN, are closely related to M2 macrophages and play a crucial role in the occurrence and immune microenvironment of endometriosis. The combined AUC of ROC prediction model in test and validation sets were 0.9815 and 0.8206, respectively. We conclude that M2 macrophages play a central role in the immune-infiltrating microenvironment of EM.

Identification and validation of shared genes and key pathways in endometriosis and endometriosis-associated ovarian cancer by weighted gene co-expression network analysis and machine learning algorithms.

BACKGROUND: Epidemiological studies reported that patients with endometriosis had an increased risk of developing endometriosis-associated ovarian cancer (EAOC). The present study aimed to identify shared genes and key pathways that commonly interacted between EAOC and endometriosis. METHODS: The expression matrix of ovarian cancer and endometriosis were collected from the Gene Expression Omnibus database. The weighted gene co-expression network analysis (WGCNA) was used to construct co-expression gene network. Machine learning algorithms were applied to identify characteristic genes. CIBERSORT deconvolution algorithm was used to explore the difference in tumor immune microenvironment. Furthermore, diagnostic nomogram was constructed and evaluated for supporting clinical practicality. RESULTS: We identified 262 shared genes between EAOC and endometriosis via WGCNA analysis. They were mainly enriched in cytokine-cytokine receptor interaction. After protein-protein interaction network and machine learning algorithms, we recognized two characteristic genes (EDNRA, OCLN) and established a nomogram that presented an outstanding predictive performance. The hub genes demonstrated remarkable associations with immunological functions. Survival analysis indicated that dysregulated expressions of EDNRA and OCLN were closely correlated with prognosis of ovarian cancer patients. gene set enrichment analyses revealed that the two characteristic genes were mainly enriched in the cancer- and immune-related pathways. CONCLUSION: Our findings pave the way for further investigation of potential candidate genes and will aid in improving the diagnosis and treatment of EAOC in endometriosis patients. More research is required to determine the exact mechanisms by which these two hub genes affecting the development and progression of EAOC from endometriosis.

Histone modifications regulate pioneer transcription factor cooperativity.

Pioneer transcription factors have the ability to access DNA in compacted chromatin1. Multiple transcription factors can bind together to a regulatory element in a cooperative way, and cooperation between the pioneer transcription factors OCT4 (also known as POU5F1) and SOX2 is important for pluripotency and reprogramming2-4. However, the molecular mechanisms by which pioneer transcription factors function and cooperate on chromatin remain unclear. Here we present cryo-electron microscopy structures of human OCT4 bound to a nucleosome containing human LIN28B or nMATN1 DNA sequences, both of which bear multiple binding sites for OCT4. Our structural and biochemistry data reveal that binding of OCT4 induces changes to the nucleosome structure, repositions the nucleosomal DNA and facilitates cooperative binding of additional OCT4 and of SOX2 to their internal binding sites. The flexible activation domain of OCT4 contacts the N-terminal tail of histone H4, altering its conformation and thus promoting chromatin decompaction. Moreover, the DNA-binding domain of OCT4 engages with the N-terminal tail of histone H3, and post-translational modifications at H3K27 modulate DNA positioning and affect transcription factor cooperativity. Thus, our findings suggest that the epigenetic landscape could regulate OCT4 activity to ensure proper cell programming.

Histone modifications regulate pioneer transcription factor binding and cooperativity.

Pioneer transcription factors have the ability to access DNA in compacted chromatin. Multiple transcription factors can bind together to a regulatory element in a cooperative way and cooperation between pioneer transcription factors Oct4 and Sox2 is important for pluripotency and reprogramming. However, the molecular mechanisms by which pioneer transcription factors function and cooperate remain unclear. Here we present cryo-EM structures of human Oct4 bound to a nucleosome containing human Lin28B and nMatn1 DNA sequences, which bear multiple binding sites for Oct4. Our structural and biochemistry data reveal that Oct4 binding induces changes to the nucleosome structure, repositions the nucleosomal DNA and facilitates cooperative binding of additional Oct4 and of Sox2 to their internal binding sites. The flexible activation domain of Oct4 contacts the histone H4 N-terminal tail, altering its conformation and thus promoting chromatin decompaction. Moreover, the DNA binding domain of Oct4 engages with histone H3 N-terminal tail, and posttranslational modifications at H3K27 modulate DNA positioning and affect transcription factor cooperativity. Thus, our data show that the epigenetic landscape can regulate Oct4 activity to ensure proper cell reprogramming.

Applications and prospects of cryo-EM in drug discovery.

Drug discovery is a crucial part of human healthcare and has dramatically benefited human lifespan and life quality in recent centuries, however, it is usually time- and effort-consuming. Structural biology has been demonstrated as a powerful tool to accelerate drug development. Among different techniques, cryo-electron microscopy (cryo-EM) is emerging as the mainstream of structure determination of biomacromolecules in the past decade and has received increasing attention from the pharmaceutical industry. Although cryo-EM still has limitations in resolution, speed and throughput, a growing number of innovative drugs are being developed with the help of cryo-EM. Here, we aim to provide an overview of how cryo-EM techniques are applied to facilitate drug discovery. The development and typical workflow of cryo-EM technique will be briefly introduced, followed by its specific applications in structure-based drug design, fragment-based drug discovery, proteolysis targeting chimeras, antibody drug development and drug repurposing. Besides cryo-EM, drug discovery innovation usually involves other state-of-the-art techniques such as artificial intelligence (AI), which is increasingly active in diverse areas. The combination of cryo-EM and AI provides an opportunity to minimize limitations of cryo-EM such as automation, throughput and interpretation of medium-resolution maps, and tends to be the new direction of future development of cryo-EM. The rapid development of cryo-EM will make it as an indispensable part of modern drug discovery.

Non-canonical bivalent H3K4me3K9me3 recognition by Spindlin1/C11orf84 complex.

Bivalent chromatin with active H3K4me3 and repressive H3K27me3 was initially identified in embryonic stem cells (ESCs) to poise expression of developmental genes upon lineage commitment. Since then, many more different bivalent modifications have been demonstrated in both ESCs and fully differentiated cells. Bivalency not only spatiotemporally controls gene transcription but also acts to fine-tune the level of transcription during development. Although increasing number of studies demonstrated the functional significance of bivalent chromatin, the molecular connection of bivalent chromatin and transcriptional regulation remains largely elusive. Recently, we showed Spindlin1/C11orf84 complex prefers to recognize the non-canonical histone H3K4me3K9me3 bivalent mark, which is required for timely ribosomal RNA transcription. Here, we hypothesize the recognition of K4me3 and K9me3 at the same histone tail by Spindlin1/C11orf84 complex may serve as a general mechanism of conversion from a repressed to an active chromatin structure for transcriptional activation.

Withaferin A inhibits proliferation of human endometrial cancer cells via transforming growth factor-ß (TGF-ß) signalling.

The present study was designed to evaluate the anticancer effects of withaferin A against the human endometrial cancer via modulation of transforming growth factor-ß (TGF-ß) signalling. The results of the present study revealed that withaferin A exerts a dose and time-dependent antiproliferative effects against the human KLE endometrial cancer cells with comparatively lower toxicity against the THESCs normal cells. The IC50 of withaferin A against the KLE endometrial cancer cells was found to 10 µM. The results showed that withaferin A induced apoptosis and G2/M cell cycle arrest of the KLE cells which was associated with alteration of the apoptosis and cell cycle related proteins. In addition, the transwell assays showed that the migration and invasion of the KLE cells were inhibited by 53 and 40%, respectively. Finally, the effects of withaferin A were also examined on the TGF-ß signalling pathway. The results showed that withaferin A blocked TGF-ß-dependent Smad2 phosphorylation and expression of other TGF-ß-related proteins in KLE cells. Summing up, the results suggest that withaferin A inhibits the proliferation of the human endometrial carcinoma via TGF-ß signalling.

Structural mechanism of bivalent histone H3K4me3K9me3 recognition by the Spindlin1/C11orf84 complex in rRNA transcription activation.

Spindlin1 is a unique multivalent epigenetic reader that facilitates ribosomal RNA transcription. In this study, we provide molecular and structural basis by which Spindlin1 acts in complex with C11orf84 to preferentially recognize non-canonical bivalent mark of trimethylated lysine 4 and lysine 9 present on the same histone H3 tail (H3K4me3K9me3). We demonstrate that C11orf84 binding stabilizes Spindlin1 and enhances its association with bivalent H3K4me3K9me3 mark. The functional analysis suggests that Spindlin1/C11orf84 complex can displace HP1 proteins from H3K4me3K9me3-enriched rDNA loci, thereby facilitating the conversion of these poised rDNA repeats from the repressed state to the active conformation, and the consequent recruitment of RNA Polymerase I for rRNA transcription. Our study uncovers a previously unappreciated mechanism of bivalent H3K4me3K9me3 recognition by Spindlin1/C11orf84 complex required for activation of rRNA transcription.