Aberrant HIF1- α and SIX-1 Expression is Associated with Poor Prognosis in Acute Myeloid Leukemia Patients with Isocitrate Dehydrogenase 1 Mutations.

BACKGROUND: IDH1 mutations are common in many cancers, however, their role in promoting the Warburg effect remains elusive. This study elucidates the putative involvement of mutant-IDH1 in regulating hypoxia-inducible factor (HIF1-α) and Sine-Oculis Homeobox-1 (SIX-1) expression. METHODOLOGY: Genetic screening was performed using the ARMS-PCR in acute myeloid leukemia (AML), brain, and breast cancer (BC) cohorts, while transcript expression was determined using qPCR. Further, a meta-analysis of risk factors associated with the R132 mutation was performed. RESULTS: Approximately 32% of AML and ∼60% of glioma cases were mutants, while no mutation was found in the BC cohort. 'AA' and TT' were associated with higher disease risk (OR = 12.18 & 4.68) in AML and had significantly upregulated IDH1 expression. Moreover, downregulated HIF1-α and upregulated SIX-1 expression was also observed in these patients, suggesting that mutant-IDH1 may alter glucose metabolism. Perturbed IDH1 and HIF-α levels exhibited poor prognosis in univariate and multivariate analysis, while age and gender were found to be contributory factors as well. Based on the ROC model, these had a good potential to be used as prognostic markers. A significant variation in frequencies of R132 mutations in AML among different populations was observed. Cytogenesis (R2 = 12.2%), NMP1 mutation status (R2 = 18.5%), and ethnic contributions (R2 = 73.21%) were critical moderators underlying these mutations. Women had a higher risk of R132 mutation (HR = 1.3, P < 0.04). The pooled prevalence was calculated to be 0.29 (95% CI 0.26-0.33, P < 0.01), indicating that IDH1 mutations are a significant prognostic factor in AML. CONCLUSION: IDH1 and HIF1-α profiles are linked to poor survival and prognosis, while high SIX-1 expression in IDH1 mutants suggests a role in leukemic transformation and therapy response in AML.

IDH1 mutations are common in many types of cancer, but scientists have not fully understood how they contribute to the Warburg effect - a process that alters glucose metabolism in cells. In this study, we evaluate the association between mutant-IDH1 and HIF1 as well as SIX-1 gene expression. We analyzed genetic data from patients with brain cancer, breast cancer, and acute myeloid leukemia (AML), and found that roughly 32% of AML cases and 60% of glioma cases had IDH1 mutations, while no mutations were found in breast cancer. Patients with mutant genotypes had a higher risk of disease and showed upregulated IDH1 expression. They also had downregulated HIF1 and upregulated SIX-1 expression, suggesting that mutant-IDH1 can change glucose metabolism in cancer cells. Patients with abnormal IDH1 and HIF1 levels were more likely to have a poor prognosis. Further, we identified several risk factors that can influence IDH1 mutations, including cytogenesis, NMP1 mutation status, and ethnicity. The researchers calculated that IDH1 mutations are a significant factor in predicting outcomes for AML.

miR-128-3p alleviates airway inflammation in asthma by targeting SIX1 to regulate mitochondrial fission and fusion.

Bronchial asthma is known for airway inflammation, hyperresponsiveness, and remodeling.MicroRNAs (MiRNAs) have been involved in the development of asthma, whereas, the mechanism of various MiRNAs in asthma remains to be elucidated. In this study, we aim to explore the mechanism of miR-128-3p in asthma-related airway inflammation by targeting sine oculis homeobox homolog 1 (SIX1) to regulate the mitochondrial function. In an ovalbumin (OVA) asthma mouse model, miR-128-3p levels were found to be significantly diminished. Administration of miR-128-3p agomir decreased peribronchial inflammatory cell infiltration and improved airway inflammation. Afterwards, we used the luciferase reporter assay to predict and confirmed that SIX1 is a target gene of miR-128-3p. Overexpression of miR-128-3p attenuated IL-13-induced cellular inflammation and ROS production in bronchial epithelial cells (BEAS-2B). In vitro, overexpression of miR-128-3p and SIX1 knockdown mitigated mitochondrial fragmentation, reduced Drp1-mediated mitochondrial division, and upregulated mitochondrial membrane potential. Moreover, led to decreased production of ROS/mitochondrial ROS, P-Drp1(616) and Fis1 expression, while enhancing P-Drp1(637), MFN1, caspase-3/9, and Bax-mediated apoptosis. Our findings demonstrated that miR-128-3p could alleviate airway inflammation by downregulating SIX1 and improving mitochondrial function, positioning the miR-128-3p/SIX1/Drp1 signaling as a potential therapeutic target for asthma.

Novel LncRNA LINC02936 Suppresses Ferroptosis and Promotes Tumor Progression by Interacting with SIX1/CP Axis in Endometrial Cancer.

Endometrial cancer (EC) is a prevalent gynecological malignancy, and metabolic disorders are among its most significant risk factors. Abnormal iron metabolism is associated with the progression of cancer malignancy. Nevertheless, the involvement of iron metabolism in the EC remains uncertain. Ceruloplasmin (CP) functions as a multicopper oxidase and ferroxidase, playing a crucial role in maintaining the metabolic balance between copper and iron. Prior research has demonstrated that the dysregulated expression of CP has important clinical implications in EC. However, ​the specific underlying molecular mechanisms remains uncertain. This research examined the impact of CP on the malignant advancement of EC by suppressing ferroptosis. Next, we explored the possibility that Long non-coding RNA (lncRNA) LINC02936/SIX1/CP axis may be a key pathway for inhibiting ferroptosis and promoting cancer progression in EC. Mechanistically, SIX1 modulates the expression of CP, whereas LINC02936 interacts with SIX1 and recruits SIX1 to the CP promoter, leading to upregulation of CP, inhibition of ferroptosis, and promotion of EC progression. Administration of a small peptide cloud block the LINC02936-SIX1 interaction, thereby inhibits EC progression by promoting ferroptosis. Altogether, this is the first report on the lncRNA regulation of ferroptosis in EC. Our research enhances the knowledge of the lncRNA-mediated regulation of ferroptosis in EC progression and indicates the potential therapeutic significance of the LINC02936/SIX1/CP axis in treating EC.

GSK3ß phosphorylates Six1 transcription factor and regulates its APC/CCdh1 mediated proteosomal degradation.

Sine oculis homeobox homolog 1 (Six1) is a developmentally important transcription factor that regulates cellular proliferation, apoptosis, and dissemination during embryogenesis. Six1 overexpression as reported in multiple cancers modulates expression of a repertoire of its target genes causing an increase in proliferation, metastasis and survival of cancer cells. Six1 exists as a cell cycle regulated nuclear phosphoprotein and its cellular turnover is regulated by APC/C (Anaphase promoting complex / Cyclosome) complex mediated proteolysis. However, the kinases that regulate Six1 proteolysis have not been identified and the mechanistic details that cause its overproduction in various cancers are lacking. Here, we report that Six1 is a physiological GSK3ß substrate. GSK3ß interacts with Six1 and phosphorylates it at Ser221 within the conserved consensus sequence in its carboxy terminus. Using pharmacological inhibition, siRNA mediated knockdown and protein overexpression of GSK3ß; we show that GSK3ß regulates Six1 protein stability. Pulse chase analysis of Six1 revealed that GSK3ß regulates its ubiquitin proteolysis such that Six1 phosphomimicking mutant (Six1S221E) for Ser221 site had dramatically increased half-life than its phosphodeficient (Six1S221A) and wild type variants. Furthermore, we demonstrate that GSK3ß rescues Six1 from APC dependent proteolysis by regulating its binding with APC/C co-activator protein Cdh1. Importantly, strong positive correlation exists between GSK3ß and Six1 protein levels throughout the cell cycle and in multiple cancers indicating that GSK3ß activation may in part contribute to Six1 overproduction in a subset of human cancers.

Cytoplasmatic Localization of Six1 in Male Testis and Spermatogonial Stem Cells.

Sine oculis homeobox 1 (Six1) is an important factor for embryonic development and carcinoma malignancy. However, the localization of Six1 varies due to protein size and cell types in different organs. In this study, we focus on the expression and localization of Six1 in male reproductive organ via bioinformatics analysis and immunofluorescent detection. The potential interacted proteins with Six1 were also predicted by protein-protein interactions (PPIs) and Enrichr analysis. Bioinformatic data from The Cancer Genome Atlas and Genotype-Tissue Expression project databases showed that SIX1 was highly expressed in normal human testis, but low expressed in the testicular germ cell tumor sample. Human Protein Atlas examination verified that SIX1 level was higher in normal than that in cancer samples. The sub-localization of SIX1 in different reproductive tissues varies but specifically in the cytoplasm and membrane in testicular cells. In mouse cells, single cell RNA-sequencing data analysis indicated that Six1 expression level was higher in mouse spermatogonial stem cells (mSSCs) and differentiating spermatogonial than in other somatic cells. Immunofluorescence staining showed the cytoplasmic localization of Six1 in mouse testis and mSSCs. Further PPIs and Enrichr examination showed the potential interaction of Six1 with bone morphogenetic protein 4 (Bmp4) and catenin Beta-1 (CtnnB1) and stem cell signal pathways. Cytoplasmic localization of Six1 in male testis and mSSCs was probably associated with stem cell related proteins Bmp4 and CtnnB1 for stem cell development.

BCLAF1 drives esophageal squamous cell carcinoma progression through regulation of YTHDF2-dependent SIX1 mRNA degradation.

Esophageal cancer ranks among the most prevalent malignant tumors, and esophageal squamous cell carcinoma (ESCC) constitutes its predominant histological form. Despite its impact, a thorough insight into the molecular intricacies of ESCC's development is still incomplete, which hampers the advancement of targeted molecular diagnostics and treatments. Recently, B-cell lymphoma-2-associated transcription factor 1 (BCLAF1) has come under investigation for its potential involvement in tumor biology, yet its specific role and mechanism in ESCC remain unclear. In this study, we observed a marked increase in BCLAF1 expression in ESCC tissues, correlating with advanced tumor stages and inferior patient outcomes. Our comprehensive in vitro and in vivo studies show that BCLAF1 augments glycolytic activity and the proliferation, invasion, and spread of ESCC cells. By employing mass spectrometry, we identified YTHDF2 as a key protein interacting with BCLAF1 in ESCC, with further validation provided by colocalization, co-immunoprecipitation, and GST pull-down assay. Further investigations involving MeRIP-seq and RIP-seq, alongside transcriptomic analysis, highlighted SIX1 mRNA as a molecule significantly upregulated and modified by N6-methyladenosine (m6A) in BCLAF1 overexpressing cells. BCLAF1 was found to reduce the tumor-suppressive activities of YTHDF2, and its effects on promoting glycolysis and cancer progression were shown to hinge on SIX1 expression. This research establishes that BCLAF1 fosters glycolysis and tumor progression in ESCC through the YTHDF2-SIX1 pathway in an m6A-specific manner, suggesting a potential target for future therapeutic intervention.

Targeting sine oculis homeoprotein 1 (SIX1): A review of oncogenic roles and potential natural product therapeutics.

Sine oculis homeoprotein 1 (SIX1), a prominent representative of the homeodomain transcription factors within the SIX family, has attracted significant interest owing to its role in tumorigenesis, cancer progression, and prognostic assessments. Initially recognized for its pivotal role in embryonic development, SIX1 has emerged as a resurgent factor across a diverse set of mammalian cancers. Over the past two decades, numerous investigations have emphasized SIX1's dual significance as a developmental regulator and central player in oncogenic processes. A mounting body of evidence links SIX1 to the initiation of diverse cancers, encompassing enhanced cellular metabolism and advancement. This review provides an overview of the multifaceted roles of SIX1 in both normal development and oncogenic processes, emphasizing its importance as a possible therapeutic target and prognostic marker. Additionally, this review discusses the natural product agents that inhibit various pro-oncogenic mechanisms associated with SIX1.

Long non-coding RNA SIX1-1 promotes proliferation of cervical cancer cells via negative transcriptional regulation of RASD1.

Cervical cancer poses a significant health burden for women globally, and the rapid proliferation of cervical cancer cells greatly worsens patient prognosis. Long non-coding RNAs (lncRNAs) play a crucial role in regulating tumor cell proliferation. However, the involvement of lncRNAs in cervical cancer cell proliferation remains unclear. In this study, we investigated the lncRNA SIX1-1, which was found to be upregulated in cervical cancer tissues and cell lines. Functional assays revealed that knockdown of SIX1-1 inhibited cell proliferation in vitro and reduced tumor growth in vivo. Mechanistically, SIX1-1 was predominantly localized in the nucleus and could bind with DNMT1 protein. The expression of SIX1-1 enhanced the interaction of DNMT1 with RASD1 promoter, leading to the methylation of the promoter and decreased mRNA transcription. Then RASD1 downregulation activated the cAMP/PKA/CREB signaling pathway, promoting cell proliferation. Rescue experiments showed that knockdown of RASD1 restored the inhibited cell proliferation caused by decreased expression of SIX1-1, indicating that RASD1 acted as the functional mediator of SIX1-1. In conclusion, SIX1-1 promoted cervical cancer cell proliferation by modulating RASD1 expression. This suggests that targeting the SIX1-1/RASD1 axis could be a potential antitumor strategy for cervical cancer.

Thymic Carcinoma: Unraveling Neuroendocrine Differentiation and Epithelial Cell Identity Loss.

BACKGROUND: The histogenesis of thymic epithelial tumors (TETs) has been a subject of debate. Recent technological advancements have revealed that thymic carcinomas often exhibit a phenotype akin to tuft cells, which is a subset of medullary TECs. Here, we further explored the gene expression signatures of thymic carcinomas in relation to tuft cells and their kinships-ionocytes and neuroendocrine cells (neuroendocrine group). METHODS: We analyzed a single-cell RNA sequencing dataset from the normal human thymus. Concurrently, we examined publicly available datasets on the mRNA expression and methylation status of TECs and lung cancers. Real-time quantitative PCR was also conducted with our tissue samples. RESULTS: Thymic carcinomas displayed a neuroendocrine phenotype biased toward tuft cells and ionocytes. When exploring the possible regulators of this phenotype, we discovered that HDAC9 and NFATC1 were characteristically expressed in the neuroendocrine group in adult TECs and thymic carcinomas. Additionally, the pan-thymic epithelium markers, exemplified by PAX9 and SIX1, were significantly suppressed in thymic carcinomas. CONCLUSIONS: Thymic carcinomas might be characterized by unique neuroendocrine differentiation and loss of identity as thymic epithelial cells. Future studies investigating the role of HDAC9 and NFATC1 in thymic epithelium are warranted to explore their potential as therapeutic targets in TETs.