Mutation Research-Fundamental and Molecular Mechanisms of Mutagenesis最新文献_第2页

Repeated artificial mutagenesis of Bradyrhizobium diazoefficiens by gamma irradiation accelerates the acquisition of high-temperature tolerance 伽玛辐照对缓生重氮根瘤菌的多次人工诱变加速了其耐高温性的获得。

IF 1.9 4区医学 Q4 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Mutation Research-Fundamental and Molecular Mechanisms of Mutagenesis

Pub Date : 2025-07-01 DOI: 10.1016/j.mrfmmm.2025.111919

Yoshihiro Hase, Ikuko Nagafune, Katsuya Satoh

Effective mutant screening is critical for improving industrial microorganisms. This study was conducted to evaluate the effect of repeated mutagenesis with gamma rays on the experimental evolution of rhizobial high-temperature tolerance. Wild-type Bradyrhizobium diazoefficiens USDA110 cells grow optimally at 32−34 °C, but their growth is markedly retarded at 36 °C. Wild-type cells were subcultured in a 96-well deep-well plate for 76 or 83 days, with a gradual increase in temperature from 34.0 to 37.0 °C. Additionally, they were exposed to gamma radiation (1–120 Gy, 10 times in total) during the experimental period. The 40-Gy and 80-Gy treatments generated the most lines with high-temperature-tolerance. However, after extended subculturing without mutagenesis, tolerant lines obtained following the 80-Gy treatment produced smaller colonies than tolerant lines obtained after the 40-Gy treatment, suggesting the accumulation of deleterious mutations. These results imply that approximately 40 Gy is the appropriate dose for accumulating beneficial mutations under our experimental conditions. The two most tolerant lines obtained via the 30-Gy treatment commonly had a mutation in the 16S ribosomal RNA gene and the DNA-directed RNA polymerase subunit beta′ gene (rpoC), possibly reflecting a strong relationship with high-temperature tolerance. The optimal mutagenesis conditions for accumulating beneficial mutations were discussed based on the number of induced mutations in the population.

有效的突变体筛选是改善工业微生物的关键。本研究旨在评价伽玛射线反复诱变对根瘤菌耐高温性实验进化的影响。野生型重氮缓生根瘤菌USDA110细胞在32-34℃条件下生长最佳，但在36℃条件下生长明显迟缓。野生型细胞在96孔深孔板中传代76或83天，温度从34.0℃逐渐升高到37.0℃。此外，在实验期间，他们被暴露于γ辐射（1-120 Gy，共10次）。40 gy和80 gy处理产生的耐高温品系最多。然而，在没有诱变的情况下进行长时间传代培养后，80 gy处理后获得的耐受性系产生的菌落比40 gy处理后获得的耐受性系小，这表明有害突变的积累。这些结果表明，在我们的实验条件下，大约40 Gy是积累有益突变的适当剂量。通过30-Gy处理获得的两个最耐受性的品系通常在16S核糖体RNA基因和dna定向RNA聚合酶亚基β '基因（rpoC）中发生突变，可能反映了与高温耐受性的强烈关系。根据群体中诱导突变的数量，讨论了积累有益突变的最佳诱变条件。

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引用次数: 0

Corrigendum to “Saikosaponin-d Mediates FOXG1 to Reverse Docetaxel Resistance in Prostate Cancer through Oxidative Phosphorylation” [Mut. Res. - Fundam. Mol. Mech. Mutagenesis 829 (2024) 111875] “Saikosaponin-d通过氧化磷酸化介导FOXG1逆转前列腺癌多西紫杉醇耐药”的更正。请回答- Fundam。摩尔。机械。诱变829(2024)111875]。

IF 1.9 4区医学 Q4 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Mutation Research-Fundamental and Molecular Mechanisms of Mutagenesis

Pub Date : 2025-07-01 DOI: 10.1016/j.mrfmmm.2025.111914

Jun Meng, Bo Yang, Chang Shu, Shuai Jiang

引用次数: 0

Hypoxia-induced HIF-1α enhanced the tumorigenesis in non-small cell lung cancer by targeting GAB2 缺氧诱导的HIF-1α通过靶向GAB2促进非小细胞肺癌的肿瘤发生

IF 1.9 4区医学 Q4 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Mutation Research-Fundamental and Molecular Mechanisms of Mutagenesis

Pub Date : 2025-07-01 DOI: 10.1016/j.mrfmmm.2025.111917

Xunxia Zhu , Xiaoyong Shen , Xiaoyu Chen, Xuelin Zhang, Wen Gao

Non-small cell lung cancer (NSCLC) is a lethal disease with high morbidity and mortality rates. HIF-1α is confirmed to be involved in NSCLC. However, the detailed mechanism of its role remains unclear. NCI-H226 and SK-MES-1 cell lines were used to explore the mechanisms by which hypoxia affects the progression of NSCLC in vitro. The cellular functions were detected by transwell. The expressions of key biomarkers were examined by Real-time quantitative reverse transcription PCR (qRT-PCR) and Western Blot assays. The RNA sequencing analysis was used to explore the downstream targets of HIF-1α. Luciferase and Chromatin immunoprecipitation (ChIP) assays confirmed the interaction between HIF-1α and GAB2. What’s more, the xenograft model was used to investigate the effect of GAB2 in vivo. Hypoxia promoted the migration and invasion capabilities of NCI-H226 and SK-MES-1 cells. RNA sequencing analysis revealed that the expression of GAB2 is dramatically altered under a hypoxic environment. The bioinformatics analysis implied that the differentially expressed genes (DEGs) were enriched in the MEK/ERK signaling pathway and the significantly expressed GAB2 was associated with HIF-1α. Functionally, GAB2 regulated migration and invasion capabilities in vitro and facilitated tumor growth and lung metastasis of NSCLC in vivo. What’s more, luciferase and ChIP assays further demonstrated that HIF-1α could bind to GAB2. Hypoxia-induced HIF-1α enhanced tumor growth and lung metastasis in NSCLC by targeting GAB2, which might provide novel insights into GAB2 as a potential therapeutic target for NSCLC.

非小细胞肺癌（NSCLC）是一种发病率和死亡率都很高的致命疾病。HIF-1α被证实参与NSCLC。然而，其作用的具体机制尚不清楚。我们利用NCI-H226和SK-MES-1细胞系，探讨缺氧在体外影响NSCLC进展的机制。transwell检测细胞功能。采用实时定量反转录PCR （qRT-PCR）和Western Blot检测关键生物标志物的表达。RNA测序分析用于探索HIF-1α的下游靶点。荧光素酶和染色质免疫沉淀（ChIP）实验证实了HIF-1α和GAB2之间的相互作用。此外，采用异种移植物模型研究GAB2在体内的作用。缺氧可促进NCI-H226和SK-MES-1细胞的迁移和侵袭能力。RNA测序分析显示，在缺氧环境下，GAB2的表达发生了显著变化。生物信息学分析提示MEK/ERK信号通路中差异表达基因（DEGs）富集，GAB2显著表达与HIF-1α相关。在功能上，GAB2调节体外迁移和侵袭能力，促进体内NSCLC的肿瘤生长和肺转移。荧光素酶和ChIP实验进一步证实HIF-1α可以与GAB2结合。缺氧诱导的HIF-1α通过靶向GAB2促进非小细胞肺癌的肿瘤生长和肺转移，这可能为GAB2作为非小细胞肺癌的潜在治疗靶点提供新的见解。

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引用次数: 0

CPMFD: An algorithm for Classification of Point Mutations together with Frameshift Determination in related mRNA sequences CPMFD：一种在相关mRNA序列中进行点突变分类和移码测定的算法。

IF 1.9 4区医学 Q4 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Mutation Research-Fundamental and Molecular Mechanisms of Mutagenesis

Pub Date : 2025-07-01 DOI: 10.1016/j.mrfmmm.2025.111918

Probir Mondal , Pratyay Banerjee , Krishnendu Basuli

Mutations are responsible for the genetic origin of various diseases. Existing techniques for mutation identification often fails to detect the full spectrum of mutations in complex genomes hindering progress in diagnosis, treatment and prevention of diseases. Here we propose an algorithm to identify the location and type of mutation occurring in a mutated string with respect to a reference mRNA sequence. In addition to identifying insertion and deletion, by constructing suitable rational combinations of the prime numbers, our algorithm is able to classify point mutations in a novel way by distinguishing missense mutation from silent mutation. Amino acid transformation at each missense mutation site is identified. Moreover, the method allows to locate regions in the sequence undergoing frameshift. It turns out to be efficient when applied on sample dataset. Application of this framework to two haplotypes of the Plasmodium falciparum datasets exhibits different mutation profile to develop similar chloroquine resistance. Despite the overwhelming similarity between the

β

-globin genes of pygmy and common chimpanzees, our algorithm is able to pinpoint the minute details of the mutations occurring in them differentiating the two species. Additionally, in Alzheimer datasets, the method meticulously identifies true variations in related genes.

突变是各种疾病的遗传起源的原因。现有的突变鉴定技术往往不能检测复杂基因组中的全部突变，阻碍了疾病诊断、治疗和预防的进展。在这里，我们提出了一种算法来识别相对于参考mRNA序列在突变字符串中发生的突变的位置和类型。除了识别插入和删除外，通过构建合适的素数有理组合，我们的算法能够通过区分错义突变和沉默突变，以一种新的方式对点突变进行分类。鉴定了每个错义突变位点的氨基酸转化。此外，该方法允许定位序列中发生移码的区域。当应用于样本数据集时，结果证明该方法是有效的。将这一框架应用于恶性疟原虫的两种单倍型数据集，显示出不同的突变谱，从而产生相似的氯喹耐药性。尽管侏儒黑猩猩和普通黑猩猩的β-珠蛋白基因非常相似，但我们的算法能够精确地指出它们身上发生的区分两个物种的突变的微小细节。此外，在阿尔茨海默病的数据集中，该方法一丝不苟地识别相关基因的真实变异。

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引用次数: 0

Analysis of TERT promoter hotspot mutations using droplet digital PCR in hepatoblastoma and hepatocellular carcinoma 肝母细胞瘤和肝细胞癌中TERT启动子热点突变的微滴数字PCR分析

IF 1.9 4区医学 Q4 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Mutation Research-Fundamental and Molecular Mechanisms of Mutagenesis

Pub Date : 2025-07-01 DOI: 10.1016/j.mrfmmm.2025.111915

Yoko Hiyama , Masato Kojima , Sho Kurihara , Isamu Saeki , Ryo Touge , Takahiro Fukazawa , Takanori Harada , Eiso Hiyama

Somatic mutations in the telomerase reverse transcriptase promoter (TERTp) region are common in many cancers, including in liver cancers. Detection of TERTp mutations in tumor tissue DNAs and cell-free tumor DNAs is useful for diagnosing and monitoring cancers. Since the most common TERTp hotspot mutations, C228T and C250T, are difficult to identify using Sanger sequencing, we tested an easy and highly sensitive alternative method that targets these two sites using droplet digital PCR. Using this method, both the sensitivity and specificity for detecting these two mutations were 100 % in DNA samples derived from cell lines and liver cancer tissues, including hepatocellular carcinoma (HCC) and hepatoblastoma (HB). The detection limit for the allele frequencies of these mutations was approximately 0.1 %. This method is also widely applicable; for instance, it can be applied to DNA derived from FFPE (formalin-fixed paraffin embedded) samples. In addition, we applied this method to detecting TERTp mutations in cell-free DNA samples of patients with TERTp-mutated tumors. Finally, we found that outcomes for HB patients with TERTp mutations were significantly worse than in those without mutations, indicating the importance of this method for improving patient outcomes. In light of this, we discuss the advantages of this method for clinical implementation in the detection and monitoring of cancers.

端粒酶逆转录酶启动子（TERTp）区域的体细胞突变在许多癌症中都很常见，包括肝癌。检测肿瘤组织dna和无细胞肿瘤dna中的TERTp突变对癌症的诊断和监测是有用的。由于最常见的TERTp热点突变C228T和C250T难以用Sanger测序识别，我们测试了一种简单且高灵敏度的替代方法，使用液滴数字PCR靶向这两个位点。使用该方法，在细胞系和肝癌组织（包括肝细胞癌（HCC）和肝母细胞瘤（HB））的DNA样本中检测这两种突变的灵敏度和特异性均为100 %。这些突变的等位基因频率检测限约为0.1 %。这种方法也广泛适用；例如，它可以应用于来自FFPE（福尔马林固定石蜡包埋）样品的DNA。此外，我们将该方法应用于TERTp突变肿瘤患者的无细胞DNA样本中TERTp突变的检测。最后，我们发现TERTp突变的HB患者的预后明显差于无突变的HB患者，表明该方法对改善患者预后的重要性。鉴于此，我们讨论了该方法在癌症检测和监测中的临床实施优势。

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引用次数: 0

A novel mechanism in driving non-small cell lung cancer progression: The METTL3/FOXA1/PTK2 cascade 驱动非小细胞肺癌进展的新机制：METTL3/FOXA1/PTK2级联

IF 1.5 4区医学 Q4 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Mutation Research-Fundamental and Molecular Mechanisms of Mutagenesis

Pub Date : 2025-06-11 DOI: 10.1016/j.mrfmmm.2025.111911

Xuelin Zhang , Yizhao Chen , Lingjie Wang , Qingyue Lin , Tingjian Li , Chunya He

Background

Dysregulation of m6A modification has significant implications in human carcinogenesis. METTL3, a crucial m6A writer, acts as an oncogenic driver in non-small cell lung cancer (NSCLC). Here, we explored its mechanisms in driving NSCLC development.

Methods

Cell sphere formation, invasion, apoptosis, and proliferation were detected by sphere formation, transwell, flow cytometry, and MTT assays, respectively. Cell glycolysis was evaluated by measuring glucose consumption, lactate production, and ATP/ADP ratio. RIP, methylated RIP (MeRIP), and mRNA stability assays were used to analyze the METTL3/FOXA1 relationship. Luciferase assay and ChIP experiment were used for the evaluation of the FOXA1/PTK2 relationship. Xenograft studies were used to test the role in vivo.

Results

METTL3 was upregulated in NSCLC, and its inhibition diminished the growth, invasiveness, sphere formation ability, and glycolysis of H1299 and A549 cells. Mechanistically, METTL3 depletion caused a reduction in FOXA1 expression through the m6A modification mechanism. FOXA1 transcriptionally controlled PTK2 expression. FOXA1 upregulation reversed the effects of METTL3 inhibition on the growth, invasiveness, sphere formation ability, and glycolysis of H1299 and A549 cells. Moreover, FOXA1 increase attenuated the impact of METTL3 inhibition on the in vivo growth of A549 subcutaneous xenografts. Additionally, increased PTK2 expression counteracted the effects of FOXA1 reduction on the malignant phenotypes of H1299 and A549 cells.

Conclusion

Our finding elucidates a novel mechanism for METTL3’s oncogenic activity in NSCLC, where METTL3 upregulates FOXA1 and thus activates PTK2 transcription. Blocking this cascade may be effective for combating NSCLC.

m6A修饰的失调在人类癌变中具有重要意义。METTL3是一个重要的m6A转录因子，在非小细胞肺癌（NSCLC）中起着致癌驱动作用。在此，我们探讨了其在推动非小细胞肺癌发展中的机制。方法分别采用成球法、transwell法、流式细胞术和MTT法检测细胞的成球、侵袭、凋亡和增殖。通过测量葡萄糖消耗、乳酸生成和ATP/ADP比值来评估细胞糖酵解。使用RIP、甲基化RIP （MeRIP）和mRNA稳定性分析METTL3/FOXA1的关系。采用荧光素酶法和ChIP实验评价FOXA1/PTK2的关系。异种移植研究被用来测试在体内的作用。结果mettl3在非小细胞肺癌中表达上调，其抑制作用降低H1299和A549细胞的生长、侵袭性、成球能力和糖酵解能力。从机制上讲，METTL3缺失通过m6A修饰机制导致FOXA1表达降低。FOXA1转录控制PTK2的表达。FOXA1上调逆转了METTL3抑制对H1299和A549细胞生长、侵袭性、成球能力和糖酵解的影响。此外，FOXA1的增加减弱了METTL3抑制对A549皮下异种移植物体内生长的影响。此外，PTK2表达的增加抵消了FOXA1减少对H1299和A549细胞恶性表型的影响。我们的发现阐明了METTL3在NSCLC中致癌活性的新机制，其中METTL3上调FOXA1，从而激活PTK2转录。阻断这种级联可能对对抗非小细胞肺癌有效。

{"title":"A novel mechanism in driving non-small cell lung cancer progression: The METTL3/FOXA1/PTK2 cascade","authors":"Xuelin Zhang , Yizhao Chen , Lingjie Wang , Qingyue Lin , Tingjian Li , Chunya He","doi":"10.1016/j.mrfmmm.2025.111911","DOIUrl":"10.1016/j.mrfmmm.2025.111911","url":null,"abstract":"<div><h3>Background</h3><div>Dysregulation of m6A modification has significant implications in human carcinogenesis. METTL3, a crucial m6A writer, acts as an oncogenic driver in non-small cell lung cancer (NSCLC). Here, we explored its mechanisms in driving NSCLC development.</div></div><div><h3>Methods</h3><div>Cell sphere formation, invasion, apoptosis, and proliferation were detected by sphere formation, transwell, flow cytometry, and MTT assays, respectively. Cell glycolysis was evaluated by measuring glucose consumption, lactate production, and ATP/ADP ratio. RIP, methylated RIP (MeRIP), and mRNA stability assays were used to analyze the METTL3/FOXA1 relationship. Luciferase assay and ChIP experiment were used for the evaluation of the FOXA1/PTK2 relationship. Xenograft studies were used to test the role <em>in vivo</em>.</div></div><div><h3>Results</h3><div>METTL3 was upregulated in NSCLC, and its inhibition diminished the growth, invasiveness, sphere formation ability, and glycolysis of H1299 and A549 cells. Mechanistically, METTL3 depletion caused a reduction in FOXA1 expression through the m6A modification mechanism. FOXA1 transcriptionally controlled PTK2 expression. FOXA1 upregulation reversed the effects of METTL3 inhibition on the growth, invasiveness, sphere formation ability, and glycolysis of H1299 and A549 cells. Moreover, FOXA1 increase attenuated the impact of METTL3 inhibition on the <em>in vivo</em> growth of A549 subcutaneous xenografts. Additionally, increased PTK2 expression counteracted the effects of FOXA1 reduction on the malignant phenotypes of H1299 and A549 cells.</div></div><div><h3>Conclusion</h3><div>Our finding elucidates a novel mechanism for METTL3’s oncogenic activity in NSCLC, where METTL3 upregulates FOXA1 and thus activates PTK2 transcription. Blocking this cascade may be effective for combating NSCLC.</div></div>","PeriodicalId":49790,"journal":{"name":"Mutation Research-Fundamental and Molecular Mechanisms of Mutagenesis","volume":"831 ","pages":"Article 111911"},"PeriodicalIF":1.5,"publicationDate":"2025-06-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144312897","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

METTL14-mediated m6A modification of ETV4 inhibits tumor development in colorectal cancer mettl14介导的m6A修饰ETV4抑制结直肠癌的肿瘤发展

IF 1.5 4区医学 Q4 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Mutation Research-Fundamental and Molecular Mechanisms of Mutagenesis

Pub Date : 2025-06-11 DOI: 10.1016/j.mrfmmm.2025.111910

Xiaofeng Liao, Tao Hu

Background

Many m6A methyltransferases have been identified to regulate colorectal cancer (CRC) progression. METTL14 has been confirmed to play a negative role in CRC process, but the molecular mechanism of METTL14 in regulating CRC progression needs to be further elucidated.

Methods

The levels of METTL14, YTHDF2 and ETS translocation variant 4 (ETV4) were examined by qRT-PCR and western blot. Cell proliferation and apoptosis were determined by colony formation assay and flow cytometry. Cell glycolysis was assessed by detecting corresponding indicators. Cell ferroptosis was evaluated via measuring SOD, MDA, GSH, ROS and Fe^2 + levels. The interaction between ETV4 and METTL14 or m6A readers was confirmed by RIP assay and RNA pull-down assay. Animal experiments were performed to confirm METTL14 roles in vivo.

Results

METTL14 was downregulated in CRC tissues and cells, which overexpression inhibited proliferation and glycolysis, as well as promoted apoptosis and ferroptosis in CRC cells. METTL14 reduced the mRNA stability of ETV4 and inhibited ETV4 protein expression through m6A modification. m6A reader YTHDF2 could recognize m6A-methylated ETV4. The downregulation of ETV4 by METTL14 leads to increased apoptosis and ferroptosis in CRC cells, suggesting a critical role in tumor suppression. Moreover, METTL14 inhibited CRC tumorigenesis in vivo via reducing ETV4 expression.

Conclusion

METTL14 accelerated CRC cell apoptosis and ferroptosis via downregulating ETV4 in m6A-dependent manner, providing a molecular target for CRC treatment.

许多m6A甲基转移酶已被确定调节结直肠癌（CRC）的进展。METTL14已被证实在CRC过程中发挥负向作用，但其调控CRC进展的分子机制有待进一步阐明。方法采用qRT-PCR和western blot检测各组小鼠METTL14、YTHDF2和ETS易位变异体4 （ETV4）的表达水平。采用集落形成实验和流式细胞术检测细胞增殖和凋亡情况。通过检测相应指标评价细胞糖酵解。通过测定SOD、MDA、GSH、ROS、Fe2 +水平评价细胞铁下垂。通过RIP实验和RNA下拉实验证实了ETV4与METTL14或m6A读取器的相互作用。动物实验证实了METTL14在体内的作用。结果mettl14在结直肠癌组织和细胞中表达下调，其过表达抑制结直肠癌细胞增殖和糖酵解，促进结直肠癌细胞凋亡和铁凋亡。METTL14通过m6A修饰降低ETV4 mRNA的稳定性，抑制ETV4蛋白的表达。m6A读卡器YTHDF2可以识别m6A甲基化的ETV4。METTL14下调ETV4导致CRC细胞凋亡和铁凋亡增加，提示其在肿瘤抑制中起关键作用。此外，METTL14通过降低ETV4的表达在体内抑制CRC的肿瘤发生。结论mettl14通过下调ETV4以m6a依赖的方式加速结直肠癌细胞凋亡和铁凋亡，为结直肠癌治疗提供了分子靶点。

{"title":"METTL14-mediated m6A modification of ETV4 inhibits tumor development in colorectal cancer","authors":"Xiaofeng Liao, Tao Hu","doi":"10.1016/j.mrfmmm.2025.111910","DOIUrl":"10.1016/j.mrfmmm.2025.111910","url":null,"abstract":"<div><h3>Background</h3><div>Many m6A methyltransferases have been identified to regulate colorectal cancer (CRC) progression. METTL14 has been confirmed to play a negative role in CRC process, but the molecular mechanism of METTL14 in regulating CRC progression needs to be further elucidated.</div></div><div><h3>Methods</h3><div>The levels of METTL14, YTHDF2 and ETS translocation variant 4 (ETV4) were examined by qRT-PCR and western blot. Cell proliferation and apoptosis were determined by colony formation assay and flow cytometry. Cell glycolysis was assessed by detecting corresponding indicators. Cell ferroptosis was evaluated via measuring SOD, MDA, GSH, ROS and Fe<sup>2 +</sup> levels. The interaction between ETV4 and METTL14 or m6A readers was confirmed by RIP assay and RNA pull-down assay. Animal experiments were performed to confirm METTL14 roles <em>in vivo</em>.</div></div><div><h3>Results</h3><div>METTL14 was downregulated in CRC tissues and cells, which overexpression inhibited proliferation and glycolysis, as well as promoted apoptosis and ferroptosis in CRC cells. METTL14 reduced the mRNA stability of ETV4 and inhibited ETV4 protein expression through m6A modification. m6A reader YTHDF2 could recognize m6A-methylated ETV4. The downregulation of ETV4 by METTL14 leads to increased apoptosis and ferroptosis in CRC cells, suggesting a critical role in tumor suppression. Moreover, METTL14 inhibited CRC tumorigenesis <em>in vivo</em> via reducing ETV4 expression.</div></div><div><h3>Conclusion</h3><div>METTL14 accelerated CRC cell apoptosis and ferroptosis via downregulating ETV4 in m6A-dependent manner, providing a molecular target for CRC treatment.</div></div>","PeriodicalId":49790,"journal":{"name":"Mutation Research-Fundamental and Molecular Mechanisms of Mutagenesis","volume":"831 ","pages":"Article 111910"},"PeriodicalIF":1.5,"publicationDate":"2025-06-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144321574","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

USP7 accelerates colorectal cancer progression by up-regulating MYO6 through deubiquitination USP7通过去泛素化上调MYO6，从而加速结直肠癌的进展

IF 1.5 4区医学 Q4 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Mutation Research-Fundamental and Molecular Mechanisms of Mutagenesis

Pub Date : 2025-05-17 DOI: 10.1016/j.mrfmmm.2025.111908

Ming Jiang , Chong Xiong

Background

Ubiquitin-specific protease 7 (USP7) is one of deubiquitinases and has been reported to regulate cancer cell biological processes through removing ubiquitin modifications from protein substrates. Myosins of class VI (MYO6) is shown to be highly expressed in many of cancers, and is associated with tumor progression in several cancers by affecting cell survival. Moreover, USP7 and MYO6 have been revealed to be involved in colorectal cancer (CRC) progression. Here, we aimed to investigate the intricate interplay between MYO6 and USP7 in CRC, and whether their interaction was associated with deubiquitination.

Methods

Quantitative real-time PCR and western blot were used to for mRNA and protein detection. Functional analyses were conducted using Cell Counting Kit-8, 5-ethynyl-2-deoxy-uridine, flow cytometry, wound healing and transwell assays in vitro, and murine xenograft models in vivo. M2 macrophage polarization was determined with CD206 antibody using flow cytometry. The protein interaction between MYO6 and USP7 was determined by chromatin immunoprecipitation assay. The deubiquitination effect of USP7 was validated by cellular ubiquitination and immunoprecipitation assay.

Results

CRC tissues and cells showed high expression of MYO6. Functionally, silencing of MYO6 suppressed CRC cell proliferation, migration, invasion, angiogenesis, induced cell apoptosis and negatively affected macrophage M2 polarization in vitro, and impeded CRC growth in vivo. For a mechanism analysis, USP7 could stabilize and up-regulate MYO6 expression by inducing MYO6 deubiquitination. USP7 was also highly expressed in CRC, USP7 silencing repressed CRC cell malignant phenotypes and reduced macrophage M2 polarization, while these effects were reversed by MYO6 overexpression.

Conclusion

MYO6 promoted CRC cell tumorigenesis and macrophage M2 polarization, and the mechanism was associated with USP7-induced MYO6 deubiquitination. These results suggested new targets for the development of epigenetic-based therapy in CRC.

泛素特异性蛋白酶7 （USP7）是一种去泛素酶，据报道通过去除蛋白质底物上的泛素修饰来调节癌细胞的生物学过程。VI类肌球蛋白（MYO6）在许多癌症中高表达，并通过影响细胞存活与几种癌症的肿瘤进展相关。此外，USP7和MYO6已被发现参与结直肠癌（CRC）的进展。在这里，我们的目的是研究MYO6和USP7在CRC中复杂的相互作用，以及它们的相互作用是否与去泛素化有关。方法采用实时荧光定量PCR和western blot技术进行mRNA和蛋白的检测。使用细胞计数试剂盒- 8,5 -乙基-2-脱氧尿苷、流式细胞术、体外伤口愈合和transwell实验以及小鼠体内异种移植模型进行功能分析。用CD206抗体流式细胞术检测M2巨噬细胞极化。采用染色质免疫沉淀法测定MYO6与USP7之间的蛋白相互作用。通过细胞泛素化和免疫沉淀实验验证了USP7的去泛素化作用。结果scrc组织细胞MYO6高表达。在功能上，MYO6的沉默在体外抑制结直肠癌细胞的增殖、迁移、侵袭、血管生成，诱导细胞凋亡，并对巨噬细胞M2极化产生负面影响，在体内抑制结直肠癌的生长。机制分析表明，USP7可以通过诱导MYO6去泛素化来稳定和上调MYO6的表达。USP7在结直肠癌中也高表达，USP7沉默抑制结直肠癌细胞的恶性表型，减少巨噬细胞M2极化，而这些作用被MYO6过表达逆转。结论MYO6促进结直肠癌细胞的肿瘤发生和巨噬细胞M2极化，其机制与usp7诱导的MYO6去泛素化有关。这些结果为发展基于表观遗传学的结直肠癌治疗提供了新的靶点。

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引用次数: 0

In silico report on five high-risk protein C pathogenic variants: G403R, P405S, S421N, C238S, and I243T 五种高危蛋白C致病变异：G403R、P405S、S421N、C238S和I243T的计算机报告

IF 1.5 4区医学 Q4 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Mutation Research-Fundamental and Molecular Mechanisms of Mutagenesis

Pub Date : 2025-05-17 DOI: 10.1016/j.mrfmmm.2025.111907

Daniela Hristov , Done Stojanov

In this study, we propose reclassification of 5 out of 16 PROC VUS (variants of uncertain significance): C238S, I243T, G403R, P405S, and S421N, as pathogenic variants, associated with thrombophilia due to PROC deficiency. The obtained results are based on in silico analysis, which enables a detailed assessment of variants’ impact, despite limited clinical evidence. In particular, the G403R substitution, next to the S402-active site, is expected to reduce the flexibility of the local coil domain, affecting the catalytic activity of serine protease. The P405S substitution may imply B-factor gain (P = 0.24; p-value=0.040). On the other hand, the S421N variant causes phosphorylation site disruption at S421, which serves as a target for CK2 phosphorylation. C238S substitution alters metal binding, while the I243T variant may alter transmembrane properties (P = 0.27, P-value=0.00071). All five PROC variants hold promise as diagnostic markers for protein C deficiency and may also serve as potential drug targets for therapeutic intervention.

在这项研究中，我们提出将16个PROC VUS（意义不确定的变体）中的5个重新分类为致病变体，即C238S、I243T、G403R、P405S和S421N，这些变体与PROC缺乏导致的血栓形成相关。获得的结果基于计算机分析，尽管临床证据有限，但可以对变异的影响进行详细评估。特别是在s402活性位点旁边的G403R取代，预计会降低局部线圈结构域的柔韧性，影响丝氨酸蛋白酶的催化活性。P405S取代可能意味着b因子增益(P = 0.24；假定值= 0.040)。另一方面，S421N变异导致S421磷酸化位点破坏，S421作为CK2磷酸化的靶点。C238S取代改变金属结合，而I243T变体可能改变跨膜性质（P = 0.27，P值=0.00071）。所有五种PROC变体都有望作为蛋白C缺乏症的诊断标记物，也可能作为治疗干预的潜在药物靶点。

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引用次数: 0

SIRT6 regulates the HIPK2/P53 pathway to reduce oxidative stress and apoptosis to attenuate vancomycin-induced nephrotoxicity SIRT6调节HIPK2/P53通路，减少氧化应激和细胞凋亡，减轻万古霉素引起的肾毒性

IF 1.9 4区医学 Q4 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Mutation Research-Fundamental and Molecular Mechanisms of Mutagenesis

Pub Date : 2025-01-01 DOI: 10.1016/j.mrfmmm.2024.111897

Xiuying Feng , Yunhui Liu , Lei Su , Luyang Xu

SIRT6 is known to play a protective role in several kidney diseases; however, its role in vancomycin-induced renal injury remains unclear. This study aims to confirm the role and related mechanisms of SIRT6 in vancomycin-induced renal injury. To develop a kidney damage model, mice were given vancomycin injections for seven days. Additionally, an in vivo transfection with a SIRT6 overexpression plasmid was performed. PCR and Western blot analyses were used to assess the SIRT6 mRNA and protein expression levels in renal tissue. HE staining was performed to evaluate renal tissue damage, while Scr and BUN were measured using specialized kits. Renal tissue apoptotic cells were labeled using a TUNEL kit, and the levels of the antioxidant enzymes SOD and GSH were measured using appropriate kits. Western blot was used to identify HIPK2, p-p53, and p53 protein expression in the renal tissue. The results reveal that SIRT6 is expressed at markedly low levels in renal tissue. Furthermore, mice administered vancomycin exhibited a significant increase in Scr and BUN levels, indicating impaired renal function. Histological examination through HE staining demonstrated considerable damage to the renal tissue of the vancomycin group. Additionally, the renal tissue of the mice in the vancomycin group displayed reduced levels of the antioxidant enzymes SOD and GSH, an increased number of TUNEL-positive cells, and significantly elevated levels of HIPK2 and p-p53 protein expression. Moreover, the mice transfected with SIRT6 exhibited significant improvements in previously described symptoms. These findings imply that the inhibition of HIPK2/p53 by SIRT6 may represent a promising therapeutic strategy for alleviating vancomycin-induced nephrotoxicity.

已知SIRT6在几种肾脏疾病中发挥保护作用；然而，其在万古霉素引起的肾损伤中的作用尚不清楚。本研究旨在证实SIRT6在万古霉素致肾损伤中的作用及相关机制。为了建立肾脏损伤模型，小鼠被注射万古霉素7天。此外，用SIRT6过表达质粒进行体内转染。采用PCR和Western blot方法检测肾组织中SIRT6 mRNA和蛋白的表达水平。HE染色评估肾组织损伤，Scr和BUN用专用试剂盒测定。采用TUNEL试剂盒标记肾组织凋亡细胞，采用相应试剂盒检测抗氧化酶SOD和GSH水平。Western blot检测肾组织中HIPK2、p-p53和p53蛋白的表达。结果显示SIRT6在肾组织中表达水平明显较低。此外，给药万古霉素的小鼠Scr和BUN水平显著升高，表明肾功能受损。HE染色组织学检查显示万古霉素组肾组织损伤明显。此外，万古霉素组小鼠肾组织中抗氧化酶SOD和GSH水平降低，tunel阳性细胞数量增加，HIPK2和p-p53蛋白表达水平显著升高。此外，转染SIRT6的小鼠在先前描述的症状中表现出显着改善。这些发现表明SIRT6抑制HIPK2/p53可能是缓解万古霉素引起的肾毒性的一种有希望的治疗策略。

{"title":"SIRT6 regulates the HIPK2/P53 pathway to reduce oxidative stress and apoptosis to attenuate vancomycin-induced nephrotoxicity","authors":"Xiuying Feng , Yunhui Liu , Lei Su , Luyang Xu","doi":"10.1016/j.mrfmmm.2024.111897","DOIUrl":"10.1016/j.mrfmmm.2024.111897","url":null,"abstract":"<div><div><span><span><span>SIRT6 is known to play a protective role in several kidney diseases; however, its role in vancomycin-induced </span>renal injury<span> remains unclear. This study aims to confirm the role and related mechanisms of SIRT6 in vancomycin-induced renal injury. To develop a kidney damage model, mice were given </span></span>vancomycin<span> injections for seven days. Additionally, an in vivo transfection with a SIRT6 overexpression plasmid was performed. PCR and </span></span>Western blot analyses<span><span> were used to assess the SIRT6 mRNA and protein expression levels<span><span><span><span> in renal tissue. HE staining was performed to evaluate renal </span>tissue damage, while </span>Scr<span> and BUN<span> were measured using specialized kits. Renal tissue apoptotic cells were labeled using a TUNEL kit, and the levels of the antioxidant enzymes </span></span></span>SOD and GSH were measured using appropriate kits. Western blot was used to identify </span></span>HIPK2<span><span>, p-p53, and p53 protein expression<span> in the renal tissue. The results reveal that SIRT6 is expressed at markedly low levels in renal tissue. Furthermore, mice administered vancomycin<span> exhibited a significant increase in Scr and BUN levels, indicating impaired </span></span></span>renal function<span><span><span>. Histological examination through HE staining demonstrated considerable damage to the renal tissue of the vancomycin group. Additionally, the renal tissue of the mice in the vancomycin group displayed reduced levels of the antioxidant enzymes </span>SOD and GSH, an increased number of TUNEL-positive cells, and significantly elevated levels of </span>HIPK2<span> and p-p53 protein expression. Moreover, the mice transfected with SIRT6 exhibited significant improvements in previously described symptoms. These findings imply that the inhibition of HIPK2/p53 by SIRT6 may represent a promising therapeutic strategy for alleviating vancomycin-induced nephrotoxicity.</span></span></span></span></div></div>","PeriodicalId":49790,"journal":{"name":"Mutation Research-Fundamental and Molecular Mechanisms of Mutagenesis","volume":"831 ","pages":"Article 111897"},"PeriodicalIF":1.9,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143178151","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

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