【目的】克隆牦牛丝氨酸蛋白酶抑制剂家族E成员1(serpin family E member 1,SERPINE1)基因序列,同时探究不同嫩度牦牛背最长肌组织中SERPINE1基因核苷酸序列差异,为进一步研究其对牦牛肉嫩度的影响提供试验数据。【方法】根据GenBank中...【目的】克隆牦牛丝氨酸蛋白酶抑制剂家族E成员1(serpin family E member 1,SERPINE1)基因序列,同时探究不同嫩度牦牛背最长肌组织中SERPINE1基因核苷酸序列差异,为进一步研究其对牦牛肉嫩度的影响提供试验数据。【方法】根据GenBank中牦牛SERPINE1基因序列设计特异性引物,通过PCR扩增克隆获得高、低嫩度牦牛背最长肌中的SERPINE1基因序列全长,并对其结构及编码蛋白进行生物信息学分析,通过实时荧光定量PCR检测不同嫩度牦牛背最长肌中SERPINE1基因表达量。【结果】高、低嫩度牦牛背最长肌组织中SERPINE1基因全长分别为8 315和8 318 bp,均编码402个氨基酸且氨基酸序列完全相同。高、低嫩度背最长肌组织中SERPINE1基因非编码序列中存在3个碱基缺失及22个碱基突变(4个碱基颠换、18个碱基转换)。牦牛SERPINE1基因核苷酸序列与普通牛、瘤牛、美洲野牛、绵羊、山羊、家猪、人、小鼠的相似性分别为99.26%、99.26%、99.59%、96.94%、97.02%、90.49%、86.52%和80.10%。SERPINE1蛋白是一个含有23个氨基酸信号肽的亲水性稳定膜外蛋白,位于细胞外,具有34个潜在磷酸化位点,包含1个反应中心环(reactive center loop, RCL)。SERPINE1蛋白二级结构主要由α-螺旋(44.78%)和无规则卷曲(35.32%)构成。实时荧光定量PCR结果显示,SERPINE1基因在高嫩度牦牛背最长肌中表达量极显著高于低嫩度牦牛背最长肌(P<0.01)。【结论】本研究成功从高、低嫩度牦牛背最长肌组织中克隆SERPINE1基因全长并进行了生物信息学特征分析,为后续研究SERPINE1基因参与调控牦牛肉嫩度机制提供理论参考。展开更多
目的探讨丝氨酸蛋白酶抑制剂分支E成员1(serpin family E member 1,SERPINE1)的表达与胃癌的预后及肿瘤免疫细胞浸润的关系。方法通过肿瘤免疫估计资源(TIMER)数据库和TCGA中的胃癌数据,分析SERPINE1的表达。同时使用Kaplan-Meier plot...目的探讨丝氨酸蛋白酶抑制剂分支E成员1(serpin family E member 1,SERPINE1)的表达与胃癌的预后及肿瘤免疫细胞浸润的关系。方法通过肿瘤免疫估计资源(TIMER)数据库和TCGA中的胃癌数据,分析SERPINE1的表达。同时使用Kaplan-Meier plotter和基因表达谱互动分析(GEPIA)评估SERPINE1对临床预后的影响。使用基因集富集评分(GSEA)对基因免疫途径进行了富集分析。此外,使用TIMER、TISIDB和GEPIA分析SERPINE1表达与免疫浸润标志物的相关性。结果对来自TCGA胃癌患者的375个肿瘤组织和32个正常组织的RNA测序数据的分析表明,SERPINE1在肿瘤组织中的表达明显高于正常组织。Kaplan-Meier plotter及GEPIA分析得出SERPINE1高表达与较差的预后明显相关(Hr=1.44,95%CI=1.21~1.71,P<0.05)。TISIDB和TIMER数据库分析后发现,SERPINE1的表达与胃癌患者肿瘤免疫细胞浸润明显相关,并与免疫和基质成分增加相关。结论SERPINE1的表达与胃癌患者的预后和各种类型的免疫细胞浸润相关,可作为判断胃癌预后和免疫治疗的生物标志物。展开更多
Objective： To investigated whether epigenetic mechanisms contribute to the variable expression of variable protease nexin1（PN-1） encoded by the SERPINE2 gene in different cell types. Methods： Working with 5 human ...Objective： To investigated whether epigenetic mechanisms contribute to the variable expression of variable protease nexin1（PN-1） encoded by the SERPINE2 gene in different cell types. Methods： Working with 5 human cell lines, we determined the CpG methylation status within two CpG islands in the SERPINE2 gene by bisulphate sequencing and the PN-1 mRNA level by Q-RT PCR. Results： A CpG island spanning the transcription initiation site showed little methylation in 3 of the cell lines and substantial methylation in 2 of the cell lines. A CpG island covering the translation starting site showed full methylation in all investigated cell lines. Methylation within the CpG island was not randomly distributed, but showed accumulation at specific sites. However, we were not able to distinguish any patterns which related the methylation frequency to the gene expression level. Inhibition of CpG methylation with 5-aza-2’-deoxycytidine led to a several fold increase in PN-1 mRNA levels, but based on the results on CpG methylation in the CpG island spanning the transcript, the effect is most likely indirect. Conclusion： We have carefully mapped the CpG methylation pattern in two CpG islands in the 5’ part of the SERPINE2 gene without finding any obvious inverse correlation between methylation frequency and expression level.展开更多
Background:Liver cancer is a malignancy with high morbidity and mortality rates.Serpin family E member 2(SERPINE2)has been reported to play a key role in the metastasis of many tumors.In this study,we aimed to investi...Background:Liver cancer is a malignancy with high morbidity and mortality rates.Serpin family E member 2(SERPINE2)has been reported to play a key role in the metastasis of many tumors.In this study,we aimed to investigate the potential mechanism of SERPINE2 in liver cancer metastasis.Methods:The Cancer Genome Atlas database(TCGA),including DNA methy-lation and transcriptome sequencing data,was utilized to identify the crucial oncogene associated with DNA methylation and cancer progression in liver can-cer.Data from the TCGA and RNA sequencing for 94 pairs of liver cancer tissues were used to explore the correlation between SERPINE2 expression and clin-ical parameters of patients.DNA methylation sequencing was used to detect the DNA methylation levels in liver cancer tissues and cells.RNA sequencing,cytokine assays,immunoprecipitation(IP)and mass spectrometry(MS)assays,protein stability assays,and ubiquitination assays were performed to explore the regulatory mechanism of SERPINE2 in liver cancer metastasis.Patient-derived xenografts and tumor organoid models were established to determine the role of SERPINE2 in the treatment of liver cancer using sorafenib.Results:Based on the public database screening,SERPINE2 was identified as a tumor promoter regulated by DNA methylation.SERPINE2 expression was significantly higher in liver cancer tissues and was associated with the dismal prognosis in patients with liver cancer.SERPINE2 promoted liver cancer metas-tasis by enhancing cell pseudopodia formation,cell adhesion,cancer-associated fibroblast activation,extracellular matrix remodeling,and angiogenesis.IP/MS assays confirmed that SERPINE2 activated epidermal growth factor receptor(EGFR)and its downstream signaling pathways by interacting with EGFR.Mechanistically,SERPINE2 inhibited EGFR ubiquitination and maintained its protein stability by competing with the E3 ubiquitin ligase,c-Cbl.Additionally,EGFR was activated in liver cancer cells after sorafenib treatment,and SER-PINE2 knockdown-induced EGFR downregulation significantly enhanced the therapeutic efficacy of sorafenib against liver cancer.Furthermore,we found that SERPINE2 knockdown also had a sensitizing effect on lenvatinib treatment.Conclusions:SERPINE2 promoted liver cancer metastasis by preventing EGFR degradation via c-Cbl-mediated ubiquitination,suggesting that inhibition of the SERPINE2-EGFR axis may be a potential target for liver cancer treatment.展开更多
文摘【目的】克隆牦牛丝氨酸蛋白酶抑制剂家族E成员1(serpin family E member 1,SERPINE1)基因序列,同时探究不同嫩度牦牛背最长肌组织中SERPINE1基因核苷酸序列差异,为进一步研究其对牦牛肉嫩度的影响提供试验数据。【方法】根据GenBank中牦牛SERPINE1基因序列设计特异性引物,通过PCR扩增克隆获得高、低嫩度牦牛背最长肌中的SERPINE1基因序列全长,并对其结构及编码蛋白进行生物信息学分析,通过实时荧光定量PCR检测不同嫩度牦牛背最长肌中SERPINE1基因表达量。【结果】高、低嫩度牦牛背最长肌组织中SERPINE1基因全长分别为8 315和8 318 bp,均编码402个氨基酸且氨基酸序列完全相同。高、低嫩度背最长肌组织中SERPINE1基因非编码序列中存在3个碱基缺失及22个碱基突变(4个碱基颠换、18个碱基转换)。牦牛SERPINE1基因核苷酸序列与普通牛、瘤牛、美洲野牛、绵羊、山羊、家猪、人、小鼠的相似性分别为99.26%、99.26%、99.59%、96.94%、97.02%、90.49%、86.52%和80.10%。SERPINE1蛋白是一个含有23个氨基酸信号肽的亲水性稳定膜外蛋白,位于细胞外,具有34个潜在磷酸化位点,包含1个反应中心环(reactive center loop, RCL)。SERPINE1蛋白二级结构主要由α-螺旋(44.78%)和无规则卷曲(35.32%)构成。实时荧光定量PCR结果显示,SERPINE1基因在高嫩度牦牛背最长肌中表达量极显著高于低嫩度牦牛背最长肌(P<0.01)。【结论】本研究成功从高、低嫩度牦牛背最长肌组织中克隆SERPINE1基因全长并进行了生物信息学特征分析,为后续研究SERPINE1基因参与调控牦牛肉嫩度机制提供理论参考。
文摘目的探讨丝氨酸蛋白酶抑制剂分支E成员1(serpin family E member 1,SERPINE1)的表达与胃癌的预后及肿瘤免疫细胞浸润的关系。方法通过肿瘤免疫估计资源(TIMER)数据库和TCGA中的胃癌数据,分析SERPINE1的表达。同时使用Kaplan-Meier plotter和基因表达谱互动分析(GEPIA)评估SERPINE1对临床预后的影响。使用基因集富集评分(GSEA)对基因免疫途径进行了富集分析。此外,使用TIMER、TISIDB和GEPIA分析SERPINE1表达与免疫浸润标志物的相关性。结果对来自TCGA胃癌患者的375个肿瘤组织和32个正常组织的RNA测序数据的分析表明,SERPINE1在肿瘤组织中的表达明显高于正常组织。Kaplan-Meier plotter及GEPIA分析得出SERPINE1高表达与较差的预后明显相关(Hr=1.44,95%CI=1.21~1.71,P<0.05)。TISIDB和TIMER数据库分析后发现,SERPINE1的表达与胃癌患者肿瘤免疫细胞浸润明显相关,并与免疫和基质成分增加相关。结论SERPINE1的表达与胃癌患者的预后和各种类型的免疫细胞浸润相关,可作为判断胃癌预后和免疫治疗的生物标志物。
基金supported by the Danish National Research Foundation (26-331-6)the Danish Cancer Society (DP 07043, DP 08001)Grosserer Alfred Nielsen and Hustrus Fond
文摘Objective： To investigated whether epigenetic mechanisms contribute to the variable expression of variable protease nexin1（PN-1） encoded by the SERPINE2 gene in different cell types. Methods： Working with 5 human cell lines, we determined the CpG methylation status within two CpG islands in the SERPINE2 gene by bisulphate sequencing and the PN-1 mRNA level by Q-RT PCR. Results： A CpG island spanning the transcription initiation site showed little methylation in 3 of the cell lines and substantial methylation in 2 of the cell lines. A CpG island covering the translation starting site showed full methylation in all investigated cell lines. Methylation within the CpG island was not randomly distributed, but showed accumulation at specific sites. However, we were not able to distinguish any patterns which related the methylation frequency to the gene expression level. Inhibition of CpG methylation with 5-aza-2’-deoxycytidine led to a several fold increase in PN-1 mRNA levels, but based on the results on CpG methylation in the CpG island spanning the transcript, the effect is most likely indirect. Conclusion： We have carefully mapped the CpG methylation pattern in two CpG islands in the 5’ part of the SERPINE2 gene without finding any obvious inverse correlation between methylation frequency and expression level.
基金This work was supported by grants from the National Nat-ural Science Foundation of China(82070652 and 81870434)Department of Science and Technology of Zhejiang Province(2020C04003)+2 种基金the Chinese Academy of Medi-cal Sciences(019-I2M-5-030)the Jinan Microecological Biomedicine Shandong Laboratory(JNL-2022007B)the State Key Laboratory for Diagnosis and Treatment of Infectious Diseases(zz202302).
文摘Background:Liver cancer is a malignancy with high morbidity and mortality rates.Serpin family E member 2(SERPINE2)has been reported to play a key role in the metastasis of many tumors.In this study,we aimed to investigate the potential mechanism of SERPINE2 in liver cancer metastasis.Methods:The Cancer Genome Atlas database(TCGA),including DNA methy-lation and transcriptome sequencing data,was utilized to identify the crucial oncogene associated with DNA methylation and cancer progression in liver can-cer.Data from the TCGA and RNA sequencing for 94 pairs of liver cancer tissues were used to explore the correlation between SERPINE2 expression and clin-ical parameters of patients.DNA methylation sequencing was used to detect the DNA methylation levels in liver cancer tissues and cells.RNA sequencing,cytokine assays,immunoprecipitation(IP)and mass spectrometry(MS)assays,protein stability assays,and ubiquitination assays were performed to explore the regulatory mechanism of SERPINE2 in liver cancer metastasis.Patient-derived xenografts and tumor organoid models were established to determine the role of SERPINE2 in the treatment of liver cancer using sorafenib.Results:Based on the public database screening,SERPINE2 was identified as a tumor promoter regulated by DNA methylation.SERPINE2 expression was significantly higher in liver cancer tissues and was associated with the dismal prognosis in patients with liver cancer.SERPINE2 promoted liver cancer metas-tasis by enhancing cell pseudopodia formation,cell adhesion,cancer-associated fibroblast activation,extracellular matrix remodeling,and angiogenesis.IP/MS assays confirmed that SERPINE2 activated epidermal growth factor receptor(EGFR)and its downstream signaling pathways by interacting with EGFR.Mechanistically,SERPINE2 inhibited EGFR ubiquitination and maintained its protein stability by competing with the E3 ubiquitin ligase,c-Cbl.Additionally,EGFR was activated in liver cancer cells after sorafenib treatment,and SER-PINE2 knockdown-induced EGFR downregulation significantly enhanced the therapeutic efficacy of sorafenib against liver cancer.Furthermore,we found that SERPINE2 knockdown also had a sensitizing effect on lenvatinib treatment.Conclusions:SERPINE2 promoted liver cancer metastasis by preventing EGFR degradation via c-Cbl-mediated ubiquitination,suggesting that inhibition of the SERPINE2-EGFR axis may be a potential target for liver cancer treatment.
