目的:探讨过表达MARCH2对肝细胞癌HUH7细胞自噬水平和生长的影响及其机制。方法:MTS和EDU检测MARCH2过表达的肝细胞癌HUH7细胞的生长情况。流式细胞术检测MARCH2过表达的肝细胞癌HUH7细胞凋亡、细胞周期的情况。Western blot和激光共聚...目的:探讨过表达MARCH2对肝细胞癌HUH7细胞自噬水平和生长的影响及其机制。方法:MTS和EDU检测MARCH2过表达的肝细胞癌HUH7细胞的生长情况。流式细胞术检测MARCH2过表达的肝细胞癌HUH7细胞凋亡、细胞周期的情况。Western blot和激光共聚焦法检测MARCH2过表达的肝细胞癌HUH7细胞自噬水平。结果:MTS和EDU检测表明,MARCH2的过表达促进了肝细胞癌HUH7的生长。流式细胞术检测表明,MARCH2过表达对肝细胞癌HUH7细胞凋亡和细胞周期没有明显影响。进一步的实验表明,MARCH2过表达损害了肝细胞癌HUH7的自噬。用常见的自噬诱导剂雷帕霉素挽救自噬可以使肝细胞癌HUH7的增殖率恢复到控制水平。结论:我们的数据表明,MARCH2通过抑制自噬在肿瘤发展过程中起着至关重要的作用。因此,MARCH2是一种有前景的肿瘤治疗新靶点。Aims: To investigate the effects and mechanisms of overexpression of MARCH2 on autophagy levels and growth of hepatocellular carcinoma HUH7 cells. Methods: MTS and EDU were used to detect the growth of MARCH2 overexpressing hepatocellular carcinoma HUH7 cells. Flow cytometry was used to detect apoptosis and cell cycle in MARCH2 overexpressing hepatocellular carcinoma HUH7 cells. Western blot and laser confocal microscopy were used to detect the autophagy levels in MARCH2 overexpressing HUH7 hepatocellular carcinoma cells. Results: In this study, MTS and EDU assays showed that the overexpression of MARCH2 promoted the growth of hepatocellular carcinoma HUH7 cells. Flow cytometry detection illustrated that MARCH2 overexpression had no obvious effect on the hepatocellular carcinoma HUH7 cell apoptosis and cell cycle. Further experiments displayed that MARCH2 overexpression impaired the autophagy of hepatocellular carcinoma HUH7 cells. Rescued autophagy with the common autophagy inducer rapamycin can bring the proliferation rate of hepatocellular carcinoma HUH7 cells back to the control level. Conclusions: Our data indicates that MARCH2 plays an essential role in the progression of tumor development through inhibiting autophagy. Therefore, MARCH2 is an attractive novel target for tumor therapy.展开更多
目的探讨泛素E3连接酶膜相关锌指蛋白2(membrane-associated RING CH 2,MARCH2)在胃癌细胞株BGC823侵袭转移中的生物学作用以及在胃癌组织中的表达及与预后的关系。方法荧光定量PCR检测40例配对胃癌组织和癌旁正常组织中的MARCH2表达水...目的探讨泛素E3连接酶膜相关锌指蛋白2(membrane-associated RING CH 2,MARCH2)在胃癌细胞株BGC823侵袭转移中的生物学作用以及在胃癌组织中的表达及与预后的关系。方法荧光定量PCR检测40例配对胃癌组织和癌旁正常组织中的MARCH2表达水平。Kaplan-Meier生存曲线分析胃癌患者生存情况。利用沉默MARCH2病毒上清感染BGC823细胞,荧光定量PCR及Western blot检测感染效率;利用CCK8细胞增殖实验、Boyden小室侵袭实验及裸鼠尾静脉肺转移实验观察BGC823细胞体外增殖和侵袭能力及体内肺转移能力。结果MARCH2在胃癌组织中mRNA表达上调(P<0.05)。胃癌组织较癌旁正常组织MARCH2 mRNA高表达的患者(n=28)中位生存时间短于低表达的患者(n=12;30个月vs 43.5个月,P<0.05)。沉默MARCH2后,BGC823细胞体外增殖和侵袭能力均下降(均P<0.05),裸鼠尾静脉肺转移能力下降(P<0.05)。结论MARCH2在胃癌中是一个促癌基因,沉默MARCH2抑制胃癌细胞BGC823体外增殖与侵袭及体内肺转移能力。展开更多
目的:检测浸润性乳腺癌组织及癌旁正常乳腺组织中膜相关环指蛋白2(membrane-associated ring CH2,MARCH2)和核转录因子κB(nuclear transcription factor-κB,NF-κB)的表达,探讨MARCH2、NF-κB在浸润性乳腺癌中的表达与临床病理参数之...目的:检测浸润性乳腺癌组织及癌旁正常乳腺组织中膜相关环指蛋白2(membrane-associated ring CH2,MARCH2)和核转录因子κB(nuclear transcription factor-κB,NF-κB)的表达,探讨MARCH2、NF-κB在浸润性乳腺癌中的表达与临床病理参数之间的关系。方法:收集自贡市第四人民医院2018年1月至12月期间接受外科手术治疗的浸润性乳腺癌患者肿瘤组织及距肿瘤边缘﹥3 cm处的正常乳腺石蜡组织各40例。采用免疫组织化学Envision法对组织标本中的MARCH2蛋白、NF-κB蛋白进行标记。结果:在40例浸润性乳腺癌患者肿瘤组织中,MARCH2在癌组织中阳性表达36例(阳性表达率90.0%),癌旁组织阳性表达7例(χ^2=42.288,P﹤0.05)。NF-κB在癌组织中阳性表达30例(阳性表达率75.0%),癌旁正常组织阳性表达5例(χ^2=31.746,P﹤0.05)。关联性分析结果表明,两种蛋白的表达呈中度正相关(r﹦0.577,χ^2=13.333,P﹤0.01)。结论:MARCH2和NF-κB在浸润性乳腺癌组织中呈高表达,乳腺癌的发生发展可能与MARCH2的高表达有关,NF-κB可能是乳腺癌发生发展的重要信号通路。展开更多
目的:探讨MARCH2多克隆抗体的制备、鉴定与纯化的方法以及其在组织和细胞系中的表达情况、在亚细胞结构中的定位情况。方法:利用DNAstar软件对MARCH2蛋白的抗原性等进行分析,化学合成MARCH2短肽,制备成完全抗原免疫家兔,获取血清,纯化,...目的:探讨MARCH2多克隆抗体的制备、鉴定与纯化的方法以及其在组织和细胞系中的表达情况、在亚细胞结构中的定位情况。方法:利用DNAstar软件对MARCH2蛋白的抗原性等进行分析,化学合成MARCH2短肽,制备成完全抗原免疫家兔,获取血清,纯化,用Western blot、Elisa、免疫荧光进行鉴定。用RT-PCR检测MARCH2在细胞系中的表达情况,用Western blot检测MARCH2在组织中的表达情况。用免疫荧光法及激光共聚焦显微镜分析检测MARCH2在亚细胞结构中的定位。结果:成功制备完全抗原免疫家兔,纯化出多克隆抗体,用Western blot、Elisa、免疫荧光法证实多克隆抗体制备成功。利用半定量RT-PCR方法,在32种细胞系中检测了MARCH2 mRNA的表达水平,结果显示,MARCH2呈广泛表达,在HeLa、U2OS、HCT116、COS7细胞中表达最高,在SKBR3、HGC-27、MGC-803细胞中低表达。利用组织芯片及免疫组织化学方法进一步分析了其在正常组织及肿瘤组织中的表达情况,染色结果显示,MARCH2呈广泛表达,在前列腺、肝组织中呈高表达,在横纹肌、甲状腺组织中呈低表达,主要在胞浆中呈现弥漫均匀细颗粒状分布。此外,MARCH2表达因肿瘤类型的不同有差异。与对应的正常组织相比,MARCH2在某些肿瘤(如前列腺癌、肝癌)组织中表达降低,而在某些肿瘤(如食管癌、结肠癌)组织中表达增高。用免疫荧光法及激光共聚焦显微镜分析检测MARCH2在亚细胞结构中的定位,发现MARCH2与内质网、高尔基体共定位,与溶酶体、内体部分共定位,与线粒体没有共定位。结论:成功获得了MARCH2多克隆抗体,为进一步研究MARCH2蛋白的功能奠定了基础。MARCH2在不同类型肿瘤中的差异表达及其在亚细胞结构中的定位高度提示MARCH2在肿瘤发生发展中具有重要的潜在应用价值。Aims: To explore the preparation, identification, and purification methods of MARCH2 polyclonal antibodies, as well as their expression and subcellular structural localization in tissues and cell lines. Methods: DNAstar software was used to analyze the antigenicity of MARCH2 protein, and MARCH2 short peptides were chemically synthesized to prepare complete antigen-immunized rabbits. Serum was obtained, purified, and identified by Western blot, Elisa, and immunofluorescence. RT-PCR was used to detect the expression of MARCH2 in cell lines, and Western blot was used to detect the expression of MARCH2 in tissues. Immunofluorescence and confocal laser microscopy were used to analyze and detect the localization of MARCH2 in subcellular structures. Results: Fully antigen-immunized rabbits were successfully prepared, and polyclonal antibodies were purified. Western blot, Elisa, and immunofluorescence were used to confirm the successful preparation of polyclonal antibodies. Using semi-quantitative RT-PCR method, the expression level of MARCH2 mRNA was detected in 32 cell lines. The results showed that MARCH2 was widely expressed, with the highest expression in HeLa, U2OS, HCT116, and COS7 cells, and low expression in SKBR3, HGC-27, and MGC-803 cells. The expression of MARCH2 in normal and tumor tissues was further analyzed using tissue chips and immunohistochemical methods. The staining results showed that MARCH2 was widely expressed, with high expression in prostate and liver tissues, low expression in striated muscle and thyroid tissues, and mainly distributed in a diffuse and uniform granular form in the cytoplasm. In addition, MARCH2 expression varies depending on the type of tumor. Compared with corresponding normal tissues, MARCH2 expression is reduced in certain tumor tissues (such as prostate cancer and liver cancer), while it is increased in certain tumor tissues (such as esophageal cancer and colon cancer). Using immunofluorescence and laser confocal microscopy to analyze and detect the localization of MARCH2 in subcellular structures, it was found that MARCH2 was co-localized with the endoplasmic reticulum and Golgi apparatus, with lysosomes and endosomes partially, but not with mitochondria. Conclusions: MARCH2 polyclonal antibodies have been successfully obtained, laying the foundation for further research on the function of MARCH2 protein. The differential expression of MARCH2 in different types of tumors and its localization in subcellular structures highly suggest that MARCH2 has important potential application value in tumor occurrence and development.展开更多
目的:探讨MARCH2和CFTR在结直肠癌中的蛋白表达及与临床病理参数的相关性。方法:采用免疫组织化学SP法检测125例结直肠癌和30例正常结直肠黏膜组织中MARCH2和CFTR的表达,并复习相关文献。结果:在125例结直肠癌组织中,MARCH2和CFTR的阳...目的:探讨MARCH2和CFTR在结直肠癌中的蛋白表达及与临床病理参数的相关性。方法:采用免疫组织化学SP法检测125例结直肠癌和30例正常结直肠黏膜组织中MARCH2和CFTR的表达,并复习相关文献。结果:在125例结直肠癌组织中,MARCH2和CFTR的阳性表达率分别为84.8%和8.8%,而在非肿瘤性肠组织中,这两种表达率的阳性率分别为6.7%和90.0%。MARCH2的阳性表达与肿瘤大小、深部浸润、淋巴结转移和TNM分期密切相关。此外,CFTR表达的缺失与分化不良、淋巴结转移和TNM分期显著相关。此外,MARCH2和CFTR表达之间存在显著负相关。结论:MARCH2在结直肠癌中高度表达,与肿瘤大小、深部浸润、淋巴结转移和TNM分期等预后不良参数有关。相比之下,CFTR在结直肠癌中几乎不表达,CFTR表达缺失与分化不良、淋巴结转移和TNM分期显著相关。MARCH2可能通过与CFTR的相互作用在结直肠癌的发展中发挥重要作用。它们可能成为结直肠癌诊断、治疗和预后评估的分子标志物。Aims: This study aimed to investigate whether the expressions of MARCH2 and CFTR in colorectal carcinomas were associated with clinicopathological parameters. Methods: The expressions of MARCH2 and CFTR were examined in 125 cases of colorectal carcinomas and 30 normal colorectal tissues using immunohistochemical staining. Results: The positive rates of MARCH2 and CFTR expressions in 125 cases of colorectal carcinoma were 84.8% and 8.8% respectively, whereas the positive expression rates of MARCH2 and CFTR in non-tumor colorectal tissues were 6.7% and 90.0% respectively. The positive expressions of MARCH2 were closely related to tumor size, deep invasion, lymph node metastasis and advanced TNM stage. In addition, loss of expression of CFTR was significantly associated with poor differentiation, lymph node metastasis and advanced TNM stage. Furthermore, there were significant negative correlations between MARCH2 and CFTR expression. Conclusions: MARCH2 is highly expressed in colorectal carcinomas, and is associated with poor prognostic parameters such as tumor size, deep invasion, lymph node metastasis and advanced TNM stage. In contrast, CFTR barely expressed in colorectal carcinomas, and loss of expression of CFTR was significantly associated with poor differentiation, lymph node metastasis and advanced TNM stage. MARCH2 may play an important role in the development of colorectal carcinoma through interaction with CFTR. They may become molecular markers for diagnosis, treatment and prognostic evaluation of colorectal carcinomas.展开更多
Membrane-associated RING-CH protein 2 (MARCH2)是MARCH家族成员之一,主要负责囊泡运输,本研究构建了过表达MARCH2和沉默MARCH2的HO-8910细胞系,目的是为了研究MARCH2对卵巢癌细胞自噬及生长的影响。研究发现,沉默MARCH2可以促进自噬...Membrane-associated RING-CH protein 2 (MARCH2)是MARCH家族成员之一,主要负责囊泡运输,本研究构建了过表达MARCH2和沉默MARCH2的HO-8910细胞系,目的是为了研究MARCH2对卵巢癌细胞自噬及生长的影响。研究发现,沉默MARCH2可以促进自噬体的形成和自噬流,相反,过表达MARCH2会抑制自噬体的形成和自噬流,进一步研究发现,MARCH2缺失介导的自噬作用在ULK1和PIK3C3-BECN1复合物的上游。此外,沉默MARCH2可以抑制体外细胞增殖,亦会抑制体外裸鼠移植瘤的生长。这些效应与自噬信号的活化有关系。展开更多
T-cell immunoglobulin mucin family member-1(TIM-1,also known as HAVCR1/KIM-1)is a transmembrane glycoprotein that has been reported to act as an entry receptor for multiple flaviviruses including Zika virus(ZIKV).The ...T-cell immunoglobulin mucin family member-1(TIM-1,also known as HAVCR1/KIM-1)is a transmembrane glycoprotein that has been reported to act as an entry receptor for multiple flaviviruses including Zika virus(ZIKV).The post-translational regulation of TIM-1 and its effects on ZIKV infection are unclear.In this study,we identified the membrane-associated RING-CH-type finger(MARCH)E3 ubiquitin ligase family members MARCH2 and MARCH3 as critical negative regulators of TIM-1 under physiological conditions.MARCH2 and MARCH3 associate with TIM-1 and mediate its K48-linked polyubiquitination at K338 and K346 respectively,leading to subsequent proteasomal degradation.While deficiency of either MARCH2 or MARCH3 modestly increases TIM-1 levels and enhances ZIKV infectivity,double knockout of MARCH2/3 has a more dramatic effect.Double knockout of MARCH2/3 increased ZIKV infectivity in wild-type but not TIM-1 knockout cells,and reconstitution of TIM-1^(K338R/K346R) into TIM-1-deficient cells increases ZIKV infectivity to a higher degree than reconstitution with wild-type TIM-1.Knockout of either MARCH2 or MARCH3 increased ZIKV infectivity and pathogenesis in mice,whereas double knockout of MARCH2/3 has a more dramatic effect.These findings suggest that MARCH2 and MARCH3 target TIM-1 for K48-linked polyubiquitination and proteasomal degradation,thereby acting as redundant host restriction factors to limit ZIKV infection and pathogenesis.展开更多
Interleukin 5(IL-5)plays crucial roles in type 2-high asthma by mediating eosinophil maturation,activation,chemotaxis and survival.Inhibition of IL-5 signaling is considered a strategy for asthma treatment.Here,we ide...Interleukin 5(IL-5)plays crucial roles in type 2-high asthma by mediating eosinophil maturation,activation,chemotaxis and survival.Inhibition of IL-5 signaling is considered a strategy for asthma treatment.Here,we identified MARCH2 and MARCH3 as critical negative regulators of IL-5-triggered signaling.MARCH2 and MARCH3 associate with the IL-5 receptorαchain(IL-5Rα)and mediate its K27-linked polyubiquitination at K379 and K383,respectively,and its subsequent lysosomal degradation.Deficiency of MARCH2 or MARCH3 modestly increases the level of IL-5Rαand enhances IL-5-induced signaling,whereas double knockout of MARCH2/3 has a more dramatic effect.March2/3 double knockout markedly increases the proportions of eosinophils in the bone marrow and peripheral blood in mice.Double knockout of March2/3 aggravates ovalbumin(OVA)-induced eosinophilia and causes increased inflammatory cell infiltration,peribronchial mucus secretion and production of Th2 cytokines.Neutralization of Il-5 attenuates OVA-induced airway inflammation and the enhanced effects of March2/3 double deficiency.These findings suggest that MARCH2 and MARCH3 play redundant roles in targeting IL-5Rαfor degradation and negatively regulating allergic airway inflammation.展开更多
文摘目的:探讨过表达MARCH2对肝细胞癌HUH7细胞自噬水平和生长的影响及其机制。方法:MTS和EDU检测MARCH2过表达的肝细胞癌HUH7细胞的生长情况。流式细胞术检测MARCH2过表达的肝细胞癌HUH7细胞凋亡、细胞周期的情况。Western blot和激光共聚焦法检测MARCH2过表达的肝细胞癌HUH7细胞自噬水平。结果:MTS和EDU检测表明,MARCH2的过表达促进了肝细胞癌HUH7的生长。流式细胞术检测表明,MARCH2过表达对肝细胞癌HUH7细胞凋亡和细胞周期没有明显影响。进一步的实验表明,MARCH2过表达损害了肝细胞癌HUH7的自噬。用常见的自噬诱导剂雷帕霉素挽救自噬可以使肝细胞癌HUH7的增殖率恢复到控制水平。结论:我们的数据表明,MARCH2通过抑制自噬在肿瘤发展过程中起着至关重要的作用。因此,MARCH2是一种有前景的肿瘤治疗新靶点。Aims: To investigate the effects and mechanisms of overexpression of MARCH2 on autophagy levels and growth of hepatocellular carcinoma HUH7 cells. Methods: MTS and EDU were used to detect the growth of MARCH2 overexpressing hepatocellular carcinoma HUH7 cells. Flow cytometry was used to detect apoptosis and cell cycle in MARCH2 overexpressing hepatocellular carcinoma HUH7 cells. Western blot and laser confocal microscopy were used to detect the autophagy levels in MARCH2 overexpressing HUH7 hepatocellular carcinoma cells. Results: In this study, MTS and EDU assays showed that the overexpression of MARCH2 promoted the growth of hepatocellular carcinoma HUH7 cells. Flow cytometry detection illustrated that MARCH2 overexpression had no obvious effect on the hepatocellular carcinoma HUH7 cell apoptosis and cell cycle. Further experiments displayed that MARCH2 overexpression impaired the autophagy of hepatocellular carcinoma HUH7 cells. Rescued autophagy with the common autophagy inducer rapamycin can bring the proliferation rate of hepatocellular carcinoma HUH7 cells back to the control level. Conclusions: Our data indicates that MARCH2 plays an essential role in the progression of tumor development through inhibiting autophagy. Therefore, MARCH2 is an attractive novel target for tumor therapy.
文摘目的:探讨MARCH2多克隆抗体的制备、鉴定与纯化的方法以及其在组织和细胞系中的表达情况、在亚细胞结构中的定位情况。方法:利用DNAstar软件对MARCH2蛋白的抗原性等进行分析,化学合成MARCH2短肽,制备成完全抗原免疫家兔,获取血清,纯化,用Western blot、Elisa、免疫荧光进行鉴定。用RT-PCR检测MARCH2在细胞系中的表达情况,用Western blot检测MARCH2在组织中的表达情况。用免疫荧光法及激光共聚焦显微镜分析检测MARCH2在亚细胞结构中的定位。结果:成功制备完全抗原免疫家兔,纯化出多克隆抗体,用Western blot、Elisa、免疫荧光法证实多克隆抗体制备成功。利用半定量RT-PCR方法,在32种细胞系中检测了MARCH2 mRNA的表达水平,结果显示,MARCH2呈广泛表达,在HeLa、U2OS、HCT116、COS7细胞中表达最高,在SKBR3、HGC-27、MGC-803细胞中低表达。利用组织芯片及免疫组织化学方法进一步分析了其在正常组织及肿瘤组织中的表达情况,染色结果显示,MARCH2呈广泛表达,在前列腺、肝组织中呈高表达,在横纹肌、甲状腺组织中呈低表达,主要在胞浆中呈现弥漫均匀细颗粒状分布。此外,MARCH2表达因肿瘤类型的不同有差异。与对应的正常组织相比,MARCH2在某些肿瘤(如前列腺癌、肝癌)组织中表达降低,而在某些肿瘤(如食管癌、结肠癌)组织中表达增高。用免疫荧光法及激光共聚焦显微镜分析检测MARCH2在亚细胞结构中的定位,发现MARCH2与内质网、高尔基体共定位,与溶酶体、内体部分共定位,与线粒体没有共定位。结论:成功获得了MARCH2多克隆抗体,为进一步研究MARCH2蛋白的功能奠定了基础。MARCH2在不同类型肿瘤中的差异表达及其在亚细胞结构中的定位高度提示MARCH2在肿瘤发生发展中具有重要的潜在应用价值。Aims: To explore the preparation, identification, and purification methods of MARCH2 polyclonal antibodies, as well as their expression and subcellular structural localization in tissues and cell lines. Methods: DNAstar software was used to analyze the antigenicity of MARCH2 protein, and MARCH2 short peptides were chemically synthesized to prepare complete antigen-immunized rabbits. Serum was obtained, purified, and identified by Western blot, Elisa, and immunofluorescence. RT-PCR was used to detect the expression of MARCH2 in cell lines, and Western blot was used to detect the expression of MARCH2 in tissues. Immunofluorescence and confocal laser microscopy were used to analyze and detect the localization of MARCH2 in subcellular structures. Results: Fully antigen-immunized rabbits were successfully prepared, and polyclonal antibodies were purified. Western blot, Elisa, and immunofluorescence were used to confirm the successful preparation of polyclonal antibodies. Using semi-quantitative RT-PCR method, the expression level of MARCH2 mRNA was detected in 32 cell lines. The results showed that MARCH2 was widely expressed, with the highest expression in HeLa, U2OS, HCT116, and COS7 cells, and low expression in SKBR3, HGC-27, and MGC-803 cells. The expression of MARCH2 in normal and tumor tissues was further analyzed using tissue chips and immunohistochemical methods. The staining results showed that MARCH2 was widely expressed, with high expression in prostate and liver tissues, low expression in striated muscle and thyroid tissues, and mainly distributed in a diffuse and uniform granular form in the cytoplasm. In addition, MARCH2 expression varies depending on the type of tumor. Compared with corresponding normal tissues, MARCH2 expression is reduced in certain tumor tissues (such as prostate cancer and liver cancer), while it is increased in certain tumor tissues (such as esophageal cancer and colon cancer). Using immunofluorescence and laser confocal microscopy to analyze and detect the localization of MARCH2 in subcellular structures, it was found that MARCH2 was co-localized with the endoplasmic reticulum and Golgi apparatus, with lysosomes and endosomes partially, but not with mitochondria. Conclusions: MARCH2 polyclonal antibodies have been successfully obtained, laying the foundation for further research on the function of MARCH2 protein. The differential expression of MARCH2 in different types of tumors and its localization in subcellular structures highly suggest that MARCH2 has important potential application value in tumor occurrence and development.
文摘目的:探讨MARCH2和CFTR在结直肠癌中的蛋白表达及与临床病理参数的相关性。方法:采用免疫组织化学SP法检测125例结直肠癌和30例正常结直肠黏膜组织中MARCH2和CFTR的表达,并复习相关文献。结果:在125例结直肠癌组织中,MARCH2和CFTR的阳性表达率分别为84.8%和8.8%,而在非肿瘤性肠组织中,这两种表达率的阳性率分别为6.7%和90.0%。MARCH2的阳性表达与肿瘤大小、深部浸润、淋巴结转移和TNM分期密切相关。此外,CFTR表达的缺失与分化不良、淋巴结转移和TNM分期显著相关。此外,MARCH2和CFTR表达之间存在显著负相关。结论:MARCH2在结直肠癌中高度表达,与肿瘤大小、深部浸润、淋巴结转移和TNM分期等预后不良参数有关。相比之下,CFTR在结直肠癌中几乎不表达,CFTR表达缺失与分化不良、淋巴结转移和TNM分期显著相关。MARCH2可能通过与CFTR的相互作用在结直肠癌的发展中发挥重要作用。它们可能成为结直肠癌诊断、治疗和预后评估的分子标志物。Aims: This study aimed to investigate whether the expressions of MARCH2 and CFTR in colorectal carcinomas were associated with clinicopathological parameters. Methods: The expressions of MARCH2 and CFTR were examined in 125 cases of colorectal carcinomas and 30 normal colorectal tissues using immunohistochemical staining. Results: The positive rates of MARCH2 and CFTR expressions in 125 cases of colorectal carcinoma were 84.8% and 8.8% respectively, whereas the positive expression rates of MARCH2 and CFTR in non-tumor colorectal tissues were 6.7% and 90.0% respectively. The positive expressions of MARCH2 were closely related to tumor size, deep invasion, lymph node metastasis and advanced TNM stage. In addition, loss of expression of CFTR was significantly associated with poor differentiation, lymph node metastasis and advanced TNM stage. Furthermore, there were significant negative correlations between MARCH2 and CFTR expression. Conclusions: MARCH2 is highly expressed in colorectal carcinomas, and is associated with poor prognostic parameters such as tumor size, deep invasion, lymph node metastasis and advanced TNM stage. In contrast, CFTR barely expressed in colorectal carcinomas, and loss of expression of CFTR was significantly associated with poor differentiation, lymph node metastasis and advanced TNM stage. MARCH2 may play an important role in the development of colorectal carcinoma through interaction with CFTR. They may become molecular markers for diagnosis, treatment and prognostic evaluation of colorectal carcinomas.
文摘Membrane-associated RING-CH protein 2 (MARCH2)是MARCH家族成员之一,主要负责囊泡运输,本研究构建了过表达MARCH2和沉默MARCH2的HO-8910细胞系,目的是为了研究MARCH2对卵巢癌细胞自噬及生长的影响。研究发现,沉默MARCH2可以促进自噬体的形成和自噬流,相反,过表达MARCH2会抑制自噬体的形成和自噬流,进一步研究发现,MARCH2缺失介导的自噬作用在ULK1和PIK3C3-BECN1复合物的上游。此外,沉默MARCH2可以抑制体外细胞增殖,亦会抑制体外裸鼠移植瘤的生长。这些效应与自噬信号的活化有关系。
基金supported by grants from the State Key R&D Program of China(2024YFA1306500,2022YFA1304900)the National Natural Science Foundation of China(32188101)+2 种基金the Major Project of Guangzhou National Laboratory(GZNL2024A01014,GZNL2024A01016)the Fundamental Research Funds for the Central Universities(2042022dx0003)Natural Science Foundation of Wuhan(2024040701010031).
文摘T-cell immunoglobulin mucin family member-1(TIM-1,also known as HAVCR1/KIM-1)is a transmembrane glycoprotein that has been reported to act as an entry receptor for multiple flaviviruses including Zika virus(ZIKV).The post-translational regulation of TIM-1 and its effects on ZIKV infection are unclear.In this study,we identified the membrane-associated RING-CH-type finger(MARCH)E3 ubiquitin ligase family members MARCH2 and MARCH3 as critical negative regulators of TIM-1 under physiological conditions.MARCH2 and MARCH3 associate with TIM-1 and mediate its K48-linked polyubiquitination at K338 and K346 respectively,leading to subsequent proteasomal degradation.While deficiency of either MARCH2 or MARCH3 modestly increases TIM-1 levels and enhances ZIKV infectivity,double knockout of MARCH2/3 has a more dramatic effect.Double knockout of MARCH2/3 increased ZIKV infectivity in wild-type but not TIM-1 knockout cells,and reconstitution of TIM-1^(K338R/K346R) into TIM-1-deficient cells increases ZIKV infectivity to a higher degree than reconstitution with wild-type TIM-1.Knockout of either MARCH2 or MARCH3 increased ZIKV infectivity and pathogenesis in mice,whereas double knockout of MARCH2/3 has a more dramatic effect.These findings suggest that MARCH2 and MARCH3 target TIM-1 for K48-linked polyubiquitination and proteasomal degradation,thereby acting as redundant host restriction factors to limit ZIKV infection and pathogenesis.
基金This work was supported by grants from the National Natural Science Foundation of China(32188101,31830024 and 32070775)the CAMS Innovation Fund for Medical Sciences(2019-I2M-5-071)the Fundamental Research Funds for the Central Universities.
文摘Interleukin 5(IL-5)plays crucial roles in type 2-high asthma by mediating eosinophil maturation,activation,chemotaxis and survival.Inhibition of IL-5 signaling is considered a strategy for asthma treatment.Here,we identified MARCH2 and MARCH3 as critical negative regulators of IL-5-triggered signaling.MARCH2 and MARCH3 associate with the IL-5 receptorαchain(IL-5Rα)and mediate its K27-linked polyubiquitination at K379 and K383,respectively,and its subsequent lysosomal degradation.Deficiency of MARCH2 or MARCH3 modestly increases the level of IL-5Rαand enhances IL-5-induced signaling,whereas double knockout of MARCH2/3 has a more dramatic effect.March2/3 double knockout markedly increases the proportions of eosinophils in the bone marrow and peripheral blood in mice.Double knockout of March2/3 aggravates ovalbumin(OVA)-induced eosinophilia and causes increased inflammatory cell infiltration,peribronchial mucus secretion and production of Th2 cytokines.Neutralization of Il-5 attenuates OVA-induced airway inflammation and the enhanced effects of March2/3 double deficiency.These findings suggest that MARCH2 and MARCH3 play redundant roles in targeting IL-5Rαfor degradation and negatively regulating allergic airway inflammation.
