Aim： To determine the possible roles of the t-complex testis expressed gene 5 （Tctex5） on sperm functions, the fulllength sequence of mRNA was studied and compared in the testis between the normal wild-type and the...Aim： To determine the possible roles of the t-complex testis expressed gene 5 （Tctex5） on sperm functions, the fulllength sequence of mRNA was studied and compared in the testis between the normal wild-type and the sterile t-haplotype mutant mice. Methods： We applied rapid amplification of cDNA ends, Northern blot and reverse transcription polymerase chain reaction to analyze the full length of Tctex5 mRNAs isolated from testes of the wild-type and the t-haplotype mice. Reverse transcription polymerase chain reaction was used to semi-quantitatively compare expression of Tctex5 transcripts in the 16 tissues and 9.5 day stage embryos in the wild-type mice. E-translation was applied to estimate the amino acid sequences. Results： One long and one short transcript of Tctex5 mRNA were discovered in mouse testis of wild-type （Tctex5^long-＋ and Tctex5^short-＋） and t-haplotype （Tctex5^long-＋ and Tctex5^short-＋） mice, respectively. Being enhanced only in the testis, Tctex5^long-＋ had 17 point mutations and one 15-bp-deletion in the exon 1 region, comparing with the Tctex5^long-＋, whereas the Tctex5^short-＋ was similar to the Tctex5^short-＋. The short isoforms of Tctex5 mRNAs in the two models encoded exactly the same peptides, but the long isoforms did not. The estimated peptide encoded by Tctex5^long-＋ had significant mutations on putative sites of phosphorylation and PP1 binding. Conclusion： We established that mutations that occur in the Tctex5 long transcript of the t-haplotype mice are important for normal sperm function, whereas the short transcript of Tctex5 might have a conserved function among different tissues. （Asian J Androl 2008 Mar; 10： 219-226）展开更多
The cytosolic chaperonin T-complex protein 1-ring complex(TRiC)or chaperonin containing T-complex protein 1(CCT)is essential in de novo folding of approximately 10%of the eukaryotic,newly translated polypeptides as we...The cytosolic chaperonin T-complex protein 1-ring complex(TRiC)or chaperonin containing T-complex protein 1(CCT)is essential in de novo folding of approximately 10%of the eukaryotic,newly translated polypeptides as well as misfolded proteins.There is a close link between the TRiC/CCT cytosolic chaperonin and neurodegenerative diseases(Lopez et al.,2015).A lot of research suggests that CCT plays neuroprotective roles in neurodegenerative diseases including Huntington’s disease(Lopez et al.,2015).Either overexpression of a single or all eight subunits(CCT1-8)or treatment of the substrate-binding apical domain of yeast CCT1(ApiCCT1)prevented mutant Huntingtin aggregation and improved cellular and neuronal functions(Zhao et al.,2016).Importantly,our recent study has demonstrated that both CCT and ApiCCT could reduce mutant Huntingtin level and enhance both anterograde and retrograde axonal transport of brain-derived neurotrophic factor.These results led to restoration of the trophic status of striatal neurons from a bacterial artificial chromosome transgenic mouse model of Huntington’s disease(Zhao et al.,2016).Axonal transport is regulated by many factors including microtubule-associated protein tau,which promotes tubulin polymerization and stabilizes microtubules.Impaired interaction between tau and microtubules plays a vital role in the pathogenesis of multiple neurodegenerative diseases(Wang and Mandelkow,2016).Interestingly,tau phosphorylation is also observed in brains of several Huntington’s disease mouse models and Huntington’s disease patients(Gratuze et al.,2016).In a recent study,we explored if CCT subunit has any effect on axonal transport in a tau-dependent pathway(Chen et al.,2018b).We focused on the retrograde axonal transport of brain-derived neurotrophic factor,as neurotrophic factor-mediated signaling in the form of signaling endosome is essential in both the developing and the mature nervous system and dysregulation of trafficking of neurotrophic factors is tightly linked to disorders of the nervous system(Chen et al.,2018a).We found that the expression of a single CCT subunit(CCT5)significantly promoted retrograde axonal transport of brain-derived neurotrophic factor in primary cortical neurons.Mechanically,CCT regulated the level of cyclin-dependent kinase 5(CDK5)/p35/p25 and,subsequently contributed to CCT-induced tau phosphorylation,which induced detachment of tau from microtubules(Chen et al.,2018b)(Figure 1).展开更多
文摘Aim： To determine the possible roles of the t-complex testis expressed gene 5 （Tctex5） on sperm functions, the fulllength sequence of mRNA was studied and compared in the testis between the normal wild-type and the sterile t-haplotype mutant mice. Methods： We applied rapid amplification of cDNA ends, Northern blot and reverse transcription polymerase chain reaction to analyze the full length of Tctex5 mRNAs isolated from testes of the wild-type and the t-haplotype mice. Reverse transcription polymerase chain reaction was used to semi-quantitatively compare expression of Tctex5 transcripts in the 16 tissues and 9.5 day stage embryos in the wild-type mice. E-translation was applied to estimate the amino acid sequences. Results： One long and one short transcript of Tctex5 mRNA were discovered in mouse testis of wild-type （Tctex5^long-＋ and Tctex5^short-＋） and t-haplotype （Tctex5^long-＋ and Tctex5^short-＋） mice, respectively. Being enhanced only in the testis, Tctex5^long-＋ had 17 point mutations and one 15-bp-deletion in the exon 1 region, comparing with the Tctex5^long-＋, whereas the Tctex5^short-＋ was similar to the Tctex5^short-＋. The short isoforms of Tctex5 mRNAs in the two models encoded exactly the same peptides, but the long isoforms did not. The estimated peptide encoded by Tctex5^long-＋ had significant mutations on putative sites of phosphorylation and PP1 binding. Conclusion： We established that mutations that occur in the Tctex5 long transcript of the t-haplotype mice are important for normal sperm function, whereas the short transcript of Tctex5 might have a conserved function among different tissues. （Asian J Androl 2008 Mar; 10： 219-226）
文摘The cytosolic chaperonin T-complex protein 1-ring complex(TRiC)or chaperonin containing T-complex protein 1(CCT)is essential in de novo folding of approximately 10%of the eukaryotic,newly translated polypeptides as well as misfolded proteins.There is a close link between the TRiC/CCT cytosolic chaperonin and neurodegenerative diseases(Lopez et al.,2015).A lot of research suggests that CCT plays neuroprotective roles in neurodegenerative diseases including Huntington’s disease(Lopez et al.,2015).Either overexpression of a single or all eight subunits(CCT1-8)or treatment of the substrate-binding apical domain of yeast CCT1(ApiCCT1)prevented mutant Huntingtin aggregation and improved cellular and neuronal functions(Zhao et al.,2016).Importantly,our recent study has demonstrated that both CCT and ApiCCT could reduce mutant Huntingtin level and enhance both anterograde and retrograde axonal transport of brain-derived neurotrophic factor.These results led to restoration of the trophic status of striatal neurons from a bacterial artificial chromosome transgenic mouse model of Huntington’s disease(Zhao et al.,2016).Axonal transport is regulated by many factors including microtubule-associated protein tau,which promotes tubulin polymerization and stabilizes microtubules.Impaired interaction between tau and microtubules plays a vital role in the pathogenesis of multiple neurodegenerative diseases(Wang and Mandelkow,2016).Interestingly,tau phosphorylation is also observed in brains of several Huntington’s disease mouse models and Huntington’s disease patients(Gratuze et al.,2016).In a recent study,we explored if CCT subunit has any effect on axonal transport in a tau-dependent pathway(Chen et al.,2018b).We focused on the retrograde axonal transport of brain-derived neurotrophic factor,as neurotrophic factor-mediated signaling in the form of signaling endosome is essential in both the developing and the mature nervous system and dysregulation of trafficking of neurotrophic factors is tightly linked to disorders of the nervous system(Chen et al.,2018a).We found that the expression of a single CCT subunit(CCT5)significantly promoted retrograde axonal transport of brain-derived neurotrophic factor in primary cortical neurons.Mechanically,CCT regulated the level of cyclin-dependent kinase 5(CDK5)/p35/p25 and,subsequently contributed to CCT-induced tau phosphorylation,which induced detachment of tau from microtubules(Chen et al.,2018b)(Figure 1).
文摘目的探索加味蜈蚣败毒饮通过影响自噬上游信号通路腺苷酸活化蛋白激酶(adenosine monophosphate activated protein kinase,AMPK)—结节性硬化复合物1/2(tuberous sclerosiscomplex 1/2,TSC1/2)抑制银屑病复发的机制。方法选取30只小鼠,除空白对照组(5只)外,以咪喹莫特二次诱导的方法(每日咪喹莫特乳膏62.5 mg局部涂擦,连续8日后自然恢复30日,再在原皮损消退部位进行8日的二次涂擦)制备复发型银屑病模型小鼠,再将小鼠随机分为模型对照组、西药对照组、中药低、中、高剂量组,每组均为5只小鼠。对各组小鼠进行药物灌胃干预,中药低、中、高剂量组分别灌胃加味蜈蚣败毒饮药液(浓度分别为0.1 g/mL、0.2 g/mL、0.4 g/mL);西药对照组前4天灌胃甲氨蝶呤混悬液(浓度0.003 mg/mL),后4天灌胃生理盐水;模型对照组灌胃等体积生理盐水;空白对照组不做处理。灌胃剂量均为1 mL/20 g体质量,每日2次,连续8天(第47~54天)。灌胃结束后(第54天)取材,取小鼠背部典型皮损组织备用。采用鼠银屑病皮损面积和严重程度指数(mice psoriasis area and severity index,MPASI)对小鼠的皮损状态进行评价;采用脱氧核糖核苷酸末端转移酶介导的dUTP缺口末端标记法(terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling,TUNEL)染色小鼠皮损组织的凋亡细胞;分别采用蛋白印迹法(western blotting,WB)和免疫组化染色法对小鼠皮损组织中AMPKα、TSC1、TSC2蛋白的含量、表达及分布进行检测。结果(1)经咪喹莫特二次诱导,小鼠银屑病样皮损较一次诱导时的出现时间更早(2日<4日),造模完成时其皮损宏观表象更为严重。(2)复发模型药物干预前,其MPASI分值均在24分以上,各剂量中药均能显著改善复发型银屑病模型的MPASI分值,中药高剂量组疗效尤其显著(P<0.05)。(3)药物干预后,复发模型皮损中位于角质层、颗粒层中的凋亡细胞显著增多,细胞凋亡处于主导地位,从而改善皮损肥厚状态。(4)各剂量中药均能通过调控AMPK-TSC1/2信号通路,抑制AMPKα蛋白(P<0.05)和TSC1蛋白(P<0.05)的表达,并促进TSC2蛋白(P<0.05)在复发模型皮损组织中的含量、表达及分布,以此影响银屑病复发的相关机制通路水平,其中以中药高剂量组的作用最为显著(P<0.05),且WB法与免疫组化法检测上述各蛋白的含量及表达结果相互吻合。结论加味蜈蚣败毒饮可能通过调控自噬上游的关键信号通路AMPK-TSC1/2,影响自噬反应对组织驻留记忆T细胞的能量供给,从而抑制细胞在皮损组织中的再激活,降低银屑病的复发。
