目的:探讨羧基端缺失了30个氨基酸的乙型肝炎病毒X蛋白(hepatitis B virus X protein,HBx)突变体ΔHBx和野生型HBx对肝癌细胞Huh7生物学行为的影响。方法:脂质体介导pcDNA3ΔHBx和pcDNA3HBx重组体转染人HBV(-)的肝癌细胞Huh7。PCR扩增Ne...目的:探讨羧基端缺失了30个氨基酸的乙型肝炎病毒X蛋白(hepatitis B virus X protein,HBx)突变体ΔHBx和野生型HBx对肝癌细胞Huh7生物学行为的影响。方法:脂质体介导pcDNA3ΔHBx和pcDNA3HBx重组体转染人HBV(-)的肝癌细胞Huh7。PCR扩增Neo基因及Western blot检测转染重组体。运用细胞生长曲线绘制、平板克隆形成、流式细胞仪和裸鼠成瘤实验对转染pcDNA3ΔHBx、pcDNA3HBx和pcDNA3的细胞生物学活性进行检测。结果:pcDNA3ΔHBx组细胞生长速度较pcDNA3HBx组和pcDNA3组减慢,其倍增时间延长;pcDNA3ΔHBx组克隆形成率较pcDNA3HBx组和pcDNA3组下降;细胞周期检测显示pcDNA3ΔHBx表达使Huh7细胞G0/G1期→S期的进程明显减慢。结论:HBx羧基端缺失了30个氨基酸的突变体比野生型HBx具有明显抑制细胞增殖的能力,提示该区域对于HBx功能发挥起着至关重要的作用。展开更多
Interleukin-17 （IL-17 or IL-17A） production is a hallmark of TH17 cells, a new unique lineage of CD4^＋ T lymphocytes contributing to the pathogenesis of multiple autoimmune and inflammatory diseases. IL-17 receptor...Interleukin-17 （IL-17 or IL-17A） production is a hallmark of TH17 cells, a new unique lineage of CD4^＋ T lymphocytes contributing to the pathogenesis of multiple autoimmune and inflammatory diseases. IL-17 receptor （IL-17R or IL-17RA） is essential for IL-17 biological activity. Emerging data suggest that the formation of a heteromeric and/or homomeric receptor complex is required for IL-17 signaling. Here we show that the orphan receptor IL-17RD （Sef, similar expression to FGF genes or IL-17RLM） is associated and colocalized with IL-17R. Importantly, IL-17RD mediates IL-17 signaling, as evaluated using a luciferase reporter driven by the native promoter of 24p3, an IL-17 target gene. In addition, an IL-17RD mutant lacking the intraeellnlar domain dominant-negatively suppresses IL-17R- mediated IL-17 signaling. Moreover, IL-17RD as well as IL-17R is associated with TRAF6, an IL-17R downstream molecule. These results indicate that IL-17RD is a part of the IL-17 receptor signaling complex, therefore providing novel evidence for IL-17 signaling through a heteromeric and/or homomeric receptor complex.展开更多
The great majority of genetic disorders are caused by defects in the nuclear genome. However, some significant diseases are the result of mitochondrial mutations. Because of the unique features of the mitochondria, th...The great majority of genetic disorders are caused by defects in the nuclear genome. However, some significant diseases are the result of mitochondrial mutations. Because of the unique features of the mitochondria, these diseases display characteristic modes of inheritance and a large degree of phenotypic variability. Recent studies have suggested that mitochondrial dysfunction plays a central role in a wide range of age-related disorders and various forms of cancer.展开更多
Glycidyl methacrylate (GMA)is a recently recognized mutagen. In order to explore the mutagenicity and mechanism of GMA, plasmid PBR322 was used for in vitro binding, mutant screening, restriction enzyme mapping,and DN...Glycidyl methacrylate (GMA)is a recently recognized mutagen. In order to explore the mutagenicity and mechanism of GMA, plasmid PBR322 was used for in vitro binding, mutant screening, restriction enzyme mapping,and DNA sequencing. To explore the mechamism by which an initial premutational event is converted into a stable heritable mutation, pBR322 and GMA-bound pBR322 were transformed into E. coli HB101 , and the following results were obtained : 1) GMA-bound PBR322 induced phenotype changes in competent cells. Two stable and heritable mutants were isolated (Ap￣RTc￣S and Ap￣STc￣R). 2) When restriction enzyme mapping was used to analyze the mutant Ap￣RTc￣S , four of seven maps showed changes, but no large DNA insertion or deletion were observed.3) The frequency of deletion and insertion forms counted about 10%. Sequence specificity and hot spot regions were evident in the sequence analysis of mutated plasmid. The above results indicate that the premutagenic lesions of plasmid induced by GMA can be converted into point mutations in vivo.展开更多
Duchenne muscular dystrophy(DMD)and Becker muscular dystrophy(BMD)are caused by mutations in the DMD gene.The aim of this study is to identify pathogenic DMD variants in probands and reduce the risk of recurrence of t...Duchenne muscular dystrophy(DMD)and Becker muscular dystrophy(BMD)are caused by mutations in the DMD gene.The aim of this study is to identify pathogenic DMD variants in probands and reduce the risk of recurrence of the disease in affected families.Variations in 100 unrelated DMD/BMD patients were detected by multiplex ligation-dependent probe amplification(MLPA)and next-generation sequencing(NGS).Pathogenic variants in DMD were successfully identified in all cases,and 11 of them were novel.The most common mutations were intragenic deletions(69%),with two hotspots located in the 5'end(exons 2–19)and the central of the DMD gene(exons 45–55),while point mutations were observed in 22%patients.Further,c.1149+1G>A and c.1150?2A>G were confirmed by hybrid minigene splicing assay(HMSA).This two splice site mutations would lead to two aberrant DMD isoforms which give rise to severely truncated protein.Therefore,the clinical use of MLPA,NGS,and HMSA is an effective strategy to identify variants.Importantly,eight embryos were terminated pregnancies according to prenatal diagnosis and a healthy boy was successfully delivered by preimplantation genetic diagnosis(PGD).Early and accurate genetic diagnosis is essential for prenatal diagnosis/PGD to reduce the risk of recurrence of DMD in affected families.展开更多
Editor＇s comments The human mutation rate how often new changes appear in the DNA--is fundamental to understanding many aspects of medical genetics and human evolutionary genetics. But it is low, and has therefore b...Editor＇s comments The human mutation rate how often new changes appear in the DNA--is fundamental to understanding many aspects of medical genetics and human evolutionary genetics. But it is low, and has therefore been difficult to measure. In the past, scientists could only estimate it approximately, either by observing how often mutant phenotypes appeared, or by comparison of humans and closely related species, such as chimpanzee, where many mutations could accumulate but the time period was uncertain. Now, a new study supported by the NSFC in China and The Royal Society in the UK reports the first direct measurement of the human mutation rate at the individual letters （ nucleotides or bases） of DNA. This was possible because new （ next ）-generation sequencing technology is much more powerful than the methods available previously. The work was published in the lead- ing journal Currerzt Biology on 15th September 2009. The results were reported in the news by Nature, Science and the BBC , as well as in more than 20 Chinese newspapers and radio stations after the work first appeared online on 27th August. It was also one of the research highlights in Nature on 3rd September, which commented ＂ This direct measurement of the human mutation rate should help researchers to refine evolutionary dating and better understand the source of genetic disease＇. From the work, researchers could estimate that everyone has around 200 new mutations in their genome ; as the authors said, ＂we are all mutants＂. The ability to reliably measure rates of DNA mutation means we can begin to ask how mutation rates vary between different regions of the genome and perhaps also between different individuals.展开更多
文摘目的:探讨羧基端缺失了30个氨基酸的乙型肝炎病毒X蛋白(hepatitis B virus X protein,HBx)突变体ΔHBx和野生型HBx对肝癌细胞Huh7生物学行为的影响。方法:脂质体介导pcDNA3ΔHBx和pcDNA3HBx重组体转染人HBV(-)的肝癌细胞Huh7。PCR扩增Neo基因及Western blot检测转染重组体。运用细胞生长曲线绘制、平板克隆形成、流式细胞仪和裸鼠成瘤实验对转染pcDNA3ΔHBx、pcDNA3HBx和pcDNA3的细胞生物学活性进行检测。结果:pcDNA3ΔHBx组细胞生长速度较pcDNA3HBx组和pcDNA3组减慢,其倍增时间延长;pcDNA3ΔHBx组克隆形成率较pcDNA3HBx组和pcDNA3组下降;细胞周期检测显示pcDNA3ΔHBx表达使Huh7细胞G0/G1期→S期的进程明显减慢。结论:HBx羧基端缺失了30个氨基酸的突变体比野生型HBx具有明显抑制细胞增殖的能力,提示该区域对于HBx功能发挥起着至关重要的作用。
文摘Interleukin-17 （IL-17 or IL-17A） production is a hallmark of TH17 cells, a new unique lineage of CD4^＋ T lymphocytes contributing to the pathogenesis of multiple autoimmune and inflammatory diseases. IL-17 receptor （IL-17R or IL-17RA） is essential for IL-17 biological activity. Emerging data suggest that the formation of a heteromeric and/or homomeric receptor complex is required for IL-17 signaling. Here we show that the orphan receptor IL-17RD （Sef, similar expression to FGF genes or IL-17RLM） is associated and colocalized with IL-17R. Importantly, IL-17RD mediates IL-17 signaling, as evaluated using a luciferase reporter driven by the native promoter of 24p3, an IL-17 target gene. In addition, an IL-17RD mutant lacking the intraeellnlar domain dominant-negatively suppresses IL-17R- mediated IL-17 signaling. Moreover, IL-17RD as well as IL-17R is associated with TRAF6, an IL-17R downstream molecule. These results indicate that IL-17RD is a part of the IL-17 receptor signaling complex, therefore providing novel evidence for IL-17 signaling through a heteromeric and/or homomeric receptor complex.
基金Project supported by the Natural Science Foundation of Zhejiang Province (No. 2007C33049), China
文摘The great majority of genetic disorders are caused by defects in the nuclear genome. However, some significant diseases are the result of mitochondrial mutations. Because of the unique features of the mitochondria, these diseases display characteristic modes of inheritance and a large degree of phenotypic variability. Recent studies have suggested that mitochondrial dysfunction plays a central role in a wide range of age-related disorders and various forms of cancer.
文摘Glycidyl methacrylate (GMA)is a recently recognized mutagen. In order to explore the mutagenicity and mechanism of GMA, plasmid PBR322 was used for in vitro binding, mutant screening, restriction enzyme mapping,and DNA sequencing. To explore the mechamism by which an initial premutational event is converted into a stable heritable mutation, pBR322 and GMA-bound pBR322 were transformed into E. coli HB101 , and the following results were obtained : 1) GMA-bound PBR322 induced phenotype changes in competent cells. Two stable and heritable mutants were isolated (Ap￣RTc￣S and Ap￣STc￣R). 2) When restriction enzyme mapping was used to analyze the mutant Ap￣RTc￣S , four of seven maps showed changes, but no large DNA insertion or deletion were observed.3) The frequency of deletion and insertion forms counted about 10%. Sequence specificity and hot spot regions were evident in the sequence analysis of mutated plasmid. The above results indicate that the premutagenic lesions of plasmid induced by GMA can be converted into point mutations in vivo.
基金the National Key Research and Development Program of China(No.2016YFC1000703)the Medicine and Health Science and Technology Plan Projects in Zhejiang Province(No.2014KYA246)the National Natural Science Foundation of China(Nos.81801441 and 81300532)
文摘Duchenne muscular dystrophy(DMD)and Becker muscular dystrophy(BMD)are caused by mutations in the DMD gene.The aim of this study is to identify pathogenic DMD variants in probands and reduce the risk of recurrence of the disease in affected families.Variations in 100 unrelated DMD/BMD patients were detected by multiplex ligation-dependent probe amplification(MLPA)and next-generation sequencing(NGS).Pathogenic variants in DMD were successfully identified in all cases,and 11 of them were novel.The most common mutations were intragenic deletions(69%),with two hotspots located in the 5'end(exons 2–19)and the central of the DMD gene(exons 45–55),while point mutations were observed in 22%patients.Further,c.1149+1G>A and c.1150?2A>G were confirmed by hybrid minigene splicing assay(HMSA).This two splice site mutations would lead to two aberrant DMD isoforms which give rise to severely truncated protein.Therefore,the clinical use of MLPA,NGS,and HMSA is an effective strategy to identify variants.Importantly,eight embryos were terminated pregnancies according to prenatal diagnosis and a healthy boy was successfully delivered by preimplantation genetic diagnosis(PGD).Early and accurate genetic diagnosis is essential for prenatal diagnosis/PGD to reduce the risk of recurrence of DMD in affected families.
文摘Editor＇s comments The human mutation rate how often new changes appear in the DNA--is fundamental to understanding many aspects of medical genetics and human evolutionary genetics. But it is low, and has therefore been difficult to measure. In the past, scientists could only estimate it approximately, either by observing how often mutant phenotypes appeared, or by comparison of humans and closely related species, such as chimpanzee, where many mutations could accumulate but the time period was uncertain. Now, a new study supported by the NSFC in China and The Royal Society in the UK reports the first direct measurement of the human mutation rate at the individual letters （ nucleotides or bases） of DNA. This was possible because new （ next ）-generation sequencing technology is much more powerful than the methods available previously. The work was published in the lead- ing journal Currerzt Biology on 15th September 2009. The results were reported in the news by Nature, Science and the BBC , as well as in more than 20 Chinese newspapers and radio stations after the work first appeared online on 27th August. It was also one of the research highlights in Nature on 3rd September, which commented ＂ This direct measurement of the human mutation rate should help researchers to refine evolutionary dating and better understand the source of genetic disease＇. From the work, researchers could estimate that everyone has around 200 new mutations in their genome ; as the authors said, ＂we are all mutants＂. The ability to reliably measure rates of DNA mutation means we can begin to ask how mutation rates vary between different regions of the genome and perhaps also between different individuals.
