Assembly of the TCR/CD3 complex: CD3ϵ/δ and CD3ϵ/γ dimers associate indistinctly with both TCRα and TCRβ chains. Evidence for a double TCR heterodimer model

The TCR/CD3 complex is composed of six subunits which are expressed on the cell surface in a coordinate fashion after assembly in the endoplasmic reticulum (ER). The TCR/CD3 complex is assembled after a series of pairwise interactions involving the formation of dimers of CD3ϵ with either CD3γ or CD3δ. These dimers assemble with TCRα and TCRβ chains, and finally, the CD3ζ homodimer is added to allow export of the full complex from the ER. A model has been proposed suggesting that during assembly the CD3ϵ/CD3γ dimer interacts exclusively with TCRβ and the CD3ϵ/CD3δ dimer with TCRα to form a complex with a single TCRα/β heterodimer. We show in this study, by immunoprecipitation and two-dimensional gel electrophoresis, that in the human T cell line Jurkat as well as in total human thymocytes, this preferential interaction does not occur and instead, the CD3ϵ/CD3γ and CD3ϵ/CD3δ dimers associate with both TCR chains simultaneously and indistinctly. These data are confirmed by the analysis of the TCRα-negative T cell line MOLT-4 in which TCRβ is found separately associated with CD3ϵ/CD3γ and with CD3ϵ/CD3δ dimers. Indirectly, our results support a model of stoichiometry in which two TCRα/β heterodimers are present in a TCR/CD3 complex. Furthermore, immunoprecipitation with anti-CD3γ and anti-CD3δ antibodies from 1 % NP40 and 1 % Brij96 cell lysates showed that these subunits form independent partial complexes which are cross-linked through the CD3ζ homodimer. This suggests that CD3ζ mediates the interaction between both TCRα/β heterodimers contained in the double TCR complex. Further proof for this hypothesis is obtained after analysis of a Jurkat cell transfectant containing a point mutation in the transmembrane domain of TCRβ that impairs the association of CD3ζ. In this mutant cell line, unlike a control line with wild-type TCRβ, the CD3γ- and CD3δ-containing complexes were found completely independent. Altogether, these results support a model of TCR/CD3 assembly and stoichiometry in which two TCR-α/β heterodimers form two hemicomplexes containing either CD3ϵ/γ or CD3ϵ/δ dimers which become associated via the CD3ζ homodimer.     

钙调素对微管组装的调节作用

利用我们建成的钙调素表达可调细胞模型－ＲＣ３细胞，对ＣａＭ高表达时微管组装行为进行了研究，当用生理剂量的地塞米松处理ＲＣ３细胞，细胞内ＣａＭ水平提高，而管蛋白浓度没有变化，造成钙调素／管蛋白比值上升，ＭＴ解聚，但同时加入ＣａＭ拮抗剂三氟拉嗪处理时，则可抑制ＭＴ的解聚，Ｃ３Ｈ１０Ｔ１／２转化细胞ＣａＭ含量的增加是引起ＭＴ解聚的主要因素，ＴＦＰ处理可恢复ＭＴ组装。ＲＣ３细胞ＣａＭ高表达导致ＭＴ解聚的实     

有限内固定结合组合外固定架治疗胫骨多段开放性粉碎性骨折

目的应用有限内固定结合组合外固定架治疗胫骨多段开放性粉碎性骨折,观察其治疗效果。方法胫骨多段开放性粉碎性骨折8例,合并肠破裂1例,硬膜外血肿1例,胫骨骨折采用螺钉、钢丝结合组合外固定架固定,腓骨应用钢板内固定,对治疗结果进行总结分析。结果8例全部随访,均无感染,无关节活动障碍及肢体缩短,骨折均骨性愈合,外固定架使用时间为5~8个月,平均6个月。结论对胫骨多段开放性粉碎性骨折,采用有限内固定结合组合外固定架固定是一种有效的治疗方法。     

Musculoskeletal symptoms among truck assembly workers

BACKGROUND: Concerns were raised about the possibility of a high prevalence of musculoskeletal symptoms in a truck assembly plant. AIM: The aim of this study was to investigate the prevalence of musculoskeletal symptoms in a group of truck assembly workers. METHOD: A cross-sectional study of 461 truck assembly workers was carried out using a modified version of the Nordic questionnaire and the General Health Questionnaire (GHQ12). Employees were further subdivided into three distinct occupational subgroups: skilled line workers (252), bench subassembly workers (108) and material handlers (101). Responses were analysed according to occupational subgroup. RESULTS: Seventy per cent of 461 truck assembly workers responded to the questionnaires. Seventy-nine per cent of respondents had been troubled with musculoskeletal symptoms in the last 12 months. The commonest musculoskeletal symptoms were from the lower back (65%), neck (60%) and shoulders (57%). Musculoskeletal symptoms were related to age, length of service, occupational subgroup and GHQ12 score. CONCLUSION: There was a high reported prevalence of musculoskeletal symptoms in this group of truck assembly workers, with a differing pattern of symptom reporting depending on occupational subgroup. Risk reduction recommendations were made to the site management. A further study investigating the relationship between symptoms and specific hazards is planned.     

Premature processing of mouse mammary tumor virus Gag polyprotein impairs intracellular capsid assembly

Mouse mammary tumor virus (MMTV) is the prototypical member of the Betaretrovirus genus, but the processes of its morphogenesis are poorly characterized. In this report, we describe an unusual intracellular processing of MMTV Gag polyprotein in human 293T cells transiently expressing MMTV from heterologous promoter. The same specific cleavage products of the viral protease were seen for the wild type as well as for nonmyristylated mutant of MMTV Gag polyprotein completely defective in the particle release. Inactivation of the viral protease resulted in more stable Gag polyprotein and in accumulation of intracytoplasmic particles for nonmyristylated Gag. The intracellular processing of nonmyristylated MMTV Gag indicates that protease activation in betaretrovirus can occur independently of budding.     

Functional role of Alix in HIV-1 replication

Retroviral Gag proteins encode small peptide motifs known as late domains that promote the release of virions from infected cells by interacting directly with host cell factors. Three types of retroviral late domains, with core sequences P(T/S)AP, YPX_nL, and PPPY, have been identified. HIV-1 encodes a primary P(T/S)AP-type late domain and an apparently secondary late domain sequence of the YPX_nL type. The P(T/S)AP and YPX_nL motifs interact with the endosomal sorting factors Tsg101 and Alix, respectively. Although biochemical and structural studies support a direct binding between HIV-1 p6 and Alix, the physiological role of Alix in HIV-1 biology remains undefined. To elucidate the function of the p6–Alix interaction in HIV-1 replication, we introduced a series of mutations in the p6 Alix binding site and evaluated the effects on virus particle production and virus replication in a range of cell types, including physiologically relevant primary T cells and macrophages. We also examined the effects of the Alix binding site mutations on virion morphogenesis and single-cycle virus infectivity. We determined that the p6–Alix interaction plays an important role in HIV-1 replication and observed a particularly severe impact of Alix binding site mutations when they were combined with mutational inactivation of the Tsg101 binding site.     

应用Gibson Assembly法快速构建人组蛋白甲基化转移酶启动子荧光素酶报告载体

目的：快速构建人组蛋白甲基转移酶(enhancer of zeste homolog 2,EZH2)启动子荧光素酶报告载体并验证其活性。方法：因EZH2启动子富含GC,故应用Gibson Assembly方法设计分两段扩增,然后将片段一步法连入pGL3-Basic质粒,构建荧光素酶报告载体,进行酶切鉴定。与pRL-TK内参照质粒共转染C4-2细胞,双荧光素酶分析法检测启动子的活性。结果：经酶切鉴定证实成功构建了EZH2启动子报告载体,双荧光索酶报告基因检测结果显示构建的报告载体具有启动子活性。结论：快速成功构建了具有启动子活性的EZH2启动子荧光素酶报告载体,为进一步研究前列腺癌中EZH2表达调控机制提供必要的实验材料。     

乙型肝炎病毒在丁型肝炎病毒包装中的作用机理研究

用含ＨＢＶ全基因和ＨＢＶ大、中、小分子表面蛋白基因的真核细胞表达质粒（ＣＭＶ－ＨＢＶ、ＣＭＶ－ＬＳ、ＣＭＶ－ＭＳ、ＣＭＶ－Ｓ）分别与含ＨＤＶ ｃＤＮＡ三聚体的重组质粒共转染ＣＨＯ细胞。转染后３天在上述４种转染细胞内及培养上清中均检出了ＨＤＶ ＲＮＡ和ＨＤＡｇ表明上述４种转染细胞的培养上清中均有ＨＤＶ病毒颗粒的包装和分泌。提示：ＨＤＶ病毒的包装可能仅需ＨＢＶ Ｓ 基因及其小分子表面蛋白的辅助。     

Interaction of the adenovirus major core protein precursor, pVII, with the viral DNA packaging machinery

Adenovirus is one of the well-studied double-stranded DNA viruses. However, the mechanisms of its DNA packaging and virion assembly are still not fully understood. One of the unique features of adenovirus is that the unpackaged viral DNA is associated with core protein pVII. Packaging of viral DNA bound with proteins has not been reported from other viruses. To characterize how viral DNA bound with protein pVII is packaged, we performed experiments to see if protein pVII interacts with the known DNA packaging proteins or the packaging sequence. Our results demonstrated that protein pVII interacted with the viral IVa2 and L1 52/55 kDa proteins, which are the known viral DNA packaging proteins. Furthermore, our protein-DNA binding experiments demonstrated that the IVa2 protein mediates the specific interaction with the packaging sequence, whereas protein pVII and the L1 52/55 kDa protein bind to DNA non-specifically. Although the non-specific binding of protein pVII and the L1 52/55 kDa protein do not appear to affect the specific binding of the IVa2 protein to the packaging sequence, and the specific binding of the IVa2 protein does not appear to block the bindings of protein pVII and the L1 52/55 kDa protein to the packaging sequence, the possibility of a cooperative binding among the IVa2 protein, the L1 52/55 kDa protein and protein pVII on the packaging sequence needs to be further determined. In summary, the results indicate that the assembly of the DNA packaging initiation complex may be mediated by the specific interaction of the IVa2 protein with the packaging sequence and other viral proteins, such as protein pVII and the L1 52/55 kDa protein.     

大型综合性医院晨会应用分析和实证研究

目的 为了保证医院的可持续发展及伦理建设,通过分析晨会实施的意义及作用,不断探索创新晨会的形式.方法 采用文献研究法和资料分析法,并结合本院的实际情况进行实证研究.结果 通过一系列的研究发现,我院实施的晨会制度在传播医院文化、加强医风医德建设,传递有效信息、塑造人性化医院,协调解决问题、保证患者利益,培养前瞻能力、兼顾社会效益与经济效益方面发挥了重要作用.结论 通过资源整合与展会制度的实施,为我院创建了一个和谐、公正、公平的医疗环境,医护人员的基本需求得到满足,患者利益得到保证,取得了一定的成绩,但医院持续稳定发展还必须依靠全院员工的共同努力.