EXT1基因1633-26(C→A)突变可能致多发性外生性骨疣

目的研究1例散发多发性外生性骨疣患者的基因突变情况，确定其致病基因。方法采用聚合酶链反应结合DNA直接测序法检测EXT1以及EXT2基因的突变热点区域；并应用错配引物PCR扩增引入酶切位点结合限制性片段长度多态性方法检测和鉴定突变。结果经测序证实在患者EXT1基因的第7内含子3’剪接位点上游26bp处发现一杂合突变，此杂合突变不存在于其表型正常的父母双亲中，是一个新生突变；错配引物扩增与限制性片段长度多态性分析结果表明在150名家系外正常对照者中没有此突变。结论EXT1基因1633-26（C→A）突变可能是导致这个患者发生多发性外生性骨疣的致病突变。     

Identification of a Thymidylate Synthase Gene within the Genome of Chilo Iridescent Virus

The thymidylate synthase (TS, EC 2.1.1.45) is essential for the de novo synthesis of dTMP in pro- and eucaryotic organisms. Consequently it plays a major role in the replication of the DNA genome of a cell or a DNA virus. The gene encoding the TS of Chilo iridescent virus (CIV) was identified by nucleotide sequence analysis of the viral genome and was mapped within the EcoRI CIV DNA fragments G and R. Computer assisted analysis of the DNA nucleotide sequence between the genome coordinates 0.482 and 0.489 revealed an open reading frame (ORF) of 885 nucleotides. This ORF was found to encode a polypeptide of 295 amino acid residues (33.9 kDa) that showed significant homologies to known TS of different species including mammals, plants, fungi, protozoa, bacteria, and DNA viruses. The highest amino acid homologies were found between the CIV-TS and the TS of herpesvirus ateles (54.0%), Saccharomyces cerevisiae (51.8%), herpesvirus saimiri (51.0%), rhesus monkey rhadinovirus (50.7%), mouse (50.5%), rat (50.2%), varicella-zoster virus (50.2%), equine herpesvirus 2 (50.0%), and the human TS (48.4%). The CIV-TS contains six amino acid domains that are highly conserved in the TS of other species. Within these domains the major amino acid residues are present for which a functional role has been reported. The CIV-TS was found to be more closely related to the TS of eucaryotes than to the TS of procaryotes indicating the phylogenetic origin of the CIV-TS gene. The identification of a TS gene in the genome of CIV is the first report of a viral TS that is not encoded by a herpesvirus or a bacteriophage.     

Complete Nucleotide Sequence of the Rice Tungro Spherical Virus Genome of the Highly Virulent Strain Vt6

The complete nucleotide sequence of rice tungro spherical virus (RTSV) strain Vt6, originally from Mindanao, the Philippines, with higher virulence to resistant rice cultivars, was determined and compared with the published sequence for the Philippine-type strain A (RTSV-A-Shen). It was reported that RTSV-A was not able to infect a rice resistant cultivar TKM 6 (10). RTSV-Vt6 and RTSV-A-Shen share 90% and 95% homology at nucleotide and amino-acid levels, respectively. The N-terminal leader sequence of RTSV-Vt6 contained a 39-amino acids-region (positions 65 to 103) which was totally different from that of RTSV-A-Shen; the difference resulted from frame shifting by nucleotide insertions and deletions. To confirm the amino-acid sequence differences of the leader polypeptide, the same region was cloned and sequenced using a newly obtained variant of RTSV-type 6, which had been collected in the field of IRRI, and seven field isolates from Mindanao, the Philippines. Since all the sequences of the target region are identical to that of the Vt6 leader polypeptide, the sequence difference in the leader region seems not to correlate with the virulence of Vt6.     

Hantaviruses in Central South America: phylogenetic analysis of the S segment from HPS cases in Paraná, Brazil

We sequenced the complete S segments of hantaviruses detected from 12 HPS patients living in southern of Brazil. Samples were obtained from patients diagnosed in different years, in distinct areas, and with a broad spectrum of clinical signs. Despite these differences, all the S proteins of hantavirus from Paraná were identical, except for one amino acid substitution. Phylogenetic analyses of the complete S segment nucleotide and amino acid sequences indicated that hantaviruses from Paraná form a distinct clade from those circulating in South and North America. Other hantaviruses from Brazil were not placed in the same clade. The Oligoryzomys nigripes-associated strains ITA37 and ITA38 from Paraguay were found to belong to the same clade as the hantaviruses from Paraná. Paraguay and Paraná state are located at the same latitude and some ecosystems are similar in both places. The geographic position and common rodent hosts could explain this phylogenetic relationship.     

Sequence Analysis of the Washington/1964 Strain of Human Parainfluenza Virus Type 1 (HPIV1) and Recovery and Characterization of Wild-Type Recombinant HPIV1 Produced by Reverse Genetics

A complete consensus sequence was determined for the genomic RNA of human parainfluenza virus type 1 (HPIV1) strain Washington/20993/1964 (HPIV1 WASH/64), a clinical isolate that previously was shown to be virulent in adults. The sequence exhibited a high degree of relatedness to both Sendai virus, a PIV1 virus recovered from mice, and human PIV3 (HPIV3) with regard to cis-acting regulatory regions and protein-coding sequences. This consensus sequence was used to generate a full-length antigenomic cDNA and to recover a recombinant wild-type HPIV1 (rHPIV1). Interestingly, the rHPIV1 could be rescued from full-length antigenomic rHPIV1 cDNA using HPIV3 support plasmids, HPIV1 support plasmids, or a mixture thereof. The replication of rHPIV1 in vitro and in the respiratory tract of hamsters was similar to that of its biologically derived parent virus. The similar biological properties of rHPIV1 and HPIV1 WASH/64 in vitro and in vivo, together with the previous demonstration of the virulence of this specific isolate in humans, authenticates the rHPIV1 sequence as that of a wild-type virus. This rHPIV1 can now be used to study the biological properties of HPIV1 and as a substrate to introduce attenuating mutations for the generation of live-attenuated HPIV1 vaccine candidates.An erratum to this article can be found at     

Guanine nucleotide exchange factors of the cytohesin family and their roles in signal transduction

Summary:  Members of the cytohesin protein family, a group of guanine nucleotide exchange factors for adenosine diphosphate ribosylation factor (ARF) guanosine triphosphatases, have recently emerged as important regulators of signal transduction in vertebrate and invertebrate biology. These proteins share a modular domain structure, comprising carboxy-terminal membrane recruitment elements, a Sec7 homology effector domain, and an amino-terminal coiled-coil domain that serve as a platform for their integration into larger signaling complexes. Although these proteins have a highly similar overall build, their individual biological functions appear to be at least partly specific. Cytohesin-1 had been identified as a regulator of β2 integrin inside-out regulation in immune cells and was subsequently shown to be involved in mitogen-associated protein kinase signaling in tumor cell proliferation as well as in T-helper cell activation and differentiation. Cytohesin-3, which had been discovered to be strongly associated with T-cell anergy, was very recently described as an essential component of insulin signal transduction in Drosophila and in human and murine liver cells. Future work will aim to dissect the mechanistic details of the modes of action of the cytohesins as well as to define the precise roles of these versatile proteins in vertebrates at the genetic level.     

四引物突变特异性扩增系统法检测巨噬细胞移动抑制因子基因-173位点单核苷酸多态性分布

目的探讨中国浙江地区汉族人群巨噬细胞移动抑制因子（macmphage migration inhibitory factor，MW）基因-173位点单核苷酸多态性（single nucleotide polymorphism，SNP）分布。方法收集浙江地区142名无血缘关系健康个体的静脉血，提取DNA，分别应用四引物突变特异性扩增系统（amplification refractory mutation sysntem，ARMS）法和限制性片段长度多态性．PCR方法对MIF基因-173位点SNP多态性进行分型，并将PCR产物克隆及测序鉴定。结果MIF基因-173位点检测到3种基因型，其基因型分布皆符合Hardy-Weinberg平衡定律。四引物ARMS法和限制性片段长度多态性-PCR两种方法结果完全一致。统计分析显示，中国汉族人MIF基因-173位点等位基因和基因型频率分布与欧洲白人差异有统计学意义（P〈0．01），与日本人群的差异无统计学意义（P〉0．05）。结论四引物ARMS法是一种准确、快速和经济的SNP测定方法。MIF基因-173位点等位基因频率分布具有种族的差异性。     

A cytoplasmic component of pyridine nucleotide fluorescence in rat diaphragm: evidence from comparisons with flavoprotein fluorescence

Pyridine nucleotide (PN) and flavoprotein (Fp) fluorescence were monitored in the isolated intact rat diaphragm. A substantial increase in PN fluorescence occurred when N₂ replaced O₂ in glucose medium. This response was much reduced in pyruvate medium and/or by pretreatment with iodoacetic acid (IAA). The anaerobic levels of Fp fluorescence were less affected by substrate and IAA. Substitution of glucose by pyruvate did not alter the PN fluorescence of the resting aerobic tissue, but increased Fp fluorescence. After a tetanus with glucose present the PN of the anaerobic muscle, but not the Fp underwent a substantial transient oxidation. This oxidation was absent in pyruvate medium. It is concluded that a cytoplasmic component of the PN fluorescence is present in skeletal muscle. The levels of Fp fluorescence in the resting and contracting aerobic tissue supplied with pyruvate suggest that the resting tissue respiration was ADP limited. On this basis the level of PN fluorescence in the aerobic resting state was less than expected; the source of the PN fluorescence was both mitochondrial and cytoplasmic.