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    Proceedings of the National Academy of Sciences of the United States of America

  • 中文名称: 美利坚合众国国家科学院院刊
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  • ISSN: 0027-8424
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  • 机译 人类催乳素/生长激素家族成员完整和N末端片段对血管生成的反作用:调节血管生成的有效机制。
    摘要:Angiogenesis, the process of development of a new microvasculature, is regulated by a balance of positive and negative factors. We show both in vivo and in vitro that the members of the human prolactin/growth hormone family, i.e., human prolactin, human growth hormone, human placental lactogen, and human growth hormone variant are angiogcnic whereas their respective 16-kDa N-terminal fragments are antiangiogenic. The opposite actions are regulated in part via activation or inhibition of mitogen-activated protein kinase signaling pathway. In addition, the N-terminal fragments stimulate expression of type I plasminogen activator inhibitor whereas the intact molecules have no effect, an observation consistent with the fragments acting via separatc receptors. The concept that a single molecule eucodes both angiogenic and antiangiogenic peptides represents an efficient model for regulating the balance of positive and negative factors con- trolling angiogenesis. This hypothesis has potential physio- logical importance for the control of the vascular connection between the fetal and maternal circulatious in the placenta, where human prolactin, human placental lactogen, and hu- man growth hormone variant are expressed.
  • 机译 受体诱导的涂料聚合
    摘要:Coatomer, the coat protein complex of COPl vesicles, is involved in the budding of these vesicles, but the underlying mechanism is unknown. Toward a better under- standing of this process, the interaction between coatomer and the cytoplasmic domain of a major transmembrane protein of COPI vesicles, p23, was studied. Interaction of coatomer with this peptide domain results in a conformational change and polymerization of the complex in viyro. This changed confor- mation also is observed in vivo, i.e., on the surface of authentic, isolated COPI vesicles. An average of four peptides was found associated with one coatomer complex after polymerization. Based on these results, we propose a mechanism by which the induced conformational change of coatomer results in its polymerization, and thus drives formation of the bud on tbe Golgi membrane during biogenesis of a COPI vesicle.
  • 机译 准等价原理和欧几里得几何控制着丙酮酸脱氢酶复合物的立方和十二面体核的组装
    摘要:The pyruvate dehydrogenase multienzyme comPLex (Mr of 5-10 million) is assembled around a structural core formed of multiple copies of dihydrolipoyl acetyltrans- ferase (EZp), which exhibits the shape of either a cube or a dodecahedron, depending on the source. The crystal struc- tures of the 60-meric dihydrolipoyl acyltransferase cores of Bacillus stearothermophilus and Enterococcus faecalis pyruvate dehydrogenase complexes were determined and revealed a remarkably bollow dodecahedron with an outer diameter of apporox. = 237 A 12 large openings of approx. 52 A diameter across the fivefold axes, and an inner cavity with a diamcter of approx. = 118 A. Comparison of cubic and dodecahedral EZP assemblies shows that combining the principles of quasi-equivalence formulated by Caspar and Klug [Caspar, D. L. & KIug, A. (1962) Coid Spring Harkor Symp. Quant. Biol. 27, 1-4] with strict Euclid- ean geometric considerations results in predictions of the major features of the E2P dodecahedron matching the ob- served features almost exactly.
  • 机译 中层密度电流与大陆斜坡底部的分离
    • 作者:M.E. STERN;
    • 刊名:Proceedings of the National Academy of Sciences of the United States of America
    • 1999年第4期
    摘要:The high-salinity water flowing out of the Mediterranean Sea descends to mid depths in the density- stratified ocean, continues as a narrow jet along the lberian continental slope, and intermittently detaches large-scale eddies (called ``Meddies''). This process is important because it maintains the relatively high mean salinity of a major water mass (the ``Mediterranean intermediate Water'') in the North Atlantic. Our simplified model of this jet consists of a moving Iayer with intermediate density p2 sandwiched between mo- tionless layers of density p1 < p2 and p2 > p2. The inshore (anticyclonic) portion of the midlevel jet (in the ``p2-water'') rests on an inclined bottom (the continental slope), whercas the (cyclonic) offshore portion rests on the density interface of the stagnant deep (p3) Iayer. An inviscid, steady, and finite-amplitude long wave theory is used to show that if the cross-stream topographic slope increases gradually in the downstream direction, then the ``p2-Jet'' is deflected off the bottom slope and onto the upper density interface of the p3 layer. The computed magnitude of this separation effect is such as to produce an essentially free Jet which is removed from the stabilizing influence of the continental topography. It is therefore conjectured that time-dependent effects (ba- reclinic instability) will produce further amplification, caus- ing an eddy to detach seaward from the branch of the jet remaining on the slope.
  • 机译 Hsp90的带电区域调节N末端域的功能
    摘要:Hsp90, an abundant heat shock protein that is highly expressed even under physiological conditions, is involved in the folding of key molecules of the cellular signal transduction system such as kinases and steroid receptors. It seems to contain two chaperone sites differing in substrate spccificity. Binding of ATP or the antitumor drug geldana- mycin alters the substrate affinity of the N-terminal chaper- one site, whereas both substances show no influence on the C-terminal one. In wild-type Hsp90 the fragments containing the chaperone sites are connected by a highly charged linker of various lengths in different organisms. As this linker region represents the most striking difference between bacterial and eukaryotic Hsp90s, it may be involved in a gain of function of eukaryotic HsP90s. Here, we have analyzed a fragment of yeast Hsp90 consisting of the N-terminal domain and the charged region (N272) in comparison with the isolated N-terminal domain (N210). We show that the charged region causes an increase in the affinity of the N-terminal domain for nonnative protein and establishes a crosstalk between peptide and ATP binding. Thus, the binding of peptide to N272 decreases its affinity for ATP and geldanamycin, whereas the ATP-binding properties of the monomeric N.terminal domain N210 are not influenced by peptide binding. We propose that the charged region connecting the two chaperone domains plays an im- portant role in regulating chaperone function of Hsp90.
  • 机译 视网膜色素变性GTP酶调节剂RPGR与杆状环GMP磷酸二酯酶的δ亚基相互作用
    摘要:Recently, the retinitis pigmentosa 3 (RP3) gene has been cloned and named retinitis piginentosa GTPase regulator (RPGR). The amino-terminal half of RPGR is homologous to regulator of chromosome condensation (RCCI), the nucleotide exchange factor for the small GTP- binding protein Ran. In a yeast two-hybrid screen we identi- fied the delta subunit of rod cyclic GMP phosphodiesterase (PDES) as interacting with the RCCI-like domain (RLD) of gyGR (RPGR392). The interaction of RPGR with PDFS was conFirmed by pull-down assays and plasmon surface reso- nance. The binding affinity was determined to be 90 nM. Six missense mutations at evolutionary conserved residues within the RLD, which were found in RP3 patients, were analyzed by using the two-hybrid system. All missense mutations showed reduced interaction with PDES. A non-RP3-associated mis- sense substitution outside the RLD, V36F, did not abolish the interaction with PDES. PDES is widely expressed and highly conserved across evolution and is proposed to regulate the membrane insertion or solubilization of prenylated proteins, including the catalytic subunits of the PDE holoenzyme involved in phototransduction and small GTP-binding pro- teins of the Rab family. These results suggest that MGR mutations give rise to retinal dcgeneration by dysregulation of intracellular processes that determine protein localization and protein transport.
  • 机译 神经递质受体锚蛋白gephyrin重建细菌,植物和哺乳动物细胞中的钼辅因子生物沉淀
    摘要:The molybdenum cofactor (Moco), a highly conserved pterin compound complexing molybdenum, is re- quired for the enzymatic activities of all molybdcnum enzymes except nitrogenase. Moco is synthesized by a unique and evolu. tionarily old pathway that requires the activities of at least six gene products. Some of the proteins involved in bacterial, plant and invertebrate Moco biosynthesis show striking homologies to the primary structure of gepbyrin, a polypeptide required for the clustering of inhibitory glycine receptors in postsynaptic mem- branes in the rat central nervous system. Hcre, we show that gephyrin binds with high affinity to molybdopterin, the meta- bolic precursor of Moco. Furthermore, gephyrin expression can reconstitute Moco biosynthesis in Moco-deficient bacteria, a molybdenum-dependent mouse cell line, and a Moco-deficient plant mutant. Conversely, inhibition of gephyrin expression by antisense RNA expression in cultured murine cells reduces their Moco content significantly. These data indicate that in addition to clustering glycine receptors, gepbyrin also is involved in Moco biosynthesis and illustrate the remarkable conservation of its function in Moco biosynthesis throughout phylogeny.
  • 机译 包含插入的谷氨酰胺重复序列的二聚胰凝乳蛋白酶抑制剂2突变体的晶体结构
    摘要:We have constructed mutants of chymotryp- sin inhibitor 2 with short glutamine repeats inserted into its inhibitory loop. These mutants oligomerize when expressed in Escherichia coli. The dimer of a mutant with four glutamines now has been crystallized, and its structure has been solved hy molecular replacement by using the wild-type monomer as a search model. The structure of each half of the dimer is found to be the same as that of the wild-type monomer, except around the glutamine insertion. It was proposed that the components of the oligomers are held together by hydrogen bonds between the main-chain and side-chain amides of the glutamine repeats. Instead, they appear to form by swapping domains on folding in E. coli, and the glutamine repeats connecting the components of the dimers are disordered.
  • 机译 波动环境中的生物多样性和生态系统生产力:保险假说
    • 作者:SHIGEO YACHI; MICHEL LOREAU;
    • 刊名:Proceedings of the National Academy of Sciences of the United States of America
    • 1999年第4期
    摘要:Although the effcct of biodiversity on ecosys- tem functioning has become a major focus in ecology, its significance in a fluctuating environment is still poorly un- derstood. According to the insurance hypothesis, biodiversity insures ecosystems against declines in their functioning be. cause many species provide greater guarantees that some will maintain functioning even if others fail. Here we examine this hypothesis theoretically. We develop a general stochastic dynamic model to assess the effects of species richnes on the expected temporal mean and variance of ecosystem processes such as productivity, based on individual species productivity responses to environmental fluctuations. Our model shows two major insurance effects of species richness on ecosystem productivity: (i) a buffering effect, i.e., a reduction in the temporal variance of productivity, and (ii) a Performance- enhancing effect, i.e., an increase in the temporal mean of productivity. The strength of these insurance effects is deler- mined by three factors: (i) the way ecosystem productivity is determined by individual species responses to environmental fluctuations, (ii) the degree of asynchronicity of these re- sponses, and (iii) the detailed form of these responses. In particular, the greater the variance of the specics responscs, the lower the species richness at which the temporal mean of the ecosystem process saturates and the ecosystem becomes redundant. These results provide strong theoretical foun- dation for the insurance hypotbesis, which proves to be a fundamental principle for understanding
  • 机译 在高等植物中从1-脱氧木酮糖类萜类生物合成过程中,二甲基烯丙基焦磷酸不是异戊烯基焦磷酸的固定前体
    摘要:Cell cultures of Catharanthus roseus were supplied with [2-13,C,3-2H]-deoxyxylulose or [2-`13C,4-2H]1- deoxyxylulose. Lutein and chlorophylls were isolated from the cell mass, and hydrolysis of the chlorophyll mixtures afforded phytol. Isotope labeling patterns of phytol and lutein were determincd by 2H NMR and 1H,2H-decoupled 13C NMR. From the data it must be concluded that the deuterium atom in position 3 of deoxyxylulose was incorporated into both iso- pentenyl pyrophosphate (IPP) and dimethylallyl pyrophos- phate with a rate of 75 (with respect to the internal 13C label). The detected stereochcmical signature implies that the Iabel is located preferentially in the (E)-hydrogen atom of IPP. This preferential labeling, in turn, rules out dimethylallyl pyrophosphate as the compulsory precursor of IPP. In the expcriment with [2-13C,4-2R]l-deoxyxylulose, the 13C label was efFicicntly transferred to the terpenoids whereas the 2H Iabel was completely washed out, most probably after IPP formation as a consequence or the isomerization and elonga- tion process. In addition, the data cast light on the stereo- chemical course of the dehydrogenation and cyclization steps involved in the biosynthesis of lutdin.
  • 机译 G1细胞周期停滞和核Smad4 / Dpc4诱导的细胞凋亡:致瘤突变逆转的表型
    摘要:The tumor suppressor Smad4/DPc4 is a transcription activator that binds specific DNA sequences and whose nuclear localization is induced after exposure to type beta transforming growth factor-like cytokines. We explored an inducible system in which Smad4 protein is activated by translocation to the nucleus when cell lines that stably cxpress wild-type or mutant Smad4 proteins fused to a murine estrogen receptor domain are treated with 4-hydroxytamox- ifen. This induced Smad4-mediated tran scriptional activation and a decrease in growth rate, attributable to a cell cycle arrest at the G1 phase and an induction of apoptosis. A lumor-derived mutation affecting a residue critical for DNA-binding demonstrated an ``oncogenic'' phe- notype, having decreases in both the G1 fraction and apoptosis and, consequently, an augmentation of population growth. This model should be useful in the exploration and control of components that lie further downstream in the Smad4 tumor- suppressor pathway.
  • 机译 强制展开纤连蛋白III型模块揭示了拉伸分子识别开关
    摘要:The 10th type III module of fibronectin pos- sesses a beta-sandwich structure consisting of seven beta-strands (AG) that are arranged in two antiparallel sheets. It mediates cell adhesion to surfaces via its integrin binding motif, Arg78, GIy79, and Asp80 (RGD), which is placed at the apex of the loop connecting beta-strands F and G. Stecred molecular dynamics simulations in which tension is applied to the protein's terminal ends reveal that the beta-strand G is the first to break away from the module on forced unfolding whereas the remaining fold maintains its structural integrity. The sepa- ration of strand G from the remaining fold results in a gradual shortening of the distance between the apex of the RGD- containing loop and the module surface, which potentially reduces the loop's accessibility to surface-bound integrins. The shortening is followed by a straightening of the RGD-loop from a tight beta-turn into a linear conformation, which suggests a further decrease of affinity and selectivity to integrins. The RGD-loop therefore is located strategically to undergo strong conformational changes in the early stretching stages of the module and thus constitutes a mechanosensitive control of ligand recognition.
  • 机译 凋亡蛋白抑制剂的鼠同源物的表达与细胞增殖有关
    摘要:The inhibitor of apoptosis (IAP) proteins form a highly conserved gene family that prevents cell death in response to a variety of stimuli. Herein we describe a newly defined murinc IAP, designated Tiap, that proved to be a murine homologue of human survivin based on sequence comparison. TIAP has one baculovirus IAP repeat and lacks a C-terminal RING finger motif TIAP interacted with the processed form of caspase 3 and inhibited caspase-induced cell death. Histological examinations revealed that TIAP is expressed in growing tissues such as thymus, testis, and intestine of adult mice and many tissues of embryos. In in vitro studies, TIAP was induced in splenic T cells activated with anti-CD3 antibody or Con A, and the expression of TIAP was up-regulated in synchronized NIH 3T3 cells at S to Gz/M phase of the cell cycle. We propose that during cell prolifer- alion, cellular protective activity may be augmented with inducible IAPs such as TIAP.
  • 机译 人类乳头瘤病毒16型E7癌蛋白对M2型丙酮酸激酶活性的调节
    摘要:We report here that the E7 oncoprotein encoded by the oncogenic human papillomavirus (HPW type 16 binds to the glycolytic enzyne type M2 pyruvate kinase (M2-PK). M2.PK occurs in a tetrameric form with a high affinity to its substrate phosphoenolpyruvate and a dimeric form with a low affinity to phosphoenolpyruvate, and the transition between both conformations regulates the glyco- Iytic flux in tumor cells. The glycolytic intermcdiate fructose l,6-bisphosphate induces the reassociation of the dimeric to the tetrameric form of MZ-PK. The expression of E7 in an experimental cell line shifts the equilibrium to the dimeric state despite a significant increase in the fructose 1,6- bisphosphate levels. Investigations of HPV-1 6 E7 mutants and the nononcogenic RPV-ll subtype suggest that the interaction of HPV-16 E7 with MZ-PK may be linked to the transforming potential of the viral oncoprotein.
  • 机译 细胞Src与表皮生长因子受体之间的生物协同作用机理
    摘要:Overexpression of both cellular Src (c-Src) and the epidermal growth factor receptor (EGFR) occurs in many of the same human tumors, suggesting that they may functionally interact and contribute to the progression of cancer. Indeed, in murine fibroblasts, overexpression of c-Src bas been shown to potentiate the mitogenic and tumorigenic capacity of the overexpressed EGFR. Potentiation correlated with the ability of c-Src to physically associate with the activated EGFR and the appearance of two unique in vivo phosphorylations on the receptor (Tyr-845 and Tyr-1101). Using stable cell lines of C3H10T1/2 murine fibroblasts that contain kinase-deficient (K-) c-Src and overexpressed wild. type EGFR, we show that the kinase activity of c-Src is required for both the biological synergy with the receptor and the phosphorylations on the receptor, but not for the associ. ation of c-Src with the receptor. In transient transfection assays not only epidermal growth factor but also serum- and lysophosphatidic acid-induced DNA synthesis was ablated in a dominant-negative fashion by a Y845F mutant of the EGFR, indicating that c-Src-induced phosphorylation of Y845 is critical for the mitogenic response to both the EGFR and a G protein-coupled receptor (lysophosphatidic acid receptor). Unexpectedly, the Y845F mutant EGFR was found to retain its full kinase activity and its ability to activate the adapter protein SHC and extracellular signal-regulated kinase ERKZ in response to EGF, demonstrating that the mitogenic pathway involving phosphorylation of Y845 is independent of ERK2. activation. T
  • 机译 感觉神经元中CI因子(宿主细胞因子)的核定位与潜伏期单纯疱疹病毒的重新激活有关
    摘要:After a primary infection, herpes simplex virus is maintained in a latent state in neurons of sensory ganglia until complex stimuli reactivate viral lytic replication. Although the mechanisms governing reactivation from the latent state remain unkuown, the regulated expression of the viral immediate early genes represents a critical point in this process. These genes are controlled by transcription enhancer complexes whose assembly requires and is coordinated by the cellular Cl factor (host cell factor). In contrast to other tissues, the CI factor is not detected in the nuclei of sensory neurons. Experimental conditions that induce the reactiva- tion of herpes simplex virus in mouse model systems result in rapid nuclear localization of the protein, indicating that the CI factor is sequestered in these cells until reactivation signals induce a redistribution of the protein. The regulated local- ization suggests that CI is a critical switch determinant of the viral lytic-Iatent cycle.
  • 机译 果蝇和哺乳动物基因组中核苷酸吸收的模式
    摘要:To estimate patterns of molecular evolution of unconstrained DNA sequences, we used maximum parsi- mony to separate phylogenetic trees of a non-long terminal repeat retrotransposable element into either internal branches, representing mainly the constrained evolution of active lineages, or into terminal branches, representing mainly nonfunctional ``dead-on-arrival,, copies that are uncon- strained by selection and evolve as pseudogenes. The pattern of nucleotide substitutions in unconstrained sequences is expected to be congruent with the pattern of point mutation. We examined the retrotransposon Helena in the Drosophila virilis species group (subgenus Drosophila) and the Drosophila melanogaster species suhgroup (subgenus Sophophora). The patterns of point mutation are indistinguishable, suggesting considerable stability over evolutionary time (40-60 million years). The relative frequencies of different point mutations are unequal, but the ``transition bias'' results largely from an approx. = 2-fold excess of G.C to A.T substitutions. Spontaneous mutation is biased toward A.T base pairs, with an expected mutational equilibrium of approx. = 65 A + T (quite similar to that of long introns). These data also enable the first detailed comparison of patterns of point mutations in Drosophila and mammals. Although the patterns are different, all of the statistical significance comes from a much greater rate of G.C to A.T substitution in mammals, probably because of meth- ylated cytosine ``hotspots.'' When the G.C to A.T substitutions are discounted, the remaining differences are
  • 机译 棘皮动物Hox基因簇的组织
    摘要:The Strongylocentrotus purpuratus genome contains a single tEn-gene Hox complex >0.5 megabase in length. This complex was isolated on overlapping bacterial artificial chromosome and PI artificial chromosome genomic recombinants by using probes for individual genes and by genomic walking. Echinoderm Hox genes of Paralog Groups (PG) I and 2 are reported. The cluster includes genes repre- senting all paralog groups of vertebrate Hox clusters, except that there is a single gene of the PG4-5 types and only three genes of the PGg-12 types. The echinoderm Hox gene cluster is essentially similar to those of the bilaterally organized chordates, despite the radically altered pentameral body plans of these animals.
  • 机译 G alpha o 1的翻译后修饰产生G alpha o 3,这是大脑中丰富的G蛋白
    摘要:G ALPHA O , the most abundant G protein in mam- malian brain, occurs at least in two subforms, i.e., G alpha o 1 and G alpha o 2, derived by alternative splicing of the mRNA. A third G alpha o 1-related isoform, G alpha o3, has been purified, representing about 30 of total Go in brain. Initial studies rcvealed distinct biochemical properties of G alpha o 3 as compared with other G alpha o isoforms. In matrix-assisted laser desorption/ionization pep- tide mass mapping of gel-isolatcd G alpha o 1 and Galpha o 3, C-terminal peptides showed a difference of +l Da for G alpha o3. Nanoclec- trospray tandem mass spectrometry sequencing revealed an Asp instead of an Asn at position 346 of G alpha o 3. Gel clectro. phoretic analysis of recombinanl G alpha o3 showed the same mobility as native G alpha o3 but distinct to G alpha o 1. The conversion of 346Asn -> Asp changed the signaling properties, including the velocity of the basal guanine nucleotide-exchange reaction, which points to the involvement of the C terminus in basal guanosine 5'- [r-thio] triphosphate binding. No cDNA coding for G alphaa o 3 was detectcd, suggesting an enzymatic deamidation of G alpha o1 by a yet-unidentified activity. Therefore, G alpha hetero- geneity is generated not only at the DNA or RNA levels, but also at the protein level. The relative amount of G alpha o1 and G alpha o 3 different from cell type to cell type, indicating an additional principle of G protein regulation.
  • 机译 组蛋白修饰控制酿酒酵母中复制无关的染色质组装途径的细胞周期调控
    摘要:We describe a replication-independent, cell cycle-regulated chromatin assembly pathway in budding yeast. The activity of this pathway is low in S phase extracts but is very high in G2, M, and G1 cell extracts, with peak activity in late M/early G1. The cell cycle regulation of this pathway requires a specific pattern of posttranslational mod- ification of histones H3 and/or H4, which is distinct for H3/H4 present in S phase versus M and G1 phase cell extracts. Histone H3/H4 modification is therefore important for the reciprocal control of replication-dependent and -independent chromatin assembly pathways during the cell cycle.
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