外源突变基因AGM3过表达对烟草开花的抑制和花器官发育的影响

为了解AGM3基因在异源植物中对开花及花器官发育的影响,采用农杆菌介导法将dominant negative mutation(DNM)结构基因35S-AGM3-E9导入烟草(Nicotiana tabacum),经PCR和Southern检测获得了一批阳性转化植株.荧光定量分析结果显示,AGM3在各个转基因株系中均有...     

The Sir2-Sum1 Complex Represses Transcription Using Both Promoter-Specific and Long-Range Mechanisms to Regulate Cell Identity and Sexual Cycle in the Yeast Kluyveromyces lactis

Deacetylases of the Sir2 family regulate lifespan and response to stress. We have examined the evolutionary history of Sir2 and Hst1, which arose by gene duplication in budding yeast and which participate in distinct mechanisms of gene repression. In Saccharomyces cerevisiae, Sir2 interacts with the SIR complex to generate long-range silenced chromatin at the cryptic mating-type loci, HMLα and HMR a. Hst1 interacts with the SUM1 complex to repress sporulation genes through a promoter-specific mechanism. We examined the functions of the non-duplicated Sir2 and its partners, Sir4 and Sum1, in the yeast Kluyveromyces lactis, a species that diverged from Saccharomyces prior to the duplication of Sir2 and Hst1. KlSir2 interacts with both KlSir4 and KlSum1 and represses the same sets of target genes as ScSir2 and ScHst1, indicating that Sir2 and Hst1 subfunctionalized after duplication. However, the KlSir4-KlSir2 and KlSum1-KlSir2 complexes do not function as the analogous complexes do in S. cerevisiae. KlSir4 contributes to an extended repressive chromatin only at HMLα and not at HMR a. In contrast, the role of KlSum1 is broader. It employs both long-range and promoter-specific mechanisms to repress cryptic mating-type loci, cell-type–specific genes, and sporulation genes and represents an important regulator of cell identity and the sexual cycle. This study reveals that a single repressive complex can act through two distinct mechanisms to regulate gene expression and illustrates how mechanisms by which regulatory proteins act can change over evolutionary time.     

The MDM2 RING Domain and Central Acidic Domain Play Distinct Roles in MDM2 Protein Homodimerization and MDM2-MDMX Protein Heterodimerization

The oncoprotein murine double minute 2 (MDM2) is an E3 ligase that plays a prominent role in p53 suppression by promoting its polyubiquitination and proteasomal degradation. In its active form, MDM2 forms homodimers as well as heterodimers with the homologous protein murine double minute 4 (MDMX), both of which are thought to occur through their respective C-terminal RING (really interesting new gene) domains. In this study, using multiple MDM2 mutants, we show evidence suggesting that MDM2 homo- and heterodimerization occur through distinct mechanisms because MDM2 RING domain mutations that inhibit MDM2 interaction with MDMX do not affect MDM2 interaction with WT MDM2. Intriguingly, deletion of a portion of the MDM2 central acidic domain selectively inhibits interaction with MDM2 while leaving intact the ability of MDM2 to interact with MDMX and to ubiquitinate p53. Further analysis of an MDM2 C-terminal deletion mutant reveals that the C-terminal residues of MDM2 are required for both MDM2 and MDMX interaction. Collectively, our results suggest a model in which MDM2-MDMX heterodimerization requires the extreme C terminus and proper RING domain structure of MDM2, whereas MDM2 homodimerization requires the extreme C terminus and the central acidic domain of MDM2, suggesting that MDM2 homo- and heterodimers utilize distinct MDM2 domains. Our study is the first to report mutations capable of separating MDM2 homo- and heterodimerization.     

SRPX2的功能及其在肿瘤发生发展中的作用

含sushi重复蛋白X连锁2（sushi repeat-containing protein X-linked 2,SRPX2）是一种分子质量约为53 kD,具有细胞外基质蛋白属性的硫酸软骨素蛋白聚糖,在大脑语言中枢的发育过程中发挥重要作用。目前研究发现,SRPX2在多种恶性肿瘤组织中呈高表达,参与肿瘤细胞的增殖、迁移、黏附和侵袭等生物学行为,促进肿瘤的发生和发展,与肿瘤患者的不良预后密切相关。另有研究显示,SRPX2可通过诱导内皮细胞迁移和血管网形成,从而促进血管生成。因此认为,SRPX2是一个潜在的肿瘤治疗靶点。本文对SRPX2 的结构、生物学特征、相关疾病以及在肿瘤中的生物学作用及其机制进行综述。     

APOBEC3G与Vif在HIV-1感染中的作用

载脂蛋白B mRNA编辑催化多肽样（apolipoprotein B mRNA-editing catalytic polypeptide-like,APOBEC）蛋白是一组胞嘧啶脱氨基酶,具有天然的抗病毒活性,对多种病毒具有抑制作用,特别是逆转录病毒. APOBEC3蛋白能够抑制人类免疫缺陷病毒(HIV-1)的感染,其中APOBEC3G和APOBEC3F的作用最强. APOBEC3G能够通过胞嘧啶脱氨基作用和非胞嘧啶脱氨基作用抑制病毒感染. HIV-1病毒感染因子(Vif) 蛋白主要经泛素-蛋白酶体途径介导APOBEC3G降解,从而拮抗其抗病毒作用. APOBEC3G和Vif之间相互作用的研究对于寻求新的抗HIV治疗靶点具有重要意义.     

烟草TTG2蛋白质对NPRI介导的抗病防卫反应与ARF8影响的生长发育进行交叉调控的机制

NPR1蛋白是水杨酸信号和系统获得性抗性的转录调节因子,它的功能受蛋白质降解酶体CUL3．E3的控制。植物的发育主要受生长素信号通路的控制,生长素反应因（ARF）参与生长素信号转导转录调控。植物转录因于NPR1和ARF8分别在蛋白质降解酶体CUL3-E3与CUL1-E3控制下,调控抗病防卫与生长发育。烟草TTG2促进ARF8从细胞质向细胞核转运及其转录调控作用,因此促进生长发育;相反,TTG2把NPR1扣留在细胞质,阻止它对防卫反应基因的转录调控作用,从而抑制抗病性。TTG2与NPR1或ARF8并不直接互作,说明存在协助因子。     

The Arabidopsis BE1 Gene,Encoding a Putative Glycoside Hydrolase Localized in Plastids,Plays Crucial Roles during Embryogenesis and Carbohydrate Metabolism

Carbohydrate metabolism is central to plant growth and development. However, little is known about its role in embryogenesis. Here, we report the characterization of multiple alleles of the BRANCHING ENZYME1 (BE1) gene (also known as EMB2729). The weak allele of be1-3, characterized by positional cloning, carries a single-nucleotide substitution in an exon-intron junction and shows various develop- mental defects during post-germination growth. This mutation causes a reduced level of BE1 mRNA that, likely g...     

The Arabidopsis BE1 Gene, Encoding a Putative Glycoside Hydrolase Localized in Plastids, Plays Crucial Roles during Embryogenesis and Carbohydrate Metabolism

Carbohydrate metabolism is central to plant growth and development. However, little is known about its role in embryogenesis. Here, we report the characterization of multiple alleles of the BRANCHING ENZYME1 (BE1) gene (also known as EMB2729). The weak allele of be1-3, characterized by positional cloning, carries a single-nucleotide substitution in an exon-intron junction and shows various developmental defects during post-germination growth. This mutation causes a reduced level of BE1 mRNA that, likely generated from cryptically spliced pre-mRNA, contains a Glu-to-Lys substitution at codon 366. In four null alleles, BE1 is disrupted by T-DNA insertions, causing embryo developmental arrests at the heart stage. Light microscopy reveals reduced cell divisions and abnormal cell differentiation, thereby leading to defects in setting up the shoot apical meristem, embryonic vascular tissues and cotyledons. Overexpression of BE1 results in a pleiotropic phenotype, indicating that the fine-tuned BE1 level is crucial for plant growth and development. BE1 encodes a putative glycoside hydrolase that is highly conserved in higher plants. A BE1-GFP fusion protein, which is fully functional in complementing be1 mutants, is localized in plastids. The be1-3 phenotype can be partially rescued by glucose, fructose or sucrose, implying the involvement of BE1 in carbohydrate metabolism in plastids.     

The Not4 E3 Ligase and CCR4 Deadenylase Play Distinct Roles in Protein Quality Control

Eukaryotic cells control their proteome by regulating protein production and protein clearance. Protein production is determined to a large extent by mRNA levels, whereas protein degradation depends mostly upon the proteasome. Dysfunction of the proteasome leads to the accumulation of non-functional proteins that can aggregate, be toxic for the cell, and, in extreme cases, lead to cell death. mRNA levels are controlled by their rates of synthesis and degradation. Recent evidence indicates that these rates have oppositely co-evolved to ensure appropriate mRNA levels. This opposite co-evolution has been correlated with the mutations in the Ccr4-Not complex. Consistently, the deadenylation enzymes responsible for the rate-limiting step in eukaryotic mRNA degradation, Caf1 and Ccr4, are subunits of the Ccr4-Not complex. Another subunit of this complex is a RING E3 ligase, Not4. It is essential for cellular protein solubility and has been proposed to be involved in co-translational quality control. An open question has been whether this role of Not4 resides strictly in the regulation of the deadenylation module of the Ccr4-Not complex. However, Not4 is important for proper assembly of the proteasome, and the Ccr4-Not complex may have multiple functional modules that participate in protein quality control in different ways. In this work we studied how the functions of the Caf1/Ccr4 and Not4 modules are connected. We concluded that Not4 plays a role in protein quality control independently of the Ccr4 deadenylase, and that it is involved in clearance of aberrant proteins at least in part via the proteasome.     

The Ubiquitin E3 Ligase NOSIP Modulates Protein Phosphatase 2A Activity in Craniofacial Development

Holoprosencephaly is a common developmental disorder in humans characterised by incomplete brain hemisphere separation and midface anomalies. The etiology of holoprosencephaly is heterogeneous with environmental and genetic causes, but for a majority of holoprosencephaly cases the genes associated with the pathogenesis could not be identified so far. Here we report the generation of knockout mice for the ubiquitin E3 ligase NOSIP. The loss of NOSIP in mice causes holoprosencephaly and facial anomalies including cleft lip/palate, cyclopia and facial midline clefting. By a mass spectrometry based protein interaction screen we identified NOSIP as a novel interaction partner of protein phosphatase PP2A. NOSIP mediates the monoubiquitination of the PP2A catalytic subunit and the loss of NOSIP results in an increase in PP2A activity in craniofacial tissue in NOSIP knockout mice. We conclude, that NOSIP is a critical modulator of brain and craniofacial development in mice and a candidate gene for holoprosencephaly in humans.     

A Comparison of the Roles of Protein Kinase C in Long-Term Potentiation in Rat Hippocampal Areas CA1 and CA3

1. Using agonists and antagonists with specificity toward various isozymes, we have examined the role of protein kinase C (PKC) in long-term potentiation (LTP) in rat hippocampal areas CA1 and CA3.2. Agonists (indolactum V but not phorbol ester) and antagonists (sphingosine, staurosporine, chelerytherene) acting at all PKC isozymes reduce or block LTP induction at both sites.3. However ingenol, a relatively specific agonist at the δ and ε isozymes, blocks LTP in the MF-CA3 pathway, but not in the SC-CA1 pathway.4. Go6976, a relatively specific antagonist of the α and β isozymes, blocks LTP in the SC-CA1 pathway at both ages tested (30- and 60-day-old animals), but blocks LTP in the MF-CA3 in 60 but not 30-day-old animals.5. Our studies indicate that different PKC isozymes are crucial to LTP induction in these two areas of hippocampus, and that there are development changes in the profile of isozymes.     

枣ZjMADS1-box基因克隆与表达分析

以‘金丝4号’枣花蕾和叶片为材料,通过抑制消减杂交文库(SSH文库)的构建、RACE技术和半定量RT-PCR等方法,克隆了枣花发育基因并研究了其时空表达模式,为枣花发育机理研究奠定基础。结果表明:(1)从枣花SSH文库中鉴定出476bp的花发育B类PISTILLATA(PI)基因的EST序列,命名为ZjMADS1-box;通过RACE技术获得了枣ZjMADS1-box基因的全长cDNA序列。(2)枣ZjMADS1-box基因表现出PI基因的序列结构特征。其全长cDNA长度为957bp,编码220个氨基酸残基;在氨基酸序列水平上,该基因与巨桉EgM2(Eu-calyptus grandis M2)和白千层MqPI(Melaleuca quinquenervia PI)的相似性最高为72.6%,与辣椒CaPI(Capsi-cum annuum PI)的相似性最低为61.8%;系统进化树分析显示,枣ZjMADS1-box与MqPI和EgM2基因聚在一起;枣ZjMADS1-box基因编码蛋白的等电点为9.75,分子量为25 587.5Da,属亲水性的非分泌性蛋白。(3)Zj-MADS1-box基因在花器官中特异表达,在嫩茎、成熟叶、腋芽等营养器官中不表达;在4月30日和5月8日花蕾中的表达量略高于5月4日、5月12日、5月16日的表达量,但差异不显著。