一种拟南芥ZIP蛋白全长cDNA的克隆及表达

利用异质DNA探针 ,从拟南芥叶片cDNA文库中筛选到一个阳性cDNA克隆———AT10 3。DNA序列分析表明 ,其编码产物含有一个亮氨酸拉链结构域和一个核定位信号肽。GenBank数据库搜寻结果显示 ,AT10 3未与任何已知功能基因有同源性 ,但与裂叶牵牛PNZIP、集胞藻的一个ORF、紫菜的一个ORF ,构成了一个在进化上非常保守的含有亮氨酸拉链结构域的新家族 ,极有可能是一新的核基因转录因子。AT10 3是一个单拷贝基因 ,在叶片组织中强烈表达并受光调节     

JLP associates with kinesin light chain 1 through a novel leucine zipper-like domain

Scaffolding proteins exist in eukaryotes to properly assemble signaling proteins into specific multimeric functional complexes. JLP is a novel leucine zipper protein belonging to a family of scaffolding proteins that assemble JNK signaling modules. JLP is a proline-rich protein that contains two leucine zipper domains and a highly conserved C-terminal domain. We have identified kinesin light chain 1 (KLC1) as a binding partner for the second leucine zipper domain of JLP using yeast two-hybrid screening. The interaction domain of KLC1 was mapped to its tetratripeptide repeat, which contains a novel leucine zipper-like domain that is crucial for the interaction with JLP. Mutations of Leu-280, Leu-287, Val-294, and Leu-301 within this domain of KLC1 disrupted its ability to associate with JLP. Immunofluorescence studies showed that JLP and KLC1 co-localized in the cytoplasm and that the localization of JLP was dependent on its second leucine zipper. Ectopic expression of a dominant negative form of KLC1 resulted in the mislocalization of endogenous JLP. Moreover, the association between JLP and KLC1 occurred in vivo and was important in the formation of ternary complex with JNK1. These results identify a novel protein-protein interaction between KLC1 and JLP that involves leucine zipper-like domains and support the role of motor proteins in the spatial regulation of signaling modules.     

Mutational analysis of the leucine zipper-like motif of the human immunodeficiency virus type 1 envelope transmembrane glycoprotein.

The N-terminal region of the envelope (env) transmembrane protein of human immunodeficiency virus type 1 (HIV-1) has a leucine zipper-like motif. This highly conserved zipper motif, which consists of a heptad repeat of leucine or isoleucine residues, has been suggested to play a role in HIV-1 env glycoprotein oligomerization. This hypothesis was tested by replacing the highly conserved leucine or isoleucine residues in the zipper motif with a strong alpha-helix breaker, proline. We report here that such substitutions did not abolish the ability of env protein to form oligomers, indicating that this highly conserved zipper motif does not have a crucial role in env protein oligomerization. However, the mutant viruses all showed impaired infectivity, suggesting that this conserved zipper motif can have an important role in the virus life cycle.     

Inhibition of HIV-1 entry before gp41 folds into its fusion-active conformation

HIV-1 entry into its host cell is modulated by its transmembrane envelope glycoprotein (gp41). The core of the activated conformation of gp41 consists of a trimer of heterodimers comprising a leucine/isoleucine zipper sequence (represented here by the synthetic peptide N36 or by the longer N51 peptide) and a C-terminal highly conserved region (represented here by C34). A correlation was found between the action of DP178, which is a potent inhibitor of HIV-1 entry into its host cell, and its ability to interact with the leucine/isoleucine zipper sequence. This correlation was further tested and confirmed by circular dichroism spectroscopy. We found that whereas DP178 perturbs the partial alpha-helix nature of peptides corresponding to the leucine/isoleucine zipper sequence (N36 or N51), it cannot perturb the trimer of heterodimers conformation, modeled by the complex of N36 or N51 with C34. Therefore, we suggest that the already formed trimer of heterodimers is not the target of inhibition by DP178. Our results are consistent with a model in which DP178 acquires its inhibitory activity by binding to an earlier intermediate of gp41, in which the N and C peptide regions are not yet associated, thus allowing DP178 to bind to the leucine/isoleucine zipper sequence and consequently to inhibit transition to the fusion-active conformation.     

小黑杨HD-Zip转录因子家族生物信息学及应答盐胁迫分析

HD-Zip转录因子基因是植物中特有的一类蛋白家族,在植物生长发育和逆境应答胁迫过程中发挥重要作用。HD-Zip转录因子基因是由高度保守的同源异型结构域（HD）和亮氨酸拉链域（LZ）结构域构成的特殊结构模型。杨树HD-Zip转录因子家族共有63个基因,可被分为HD-ZipⅠ、HD-ZipⅡ、HD-ZipⅢ和HD-ZipⅣ四个亚家族。本文利用RNA-Seq分析了盐胁迫条件下HD-Zip基因家族在小黑杨根、茎、叶等不同组织的基因表达差异,从转录组水平揭示其应答胁迫环境的分子机制,结果表明,盐胁迫下在叶中有25个HD-Zip基因下调表达,21个基因上调表达;茎中有42个基因下调表达,11个基因上调表达;根中有26个基因下调表达,24个基因上调表达。另外,本文根据拟南芥HD-Zip转录因子家族基因的已知功能,预测了杨树HD-Zip转录因子同源基因的功能,并利用生物信息学方法分析了杨树HD-Zip转录因子蛋白序列的保守结构域、氨基酸组成和理化性质等,为进一步研究杨树HD-Zip转录因子基因功能提供参考。     

Chlorophyllase is a rate-limiting enzyme in chlorophyll catabolism and is posttranslationally regulated

Chlorophyll is a central player in harvesting light energy for photosynthesis, yet the rate-limiting steps of chlorophyll catabolism and the regulation of the catabolic enzymes remain unresolved. To study the role and regulation of chlorophyllase (Chlase), the first enzyme of the chlorophyll catabolic pathway, we expressed precursor and mature versions of citrus (Citrus sinensis) Chlase in two heterologous plant systems: (1) squash (Cucurbita pepo) plants using a viral vector expression system; and (2) transiently transformed tobacco (Nicotiana tabacum) protoplasts. Expression of full-length citrus Chlase resulted in limited chlorophyll breakdown in protoplasts and no visible leaf phenotype in whole plants, whereas expression of a Chlase version lacking the N-terminal 21 amino acids (ChlaseDeltaN), which corresponds to the mature protein, led to extensive chlorophyll breakdown in both tobacco protoplasts and squash leaves. ChlaseDeltaN-expressing squash leaves displayed a dramatic chlorotic phenotype in plants grown under low-intensity light, whereas under natural light a lesion-mimic phenotype occurred, which was demonstrated to follow the accumulation of chlorophyllide, a photodynamic chlorophyll breakdown product. Full-length and mature citrus Chlase versions were localized to the chloroplast membrane fraction in expressing tobacco protoplasts, where processing of the N-terminal 21 amino acids appears to occur. Results obtained in both plant systems suggest that Chlase functions as a rate-limiting enzyme in chlorophyll catabolism controlled via posttranslational regulation.     

双亮氨酸拉链激酶DLK对JNK信号通路的调控作用及机制

c-Jun氨基末端激酶(c-Jun NH₂-ternimal kinase, JNK)属于进化上相当保守的促分裂原活化蛋白激酶 (Mitogen-activated protein kinase, MAPK)超家族。大量的研究揭示, JNK在细胞增殖、分化、迁移、凋亡和形态建成中起着关键作用, 并与多种人类疾病的发生与发展密切相关。双亮氨酸拉链激酶(DLK)在结构上属于MLK(Mixed lineage kinase)家族, 功能上则是MAPKKK(MAP kinase kinase kinase)中一员, 可通过MAPKK(MAP kinase kinase)对JNK的活性进行调节, 从而参与细胞凋亡、迁移、分化等一系列重要细胞反应。文章结合DLK与JNK的研究历史与最新进展, 就DLK-JNK通讯所参与的细胞凋亡、迁移及分化等活动做一简要综述。     

低温弱光胁迫对野生大豆和大豆栽培种光系统功能的影响

以野生大豆和栽培大豆为材料,通过同时测定大豆叶片的叶绿素荧光快速诱导动力学曲线和对820nm光的吸收曲线,以及测定超氧化物歧化酶（SOD）和抗坏血酸过氧化物酶（APX）的活性,分析了低温弱光胁迫及常温弱光恢复下这2种大豆光系统Ⅱ（PSⅡ）和光系统Ⅰ（PSI）功能的变化。结果表明,低温弱光胁迫对这2种大豆的PSI和PSⅡ的功能都造成伤害;在低温弱光胁迫下,维持较高的SOD和APX活性和维持PSI和PSⅡ的协调性是野生大豆比栽培大豆耐低温的一个重要原因。