Strawberry vein banding virus P6 protein intracellular transport and an important domain identification     

Yuan PAN  Xiu-hong ZHOU  Shuai LI  Ming-feng FENG  Man-ling SHI  Deng-pan ZUO  Xi-zi JIANG  Jing CHEN  Ya-hui HU  Xiang-xiang ZHANG  Tong JIANG 《农业科学学报》2018,17(9):2031-2041

Strawberry vein banding virus(SVBV)-infected strawberry cells contain cytoplasmic inclusions with isometric particles.To identify the components of the inclusions,green fluorescent protein(GFP)was fused to the carboxy-terminus(C-terminus)of SVBV open reading frames,these constructs were separately transformed into Agrobacterium tumefaciens and infiltrated into Nicotiana benthamiana leaves.Results showed that the SVBV P6 protein assembled into prominent and amorphous inclusion bodies(IBs).To investigate P6 subcellular localization,P6-GFP was ectopically expressed in N.benthamiana leaves by agroinfiltration and then stained with 4′,6-diamidino-2-phenylindole(DAPI).We found the P6 protein accumulated in the nuclei and also formed cytoplasmic IBs with different sizes.To further determine the location of P6 IBs in the cytoplasm,and explore whether the P6 IBs move freely or depend on cytoskeleton and endoplasmic reticulum(ER),the microfilament marker protein(GFP-ABD2-GFP),microtubules marker protein(m Cherry-MAP65-1)and ER marker protein(m Cherry-HDEL)were separately coexpressed with P6-GFP and into N.benthamiana leaves by agroinfiltration,exhibiting that P6 IBs aligned with cytoskeleton and endoplasmic reticulum.Meanwhile,coinfiltration of P1 and P6 indicated the P6colocalized with the P1 protein at periphery of cells.The P6 protein contains one C-terminal nuclear localization signal(NLS)region,a P6 protein mutant with a deleted NLS did not localize in the nucleus,did not form IBs,and was unable to facilitate exogenous GFP expression.These results demonstrate that the deleted NLS region is an important P6 domain required for biological functions.In summary,the mobile P6 IBs are associated with ER,microfilaments and microtubules and move along microfilaments to the SVBV P1 protein in the PD.  相似文献   

巴西橡胶树叶肉细胞原生质体中细胞骨架结构及动态观察     

王倩男  张晓东  罗红丽  安邦 《热带作物学报》2018,39(3):494-501

细胞骨架通过结构的重排在植物响应逆境胁迫的过程中起着重要作用。本研究克隆真菌微丝结合蛋白的一段保守结构域lifeact及小鼠MAP4的微管结合结构域MBD,构建与荧光蛋白EGFP融合表达的瞬时表达载体,利用原生质体瞬时表达体系成功观察到了橡胶树叶肉细胞中微丝和微管骨架的结构,并通过对原生质体细胞进行不同处理观察胞内微丝和微管骨架的结构变化。此外,本研究还成功的捕捉到橡胶树原生质体细胞中微丝及微管的动态行为。   相似文献   

Effects of Microtubule Polymerization Inhibitor on the Hypersensitive Response of Wheat Induced by the Non-Host Pathogen Sphaerotheca fuliginea   总被引：1，自引：0，他引：1  

LI Hong-li  WANG Hai-yan  HAO Xin-yuan  SONG Xiao-he  MA Qing 《中国农业科学(英文版)》2010,9(3):378-382

The plant cytoskeleton is a highly dynamic and versatile intracellular scaffold composed of microtubules and microfilaments, serving a multiplicity of functions in plant cells. To reveal the relationship between the cytoskeleton in wheat （Triticum aestivum L.） cv. Suwon 11 attacked by the non-host pathogen Sphaerotheca fuliginea and the initiation of the hypersensitive response, the microtubule inhibitor oryzalin was injected into the wheat leaves immediately prior to inoculation. The incidence of hypersensitive cell death was significantly lower than that in water-treated control. In addition, the occurrence of hypersensitive cell death was also delayed and S. fuliginea was able to penetrate and form haustoria in epidermal tissues of wheat. All the results above indicated that hypersensitive cell death was associated with depolymerisation of microtubules, suggesting that microtubules might play an important role in the expression of non-host resistance of wheat.  相似文献   

马齿苋多糖对tau蛋白磷酸化影响的研究   总被引：1，自引：0，他引：1  

康洁 《中国农学通报》2010,26(14):113-116

目的是研究马齿苋多糖对神经细胞骨架微管结合蛋白-tau蛋白的磷酸化影响。方法通过构建基因表达载体pEGFP- tau-s3和转染表达载体pEGFP- tau-s3到3T3细胞,创造体外神经细胞骨架研究模型;通过共转染pEGFP- tau-s3与pcDNA-GSK-3β到3T3细胞、荧光显微观察和Wstern blotting实验,证明了马齿苋多糖对tau蛋白的磷酸化有影响,可以抑制GSK激酶对tau蛋白的磷酸化。结论马齿苋多糖对神经细胞微管骨架有保护作用  相似文献   

Cdc42调控细胞骨架影响破骨细胞分化的研究     

宋瑞龙  徐天祺  梁琰  曹莹  刘宗平 《畜牧与兽医》2020,(3):50-55

研究Cdc42在破骨细胞分化成熟过程中的作用及分子机制。采用慢病毒干扰技术构建Cdc42沉默的RAW264.7细胞株,经嘌呤霉素筛选出稳转株。利用荧光定量PCR与Western blot检测其沉默效率。采用TRAP染色观察细胞分化能力。利用激光共聚焦显微镜观察Cdc42沉默经微丝和微管对细胞伪足形成的影响。利用荧光定量PCR与Western blot检测TRAP、PAK4、Cofilin转录和蛋白表达水平。与对照组比较,Cdc42沉默组Cdc42基因的转录极显著下降,蛋白表达水平显著下调,基因沉默效率在50%以上。TRAP阳性破骨细胞数量也急剧减少,细胞呈圆形。丝状伪足和板状伪足减少,TRAP、PAK4、Cofilin转录水平极显著下降,关键蛋白PAK4和Cofilin蛋白表达显著或极显著下调。Cdc42可通过调控细胞骨架蛋白,经丝状伪足和板状伪足,影响破骨细胞的分化。  相似文献   

Fenoxaprop‐ethyl affects mouse oocyte quality and the underlying mechanisms     

Ya‐Ting He  Lei‐Lei Yang  Yong Zhao  Wei Shen  Shen Yin  Qing‐Yuan Sun 《Pest management science》2019,75(3):844-851

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Depolymerization of the actin cytoskeleton induces defense responses in tobacco plants     

Yuhko Kobayashi  Issei Kobayashi 《Journal of General Plant Pathology》2007,73(5):360-364

Tobacco leaf sections were treated with actin inhibitors, i.e., cytochalasins, to determine the effects of actin depolymerization on tobacco defense responses. Inoculation of the leaf sections with the pathogen Erysiphe cichoracearum, depolymerized the actin cytoskeleton, priming the cells for a hypersensitive response-like cell death. Further, expression of the acidic PR1 and PR2 genes were induced in cytochalasin-treated leaf sections. The intensity of the cytochalasin effects on the defense responses was closely correlated with the extent of actin depolymerization. This suggests that plant cells may perceive perturbation of the actin cytoskeleton, and this stimulus may trigger plant defense responses.  相似文献   

Influence of seminal plasma during different stages of bovine sperm cryopreservation     

Gabriela Bertaiolli Zoca  Eneiva Carla Carvalho Celeghini  Guilherme Pugliesi  Carla Patricia Teodoro de Carvalho  Mayra Elena Ortiz D'Avila Assumpção  Adriano Felipe Perez Siqueira  Leticia Zoccolaro Oliveira  Renata Lançoni  Rubens Paes de Arruda 《Reproduction in domestic animals》2021,56(6):872-883

This study aimed to evaluate the effect of seminal plasma on bovine sperm cryopreservation and to assess the integrity of plasma and acrosomal membranes, mitochondrial potential, remodelling of F-actin cytoskeleton and sperm chromatin fragmentation during the cooling, equilibrium and freezing/thawing stages. Six ejaculates collected from seven Nelore bulls (n = 42) were used in this study. Each ejaculate was divided into two aliquots (with seminal plasma = SP group; without seminal plasma = NSP group) and packed to a final concentration of 50 × 10⁶ sperm per straw. Statistical analyses were performed using SAS software (version 9.3), and p ≤ .05 was considered significant. A time effect was observed for all sperm characteristics (p < .05), except for chromatin fragmentation (p > .05). The presence of seminal plasma better preserved the acrosomal integrity (SP = 75.2% and NSP = 71.7%; p < .05) and also provided lower F-actin remodelling during cryopreservation process (SP = 29.9% and NSP = 32.4%; p < .05). Regarding to the cryopreservation stages, it was observed that cooling step induced higher remodelling of F-actin than the equilibrium and freezing/thawing stages (56.3%, 32.2% and 23.9%, respectively; p < .05). The equilibrium step had minor influence on overall sperm characteristics while the freezing/thawing stage was responsible for the highest percentage of damage in plasma membrane (−65.2%), acrosomal membrane (−34.0%) and mitochondrial potential (−48.1%). On the other hand, none of the cryopreservation stages affected chromatin integrity. It was concluded that the presence of seminal plasma provides increased acrosomal integrity and reduced remodelling of F-actin cytoskeleton. Higher F-actin remodelling is observed after the cooling step while the freezing/thawing step is most damaging to sperm membranes and mitochondrial potential during bovine sperm cryopreservation.  相似文献   

Identification of Dynamic Changes in Proteins Associated with the Cellular Cytoskeleton after Exposure to Okadaic Acid     

Jill A. Opsahl  Sonja Ljostveit  Therese Solstad  Kristin Risa  Peter Roepstorff  Kari E. Fladmark 《Marine drugs》2013,11(6):1763-1782

Exposure of cells to the diarrhetic shellfish poison, okadaic acid, leads to a dramatic reorganization of cytoskeletal architecture and loss of cell-cell contact. When cells are exposed to high concentrations of okadaic acid (100–500 nM), the morphological rearrangement is followed by apoptotic cell death. Okadaic acid inhibits the broad acting Ser/Thr protein phosphatases 1 and 2A, which results in hyperphosphorylation of a large number of proteins. Some of these hyperphosphorylated proteins are most likely key players in the reorganization of the cell morphology induced by okadaic acid. We wanted to identify these phosphoproteins and searched for them in the cellular lipid rafts, which have been found to contain proteins that regulate cytoskeletal dynamics and cell adhesion. By using stable isotope labeling by amino acids in cell culture cells treated with okadaic acid (400 nM) could be combined with control cells before the isolation of lipid rafts. Protein phosphorylation events and translocations induced by okadaic acid were identified by mass spectrometry. Okadaic acid was shown to regulate the phosphorylation status and location of proteins associated with the actin cytoskeleton, microtubules and cell adhesion structures. A large number of these okadaic acid-regulated proteins have previously also been shown to be similarly regulated prior to cell proliferation and migration. Our results suggest that okadaic acid activates general cell signaling pathways that induce breakdown of the cortical actin cytoskeleton and cell detachment.  相似文献   

银灰杨花粉母细胞减数分裂与微管骨架变化     

李燕杰  田梦迪  刘燕  张媛  张平冬 《北京林业大学学报》2017,39(5):9-16

减数分裂是植物有性生殖过程中配子形成的必要阶段,是生殖生物学研究领域的热点。本文以银灰杨雄花枝为研究材料,运用醋酸洋红染色法和间接免疫荧光技术,开展银灰杨花粉母细胞减数分裂及其微管骨架动态变化研究。结果表明:1)银灰杨花药颜色变化可作为初步判别其花粉母细胞减数分裂时期的形态学标记,花粉母细胞从减数分裂细线期发育至四分体时期,花药颜色从嫩绿色,经由黄绿色、微红色、浅红色、鲜红色转为深红色。2)银灰杨花粉母细胞减数分裂末期Ⅰ可观察到与着丝粒微管紧密相连的落后染色体,由于染色体两侧着丝粒微管的均衡牵引而致使其停滞于细胞板中央,不移向两极而被遗弃,造成配子染色体的不平衡,可能是导致银灰杨花粉部分败育的机制。3)银灰杨的胞质分裂属于典型的连续型胞质分裂类型,受辐射状微管系统的调节,从细胞周沿向心扩展,直至细胞质完全分离。4)银灰杨花粉中存在天然2n花粉,其发生可能与减数分裂中期Ⅱ的平行纺锤体和三极纺锤体有关。本研究首次报道了银灰杨花药颜色变化与其花粉母细胞减数分裂进程的关系,从新的角度阐释了银灰杨落后染色体的形成原因及其花粉败育机制,并探索了银灰杨天然2n花粉的发生机理,为银灰杨遗传改良策略的制定奠定了基础。   相似文献