Arabidopsis MPK6 is involved in cell division plane control during early root development,and localizes to the pre‐prophase band,phragmoplast, trans‐Golgi network and plasma membrane

The proper spatial and temporal expression and localization of mitogen‐activated protein kinases (MAPKs) is essential for developmental and cellular signalling in all eukaryotes. Here, we analysed expression, subcellular localization and function of MPK6 in roots of Arabidopsis thaliana using wild‐type plants and three mpk6 knock‐out mutant lines. The MPK6 promoter showed two expression maxima in the most apical part of the root meristem and in the root transition zone. This expression pattern was highly consistent with ‘no root’ and ‘short root’ phenotypes, as well as with ectopic cell divisions and aberrant cell division planes, resulting in disordered cell files in the roots of these mpk6 knock‐out mutants. In dividing root cells, MPK6 was localized on the subcellular level to distinct fine spots in the pre‐prophase band and phragmoplast, representing the two most important cytoskeletal structures controlling the cell division plane. By combining subcellular fractionation and microscopic in situ and in vivo co‐localization methods, MPK6 was localized to the plasma membrane (PM) and the trans‐Golgi network (TGN). In summary, these data suggest that MPK6 localizing to mitotic microtubules, secretory TGN vesicles and the PM is involved in cell division plane control and root development in Arabidopsis.     

人类GABARAPL2基因的亚细胞定位

为了对GABARAPL2（GABAA受体相关蛋白相似蛋白2）基因的功能进行初步分析,首先通过同源比较的方法将序列与其同源物进行比较,发现GABARAPL2的氨基酸序列与GABARAP（GABAA受体相关蛋白）高度同源,而GABARAP已证实通过结合细胞骨架的微蛋白,使GABAA受体聚集,定位在细胞膜上,本文采用PCR法从人脑组织的cDNA文库中扩增出GABARAPL2的cDNA,克隆至T质粒载体中进行测序验证,然后以此为模板引物中引入酶切位点再次PCR,扩增出GABARAPL2的开放阅读框,并将其插入到加强型绿色荧光蛋白融合表达载体EGFP中,将绿色荧光蛋白标记的GABARAPL2和GABARAP分别转染HLF细胞株,结果两种蛋白的分布情况基本一致,在细胞质内和核内均有分布,而且核内的分布较胞质为多,结构功能域分析表明,GABARAPL2含有蛋白激酶C磷酸化位点和酪氨酸激酶磷酸化位点,可能通过磷酸化参与细胞骨架的变化,结论 GABARAPL2和GABARAP不仅在胞质中作为受体相关蛋白协助受体的聚集、定位,还参与体内许多其它重要的生理过程。     

龙眼组蛋白去乙酰化酶基因DlHDT1的克隆与特性分析

组蛋白去乙酰化是植物表观遗传调控的重要组成部分,对染色体结构修饰和基因表达调控发挥着重要的作用。为深入探究组蛋白去乙酰化酶基因(histone deacetylase 1,HDT1)在龙眼体胚发生过程中的功能,该研究结合龙眼基因组数据,采用RT-PCR方法克隆得到龙眼组蛋白去乙酰化酶基因(DlHDT1),对其进行生物信息学分析及亚细胞定位观察,同时结合转录组数据分析DlHDT1在体胚发生过程中的FPKM值,并利用qRT-PCR技术检测PEG6000和NaCl处理下DlHDT1的表达模式。结果表明:(1)DlHDT1基因CDS序列全长918 bp,编码305个氨基酸,该蛋白为不稳定亲水性蛋白,不含信号肽和跨膜结构,共含43个磷酸化位点,相对分子量为32 585.54 Da,等电点为4.65;进化树分析显示龙眼DlHDT1与漾濞槭亲缘关系最近(78.76%)。(2)亚细胞定位显示,DlHDT1蛋白定位于细胞核中;顺式作用元件分析发现DlHDT1基因含有大量光响应元件和脱落酸、茉莉酸甲酯等激素及逆境胁迫响应元件;转录组数据显示,DlHDT1在龙眼体胚发生不同时期均有表达,在胚性愈伤组织(EC)阶段表达最低,在球形胚(GE)阶段表达最高。(3)qRT-PCR显示,在PEG6000和NaCl处理下DlHDT1基因,呈下调表达趋势,推测DlHDT1可能参与调控龙眼对干旱及盐胁迫的响应,并存在负调控关系。研究认为,DlHDT1为核定位基因,可能参与龙眼体胚形态建成并在龙眼响应非生物逆境胁迫过程中发挥重要作用。     

空气重金属元素在悬铃木叶中的亚细胞分布及其区隔化效应

以南京市常见行道树二球悬铃木为试材,研究了交通繁忙区和相对清洁区道路两边悬铃木叶内6种重金属元素的亚细胞分布及其区隔化效应.结果显示:交通污染区悬铃木叶内各亚细胞组分中Cr、Cu、Ni、Pb和Zn 5种重金属元素的含量均明显高于对照区,交通空气污染是影响其含量增加的主要原因之一.相对清洁区和交通污染区5种重金属元素在悬铃木叶片、叶柄的细胞壁组分中含量最高,胞外隔离系数和污染指数均大于0.900,细胞壁是大气重金属元素重要的吸滞器官,并对重金属有明显的阻隔效应;胞内细胞器对Pb和Cu的隔离系数和污染指数最大,细胞器双层膜能在一定程度上抵御重金属元素进入细胞内.悬铃木叶片和叶柄亚细胞组分的污染指数表现为胞质组分＞细胞壁组分＞细胞器组分,即包括液泡液在内的胞质组分是囤积重金属元素的场所.研究表明,悬铃木叶片、叶柄各亚细胞组分对重金属均有不同程度的累积能力,叶内胞质组分的囤积作用以及细胞壁、质膜与细胞器双层膜的区隔化作用可能是悬铃木叶解除重金属元素毒害的重要原因.     

Topological analysis of the fusion process between cellular and subcellular compartments

The study presents an application of the theory of homeomorphic transformations of topological manifolds and the operation of the connected sum of manifolds for topological analysis of membrane transformations during the fusion process between cellular and subcellular compartments. The biological cell and the subcellular structures in the form of vesicles are modelled by an arrangement of two concentric spheres corresponding to the inner and outer layer of the membrane bounding the vesicles. The analysis shows eight succeeding topological stages of membrane transformations during the fusion process and these stages are characterized. It is concluded that there is a vectorial translocation of lipid molecules from the outer layers of the membranes before the fusion process to the internal layer of the membrane bounding the vesicle after the fusion process and there is no lipid translocation in the reverse direction.     

Subcellular Distribution and Chemical Forms of Cadmium in the Medicine Food Homology Plant <i>Platycodon grandiflorum</i> (Jacq.) A.DC.

Although Platycodon grandiflorum (Jacq.) A.DC. is a renowned medicine food homology plant, reports of excessive cadmium (Cd) levels are common, which affects its safety for clinical use and food consumption. To enable its Cd levels to be regulated or reduced, it is necessary to first elucidate the mechanism of Cd uptake and accumulation in the plant, in addition to its detoxification mechanisms. This present study used inductively couple plasma-mass-spectrometry to analyze the subcellular distribution and chemical forms of Cd in different tissues of P. grandiflorum. The experimental results showed that Cd was mainly accumulated in the roots [predominantly in the cell wall (50.96%–61.42%)], and it was found primarily in hypomobile and hypotoxic forms. The proportion of Cd in the soluble fraction increased after Cd exposure, and the proportion of insoluble phosphate Cd and oxalate Cd increased in roots and leaves, with a higher increase in oxalate Cd. Therefore, it is likely that root retention mechanisms, cell wall deposition, vacuole sequestration, and the formation of low mobility and low toxicity forms are tolerance strategies for Cd detoxification used by P. grandiflorum. The results of this study provide a theoretical grounding for the study of Cd accumulation and detoxification mechanisms in P. grandiflorum, and they can be used as a reference for developing Cd limits and standards for other medicine food homology plants.     

拟南芥新型脂转移蛋白AtDHyPRP1的亚细胞定位及其对灰霉菌的抗性

通过遗传转化技术研究了拟南芥脂转移蛋白AtDHyPRP1在细胞中的定位及其对真菌病原体的抗性。采用PCR方法从拟南芥Ws生态型克隆了AtDHyPRP1基因,构建产生pRI101-AN-AtDHyPRP1植物双元表达载体和pCAMBIA1302-AtDHyPRP1-GFP融合表达载体,经农杆菌介导的叶盘和浸花法得到烟草和拟南芥转基因植株。AtDHyPRP1基因能够明显增加烟草对灰霉菌的抗性,转AtDHyPRP1烟草叶片的被侵染部位有大量H2O2积累,激光共聚焦显微观察发现AtDHyPRP1蛋白定位于细胞表面。说明AtDHyPRP1蛋白在合成后被分泌到细胞外执行特殊的功能,与植物抗病防御机制有关。     

Role of cyclooxygenase-2 in Trypanosoma cruzi survival in the early stages of parasite host-cell interaction

Chagas disease, caused by the intracellular protozoan Trypanosoma cruzi, is a serious health problem in Latin America. During this parasitic infection, the heart is one of the major organs affected. The pathogenesis of tissue remodelling, particularly regarding cardiomyocyte behaviour after parasite infection and the molecular mechanisms that occur immediately following parasite entry into host cells are not yet completely understood. When cells are infected with T. cruzi, they develop an inflammatory response, in which cyclooxygenase-2 (COX-2) catalyses rate-limiting steps in the arachidonic acid pathway. However, how the parasite interaction modulates COX-2 activity is poorly understood. In this study, the H9c2 cell line was used as our model and we investigated cellular and biochemical aspects during the initial 48 h of parasitic infection. Oscillatory activity of COX-2 was observed, which correlated with the control of the pro-inflammatory environment in infected cells. Interestingly, subcellular trafficking was also verified, correlated with the control of Cox-2 mRNA or the activated COX-2 protein in cells, which is directly connected with the assemble of stress granules structures. Our collective findings suggest that in the very early stage of the T. cruzi-host cell interaction, the parasite is able to modulate the cellular metabolism in order to survives.     

Novel mannose-sequestration technique reveals variation in subcellular orthophosphate pools do not explain the effects of phosphorus nutrition on photosynthesis in Eucalyptus globulus seedlings

Although only a small proportion of plant phosphorus (P) is used for photosynthesis, the relationships between P and photosynthesis can be strong. It was hypothesized, in this study, that variation in the allocation of orthophosphate (Pi) between active (cytoplasmic) and nonactive (vacuolar) pools would underpin differences in rates of photosynthesis in 4-month-old Eucalyptus globulus seedlings grown with a varying P supply. Photosynthetic biochemistry was assessed by the response of net photosynthesis to increasing intercellular [CO2]. Cytoplasmic Pi was sequestered as mannose 6-phosphate. Total P and the proportion of P as Pi were positively related to P supply. The ratios of active : stored Pi (10-24%) varied little over the range of treatments. Active Pi was positively related to P supply, as was photosynthesis (7 micromol CO2 m(-2) s(-1) with 0 mM P vs. 16 micromol CO2 m(-2) s(-1) with 0.32 mM P). Positive relationships between P supply and photosynthesis were explained best by leaf P content, not by active pools of Pi. The distribution of Pi between the vacuole and the cytoplasm had little impact on the photosynthetic phosphorus-use efficiency (PPUE), and reductions in cytoplasmic Pi had little effect on photosynthesis. Hence, PPUE is an unsuitable guide for assessing plant responses to increasingly unavailable P in the environment.