PLD 的生化特性及在信号传导中的作用

磷脂酶 D(PLD)是一种分解磷脂的多功能酶,磷脂酶可激活调控许多重要的细胞生理功能,在信号转导、小泡运输、有丝分裂、激素作用的发挥、细胞骨架组装、防御反应以及种子萌发和衰老过程中都起重要作用．主要介绍了磷脂酶基因的生化特性及在植物信号转导中的作用．     

磷脂酶D（PLD）基因的结构、表达及其表达产物在信号转导中的作用

磷脂酶D(PLD EC 3.1.4.4)水解磷脂(PL),磷脂构成生物膜的骨架, 磷脂酶的激活不仅对细胞的结构和稳定性有很重要的作用,而且调控许多重要的细胞生理功能,例如PLD在信号转导、小泡运输、有丝分裂 、激素作用的发挥、细胞骨架组装、防御反应以及种子萌发和衰老过程中都起重要作用。近年来它在跨膜信号转导中的重要作用，越来越引起人们的重视，成为新的研究热点。介绍了磷脂酶基因的结构特点、亚细胞定位、表达的激活抑制以及其表达产物作为胞内信号分子在植物信号转导中的重要作用。     

磷脂酶D(PLD)基因的结构、表达及其表达产物在信号转导中的作用

磷脂酶D(PLDEC 3 .1 .4.4)水解磷脂 (PL) ,磷脂构成生物膜的骨架 ,磷脂酶的激活不仅对细胞的结构和稳定性有很重要的作用 ,而且调控许多重要的细胞生理功能 ,例如PLD在信号转导、小泡运输、有丝分裂、激素作用的发挥、细胞骨架组装、防御反应以及种子萌发和衰老过程中都起重要作用。近年来它在跨膜信号转导中的重要作用 ,越来越引起人们的重视 ,成为新的研究热点。介绍了磷脂酶基因的结构特点、亚细胞定位、表达的激活抑制以及其表达产物作为胞内信号分子在植物信号转导中的重要作用。     

桑树磷脂酶MaPLA1-2D亚型4个基因的克隆与胁迫应答分析

磷脂酶(phospholipase)是一类在植物生长发育和胁迫应答中起重要调控作用的磷脂水解酶,也是一类重要的信号转导酶。而磷脂酶A1(PLA1)在植物应答生物胁迫和非生物胁迫中的功能研究鲜见报道。研究从桑树(Morus alba L.)中克隆了磷脂酶PLA1的1个亚型MaPLA1-2D基因,对其进行了序列分析、组织表达、胁迫诱导表达和蛋白亚细胞定位分析。结果表明,桑树PLA1-2D亚型基因包括4个成员,命名为MaPLA1-2D.1~MaPLA1-2D.4。4个基因在桑树根和叶中高水平表达,蛋白亚细胞定位在叶绿体。序列和进化分析表明MaPLA1-2D基因4个成员与拟南芥AtDAD1基因的保守结构域序列具有较高相似度且进化关系紧密。MaPLA1-2D基因4个成员的启动子含有多种胁迫应答顺式元件和激素响应元件;胁迫诱导表达模式分析表明MaPLA1-2D基因表达受干旱和脱落酸处理显著诱导。以上结果说明,MaPLA1-2D基因与拟南芥DAD同源,可能在桑树非生物胁迫应答中发挥重要功能。     

磷酸肌醇磷脂酶C在DREB2表达调控中的作用

环境胁迫对植物的生长不利。转录因子DREB2对干旱、高温、低温等非生物胁迫应答基因的表达具有重要的调控作用。磷酸肌醇磷脂酶C对 DREB2 基因有双向调节机制。深入了解 DREB2 和磷酸肌醇磷脂酶C的研究进展及其在生物工程上的应用,以及磷酸肌醇磷脂酶C对 DREB2 基因的表达调控机理,可以为磷酸肌醇磷脂酶C和 DREB2 基因在提高植物胁迫耐受性中的利用提供基础。     

植物中的磷脂酶D

介绍了植物磷脂酶D(PLD)的生化性质、克隆、基因组结构、氨基酸序列结构、活性调控、信号转导和细胞生理功能的研究进展.     

表观遗传调控植物叶片衰老的研究进展

植物叶片衰老是一个非常重要的发育过程,涉及大分子的有序分解从而将营养物质从叶片转移到其他器官,对植物的生存和适应至关重要。叶片衰老主要受植物的发育调控,但同时也受内部和外部环境因素的影响,涉及高度复杂的基因调控网络和多层级的调控。近年来的研究表明表观遗传是调控植物叶片衰老的一种重要调控方式。该研究综述了植物叶片衰老过程中的表观遗传调控机制,包括组蛋白修饰、DNA甲基化、ATP依赖的染色质重塑和非编码RNA介导的调控,并对该领域今后的发展方向进行了展望。     

大豆microRNA基因GmMIR160A负调控植物叶片衰老进程

叶片衰老是受内外多种因子影响的遗传发育进程。生长素、细胞分裂素和乙烯等多种植物激素是调控叶片衰老的重要内部因子,它们通过长或短距离运输形成叶片组织内特定的区域分布和浓度梯度,从而直接或间接参与植物叶片衰老过程。分子遗传学表明,细胞分裂素和乙烯分别是叶片衰老的抑制子和正调节子,而生长素如何参与叶片衰老的分子机制目前还不清晰。植物体内成熟小分子RNA由小RNA基因转录并通过特定酶加工形成的21~23bp的双链RNA分子。这些小分子通过不完全配对方式抑制其靶基因转录和/或表达,参与植物生长发育多个过程,然而这类小RNA分子如何调控植物叶片衰老发育过程目前则还鲜有报告。大豆是重要的油料作物,具有典型的单次结实性衰老特征。研究大豆叶片衰老具有重要的科学意义和深远的应用价值。该文采用实时荧光定量PCR(qPCR)技术分析大豆(Glycine max)micro RNA基因GmMIR160A的表达模式,发现大豆第一复叶中GmMIR160A表达受外源生长素和黑暗处理的诱导,暗示该基因是生长素快速响应的叶片衰老相关基因。为进一步探究GmMIR160A在大豆叶片发育中的功能,构建了肾上腺皮质激素(Glucocorticoid,GR)类似物地塞米松(Dexamethasone,DEX)诱导表达GmMIR160A双元表达载体并通过农杆菌介导的子叶节方法转化野生型大豆。通过抗性筛选和基因组PCR鉴定并结合表型分析,共获得了4株诱导表达的稳定遗传转基因植株(株系OX-3、OX-5、OX-7和OX-8)。GmMIR160A过表达植株根、茎、叶、花和果实在形态学上与野生型相比无显著差异,但叶片的叶绿素含量增加、最大光量子效率(Fv/Fm)增强。进一步分子分析发现,转基因大豆叶片中GmARFs和衰老标记基因(GmCYSP1)表达明显下降,表明大豆Gma-miR160通过抑制靶基因GmARFs的表达来负调控植物叶片的衰老进程。该文揭示了生长素通过小分子RNA调控叶片发育一条新途径,为研究植物激素调控植物叶片衰老提供了新的思路。     

细胞分裂素对植物衰老的延缓作用

细胞分裂素是一类重要的植物激素,它可在一定程度上延缓植物的衰老。主要从3个方面综述了细胞分裂素与植物衰老之间的关系,即:(1)植物衰老过程中内源细胞分裂素含量变化;(2)外源细胞分裂素的影响;(3)转入与细胞分裂素的合成、降解相关的基因对植物衰老产生的影响。此外,还从细胞分裂素与糖、与脂质氧化反应以及与其它植物激素的关系方面探讨了细胞分裂素在延缓植物衰老中的作用机理。     

生长素与植物叶片衰老调控

作为植物叶片发育的最后一个阶段,叶片衰老的启动和进程由遗传程序严格控制,并受到内外源不同因子的协同调控.多种植物激素对叶片的衰老发挥重要的调控作用,目前认为乙烯、脱落酸、水杨酸、茉莉酸和油菜素甾醇等激素促进植物叶片衰老,而细胞分裂素和赤霉素则抑制植物叶片衰老.传统观念曾认为生长素对植物叶片衰老起负调节作用,但近年来越来越多的实验证据表明生长素是叶片衰老的正调节因子.本文旨在对生长素在叶片衰老调控中的功能和研究历程进行简要综述,为进一步理解植物叶片衰老调控中的激素功能奠定基础.     

Cloning and molecular characterization of phospholipase D (PLD) delta gene from longan (Dimocarpus longan Lour.)

Longan (Dimocarpus longan Lour.) is a non-climacteric fruit with a short postharvest life. The regulation of phospholipase D (PLD) activity closely relates to postharvest browning and senescence of longan fruit. In this study, a novel cDNA clone of longan PLDδ (LgPLDδ) was obtained and registered in GenBank (accession No. JF791814). The deduced amino acid sequence possessed all of the three typical domains of plant PLDs, a C2 domain and two catalytic HxKxxxxD motifs. The tertiary structure of LgPLDδ was further predicted. The western blot result showed that the LgPLDδ protein was specifically recognized by PLDδ antibody. The Q-RT-PCR (real-time quantitative PCR) result showed that the level of LgPLDδ mRNA expression was higher in senescent tissues than in developing tissues, which was also high in postharvest fruit. The western-blotting result further certified the different expression of LgPLDδ. These results provided a scientific basis for further investigating the mechanism of postharvest longan fruit adapting to environmental stress.     

Involvement of phospholipase D-related signal transduction in chemical-induced programmed cell death in tomato cell cultures

Phospholipase D (PLD) and its product phosphatidic acid (PA) are incorporated in a complex metabolic network in which the individual PLD isoforms are suggested to regulate specific developmental and stress responses, including plant programmed cell death (PCD). Despite the accumulating knowledge, the mechanisms through which PLD/PA operate during PCD are still poorly understood. In this work, the role of PLDα1 in PCD and the associated caspase-like proteolysis, ethylene and hydrogen peroxide (H₂O₂) synthesis in tomato suspension cells was studied. Wild-type (WT) and PLDα1-silenced cell lines were exposed to the cell death-inducing chemicals camptothecin (CPT), fumonisin B1 (FB1) and CdSO₄. A range of caspase inhibitors effectively suppressed CPT-induced PCD in WT cells, but failed to alleviate cell death in PLDα1-deficient cells. Compared to WT, in CPT-treated PLDα1 mutant cells, reduced cell death and decreased production of H₂O₂ were observed. Application of ethylene significantly enhanced CPT-induced cell death both in WT and PLDα1 mutants. Treatments with the PA derivative lyso-phosphatidic acid and mastoparan (agonist of PLD/PLC signalling downstream of G proteins) caused severe cell death. Inhibitors, specific to PLD and PLC, remarkably decreased the chemical-induced cell death. Taken together with our previous findings, the results suggest that PLDα1 contributes to caspase-like-dependent cell death possibly communicated through PA, reactive oxygen species and ethylene. The dead cells expressed morphological features of PCD such as protoplast shrinkage and nucleus compaction. The presented findings reveal novel elements of PLD/PA-mediated cell death response and suggest that PLDα1 is an important factor in chemical-induced PCD signal transduction.     

Developmental regulation of phospholipase D in tomato fruits

The catabolism of phospholipids initiated by phospholipase D (PLD, EC 3.1.4.4) is an inherent feature of developmental processes that include fruit growth and ripening. In cherry tomatoes (Lycopersicon esculentum Mill.), soluble and membrane-associated PLD activities increased during fruit development, which peaked at the mature green and orange stages. The increase in PLD activity was associated with a similar increase in the intensity of a 92 kDa band as demonstrated by western blot analysis. A full-length cDNA having 2430 bp and encoding a putative polypeptide with 809 amino acids, was isolated using tomato RNA, RT-PCR and 5' and 3' rapid amplification of cloned ends (RACE). Analysis of the primary and secondary structures showed the presence of the C2 domain, the PLD domain and several other features characteristic of PLD alpha. Microtom tomato plants transformed with antisense PLD alpha cDNA, were similar to untransformed plants and showed normal fruit set and development. The ethylene climacteric was delayed by over 7 d in the antisense PLD fruits, indicative of a slower ripening process. The leaves and unripened fruits of antisense PLD microtom plants possessed lowered PLD activity and PLD protein, as demonstrated by western blotting. However, during ripening, PLD activity in the transgenic fruits was maintained at a higher level than that in the untransformed control. Immunolocalization of PLD in microtom tomato fruits revealed the cytosol-membrane translocation of PLD during fruit development. The ripe fruits of antisense PLD celebrity plants possessed lowered PLD expression and activity and showed increased firmness and red colour. These results suggest that the expression of antisense PLD cDNA could be variable in different tomato varieties. The potential role of PLD in ethylene signal transduction events is discussed.     

Identification and characterization of phospholipase D and its association with drought susceptibilities in peanut (Arachis hypogaea)

Preharvest aflatoxin contamination has been identified by the peanut industry as a serious issue in food safety and human health because of the carcinogenic toxicity. Drought stress is the most important environmental factor exacerbating Aspergillus infection and aflatoxin contamination in peanut. The development of drought-tolerant peanut cultivars could reduce aflatoxin contamination and would represent a major advance in the peanut industry. In this study, we identified a novel PLD gene in peanut (Arachis hypogaea), encoding a putative phospholipase D (PLD, EC 3.1.4.4). The completed cDNA sequence was obtained by using the consensus-degenerated hybrid oligonucleotide primer strategy. The deduced amino acid sequence shows high identity with known PLDs, and has similar conserved domains. The PLD gene expression under drought stress has been studied using four peanut lines: Tifton 8 and A13 (both drought tolerant) and Georgia Green (moderate) and PI 196754 (drought sensitive). Northern analysis showed that PLD gene expression was induced faster by drought stress in the drought-sensitive lines than the drought tolerance lines. Southern analysis showed that cultivated peanut has multiple copies (3 to 5 copies) of the PLD gene. These results suggest that peanut PLD may be involved in drought sensitivity and tolerance responses. Peanut PLD gene expression may be useful as a tool in germplasm screening for drought tolerance. The nucleotide sequence, reported in this paper, have been submitted to GenBank under accession number AY274834.     

Identification of a novel phospholipase D gene and effects of carbon sources on its expression in <Emphasis Type="Italic">Bacillus cereus</Emphasis> ZY12

In the present study, a new strain, Bacillus cereus ZY12, producing phospholipase D (PLD) was identified. The expression of PLD in this strain was found to be induced by its substrate, phosphatidylcholine (PC), and completely silenced by other carbon sources, such as glucose, fructose, and maltose, which are generally used in microbial growth cultures, thus presenting a unique expression pattern different from other PLD-producing microorganisms. This study is the first to report on the ability of B. cereus to produce PLD, and successfully clone its PLD-coding gene and identify its function, extending the knowledge on PLD distribution and evolution in microorganisms.     

Phospholipase D in cellular senescence

Cellular senescence appears to be an important part of organismal aging. Cellular senescence is characterized by flattened enlarged morphology, inhibition of DNA replication in response to growth factors, inability to phosphorylate the pRb tumor suppressor protein, inability to produce c-fos or AP-1 and overexpression of a variety of genes, notably p21 (CIP-1/WAF-1) and p16^INK. It is now clear that certain early mitotic signals become defective with the onset of senescence. Among these is the PLD/PKC pathway. Evidence suggests that activation of PLD and PKC is critical for mitogenesis. Recent data suggest that the defect in PLD/PKC in cellular senescence is a result of elevated cellular ceramide levels which inhibit PLD activation. It appears that the elevated ceramide is a result of neutral sphingomyelinase activation. Ceramide acts to inhibit the activation of PLD by possibly three mechanisms, inhibiting activation by Rho, translocation to the membrane and gene expression. Addition of ceramide to young cells not only inhibits PLD but also recapitulates all the standard measures of cellular senescence as described above.     

Phosphatidic acid integrates calcium signaling and microtubule dynamics into regulating ABA-induced stomatal closure in Arabidopsis

Specific cellular components have been identified to function in abscisic acid (ABA) regulation of stomatal apertures, including calcium, the cytoskeleton, and phosphatidic acid. In this study, the regulation and dynamic organization of microtubules during ABA-induced stomatal closure by phospholipase D (PLD) and its product PA were investigated. ABA induced microtubule depolymerization and stomatal closure in wide-type (WT) Arabidopsis, whereas these processes were impaired in PLD mutant (pldα1). The microtubule-disrupting drugs oryzalin or propyzamide induced microtubule depolymerization, but did not affect the stomatal aperture, whereas their co-treatment with ABA resulted in stomatal closure in both WT and pldα1. In contrast, the microtubule-stabilizing drug paclitaxel arrested ABA-induced microtubule depolymerization and inhibited ABA-induced stomatal closure in both WT and pldα1. In pldα1, ABA-induced cytoplasmic Ca²⁺ ([Ca²⁺]_cyt) elevation was partially blocked, and exogenous Ca²⁺-induced microtubule depolymerization and stomatal closure were impaired. These results suggested that PLDα1 and PA regulate microtubular organization and Ca²⁺ increases during ABA-induced stomatal closing and that crosstalk among signaling lipid, Ca²⁺, and microtubules are essential for ABA signaling.