水杨酸对高温强光下小麦叶绿体蛋白酶Deg5和PSⅡ功能的调节作用

以小麦(Triticum aestivum)矮抗58为材料,采用0.1mmol/L的外源水杨酸(SA)处理小麦叶片,以清水为对照,通过Western blotting蛋白质印记技术和叶绿素荧光分析,研究了高温强光胁迫(38℃和1600μmol m-2s-1)对小麦叶绿体Deg5蛋白酶、D1蛋白和叶绿素荧光参数的影响及SA的调节作用。结果表明,高温强光胁迫导致小麦叶绿体Deg5蛋白酶、D1蛋白含量和PSⅡ最大光能转化效率(Fv/Fm)降低,原初荧光(Fo)升高。和对照相比,外源SA处理可维持较高的Deg5蛋白酶、D1蛋白、Fv/Fm水平和较低的Fo。说明外源水杨酸可减轻高温强光对Deg5蛋白酶和D1蛋白的损伤,维持较强的PSⅡ功能。     

高温强光对小麦叶绿体Deg1蛋白酶和D1蛋白的影响及水杨酸的调节作用

以小麦品种矮抗58为材料,采用0.3 mmol/L水杨酸(SA)溶液预处理灌浆期小麦叶片,以水预处理为对照,进行3种不同的光温处理:适宜温度中等光强(25℃,600 μmol m-2 s-1)2h、高温强光(38℃,1600μmol m-2 s-1)2h、高温强光2h后置于适宜温度中等光强下恢复3h.测定不同光温条件下,小麦叶绿体的Deg1蛋白酶、D1蛋白和PSⅡ功能的变化及SA的调节效应.结果表明,高温强光胁迫导致Deg1蛋白酶和D1蛋白降解,PSⅡ功能发生可逆损伤.与对照相比,水杨酸预处理不仅能够抑制高温强光下小麦叶绿体Deg1蛋白酶和D1蛋白的降解,维持较高的PSⅡ原初光化学效率(Fv/ Fm)、实际光化学效率(φPSⅡ)、电子传递速率和净光合速率(Pn),而且加快回到非逆境下PSⅡ功能的恢复.     

外源Ca2＋对高温强光胁迫下灌浆期小麦叶片光合机构运转的影响

灌浆期叶面喷施10mmol·L-1 CaCl2对高温强光胁迫下小麦叶片光合电子传递、放氧速率、叶绿素荧光参数和D1蛋白的影响结果表明,Ca2＋预处理可保护D1蛋白,削弱其降解,提高光系统I（PSI）和光系统Ⅱ（PSⅡ）子传递速率、全链电子传递速率、净光合速率（Pn）、PSII最大光化学效率（Fv/Fm）、PSII实际光化学效率（ΦPSⅡ）和光化学猝灭（qp）,维持较低的Fo,最终导致小麦适应高温强光的能力提高。     

早熟桃夏季叶色转红过程中的光化学响应特征

比较研究了‘早美’和‘春蕾’2个早熟桃品种夏季叶色转红对太阳光能的利用和光系统Ⅱ的叶绿素荧光特征的影响。结果表明：早熟桃叶片色素组成的变化会显著影响其光合和叶绿素荧光特性。叶色转红后,早熟桃净光合速率（Pn）日均值、PSII最大光化学效率（Fv/Fm）、PSII实际光化学效率（ФPSII）均上升,无显著光抑制,而绿叶对照‘红花碧桃’的电子传递速率（ETR）、Fv／Fm和ФPSII值均显著下降,7月光合明显受抑制。叶色转红程度较深的‘早美’在夏季高温强光下表现优于‘春蕾’和对照。淬灭分析表明：叶片花色素苷的积累能在短时间内增加PSII天线色素吸收的光能用于光化学反应的份额（P）与用于反应中心热耗散的相对份额（D）。转红后的叶片光化学淬灭系数（qp）显著高于绿叶,PSII光化学效率较高,但耗散过剩激发能的能力显著低于绿叶对照。     

高温胁迫对柑橘光合速率和光系统Ⅱ活性的影响

用红外CO2分析仪和叶绿素荧光仪测定了温州蜜柑和脐橙叶片的净光合速率(Pn)、初始荧光(Fo)、最大光能转换效率(Fv／Fm)及电子传递速率(ETR)．结果表明,与常温(25℃)相比,高温胁迫(38～40℃)使温州蜜柑和脐橙叶片的Pn、Fv／Fm及ETR下降,Fo升高．胁迫25d后温州蜜柑和脐橙叶片的Pn分别下降55．6％和39．8％．Fv／Fm下降22．0％和6．7％,ETR下降55．0％和41．5％,Fo分别上升了113．8％和14．9％．柑橘经高温胁迫后,在25℃下处理10d,叶片的Pn、Fv／Fm、Fo及ETR恢复明显．这些结果说明柑橘的光合速率下降与PSⅡ反应中心失活有关．     

Deg2蛋白酶在拟南芥光损伤D1降解及光保护中的作用

已有研究表明叶绿体内有200种蛋白酶,然而,多数蛋白酶的作用机制尚不清楚,尤其哪些蛋白酶参与了D1蛋白周转.其中Deg2蛋白酶体外实验证明,其参与了光损伤D1蛋白的的初步剪切.为了进一步研究Deg2蛋白酶在植物体内的作用机制,我们筛选了拟南芥Deg2蛋白酶功能缺陷型突变体.在120 μmol·m-2·s-1光照生长条件下,deg2突变体与野生型的生长曲线基本一致;在进一步的高光胁迫(1 800 μmol·m-2·s-1)处理及相同的光胁迫处理条件下,无论林可霉素存在与否,突变体PSⅡ的最大光化学效率(Fv/Fm)都和野生型没有区别;利用蛋白免疫印迹实验同样证明了光损伤D1蛋白的降解速度在deg2突变体和野生型之间也没有明显区别.我们认为Deg2蛋白酶在光抑制情况下对于光损伤D1蛋白的降解以及PSⅡ的修复不是必需的.     

强光及外源活性氧对莴苣叶绿素荧光的影响

莴苣叶绿体在强光处理下发生先抑制,主要表现为叶绿素荧光参数Fv／Fm和ΦPSⅡ降低;外源活性氧H2O2、O2·OH和1O2均能引起叶绿体PSⅡ光化学效率Fv／Fm不同程度的下降,其中以1O2影响最明显;H2O2在诱导叶绿体荧光猝灭过程中,引起荧光产量降低,而使qp、qN、ΦPSⅡ、KD上升;在H2O2诱导的叶绿体荧光猝灭过程中,Fe2 能使qN和KD低于对照;由于1O2的产生对PSⅡ反应中心造成了损伤,引起qp和ΦPSⅡ下降。     

不同供镁水平对两菊芋品种幼苗生物量、光合和叶绿素荧光特性的影响

采用砂培方式研究了不同供镁水平对"南芋1号(Nanyu No.1)"和"青芋2号(Qingyu No.2)"幼苗生物量、光合特性和叶绿素荧光特性的影响。结果表明:缺镁使菊芋叶片叶绿素含量显著降低,干物质积累减少,净光合速率、蒸腾速率和光合电子传递速率显著下降、CO2同化受抑制,同时Fv/Fm、ΦPSII、qP和qN也显著下降,加重了叶片受到光抑制的程度;随着供镁水平的增加,菊芋的干物质积累,叶绿素含量,光合参数和各系列荧光参数均呈现先增加后降低的趋势;南芋1号和青芋2号在镁离子浓度分别为3和6mmol.L-1,叶片的净光合速率、Fo、Fm、Fo’、Fm’、Fv/Fm、ΦPSII、ETR达到最高值,叶绿素a、b及总叶绿素含量也最多,干物质积累量最大;南芋1号的各系列荧光参数、光合参数、叶绿素含量和干物质积累量均显著高于青芋2号。     

细胞外ATP和水杨酸对烟草光合指标的影响

研究了细胞外ATP(eATP)和水杨酸(SA)对烟草(Nicotiana tabacum)叶片的气孔导度(GH2 O)、蒸腾速率(E)、光合作用速率(A)与叶绿素荧光参数[包括PSⅡ潜在最大光化学量子效率(Fv/Fm)、PSⅡ光适应下实际光化学效率Y(Ⅱ)、电子传递速率(ETR)、非光化学荧光淬灭(NPQ)和光化学荧光淬灭(qP)]的影响。结果表明：SA能导致A、GH2 O和E的下降,而eATP的处理能缓解SA造成的A、GH2 O和E的下降;但SA未对叶绿素荧光参数Fv/Fm、Y(Ⅱ)、NPQ、qP和ETR造成显著影响,eATP的加入也未改变SA处理下叶片叶绿素荧光参数的水平。这说明SA能导致光合作用的抑制,而eATP能明显缓解SA对光合作用的抑制,但以上作用可能均和光反应阶段无关。并对其内在机理进行了探讨。     

叶黄素循环和D1蛋白周转在毛竹叶片光保护中的作用

利用叶绿素荧光技术,对强光胁迫下以及叶黄素循环抑制剂-二硫苏糖醇(DTT)和D1蛋白合成抑制剂-硫酸链霉素(SM)处理后毛竹(Phyllostachys edulis (Carr.) Lehaie)的光抑制特征进行研究。结果显示:在夏季中午强光或人为强光胁迫下,毛竹叶片最大光化学效率Fv/Fm均显著降低;在下午光强减弱或黑暗、弱光条件下,Fv/Fm可有效恢复。DTT和SM均可抑制毛竹叶片非光化学淬灭(NPQ),且DTT效果明显优于SM。另外,在强光下,DTT和SM处理均能使毛竹叶片Fv/Fm、实际光化学效率Y(Ⅱ)和光化学淬灭qP等荧光参数下降幅度增大。研究结果表明毛竹叶片具有完善的光破坏防御机制,NPQ与叶黄素循环和D1蛋白周转紧密关联,在叶片光保护机制中具有重要作用。     

DISC1, PDE4B, and NDE1 at the centrosome and synapse

Disrupted-In-Schizophrenia 1 (DISC1) is a risk factor for schizophrenia and other major mental illnesses. Its protein binding partners include the Nuclear Distribution Factor E Homologs (NDE1 and NDEL1), LIS1, and phosphodiesterases 4B and 4D (PDE4B and PDE4D). We demonstrate that NDE1, NDEL1 and LIS1, together with their binding partner dynein, associate with DISC1, PDE4B and PDE4D within the cell, and provide evidence that this complex is present at the centrosome. LIS1 and NDEL1 have been previously suggested to be synaptic, and we now demonstrate localisation of DISC1, NDE1, and PDE4B at synapses in cultured neurons. NDE1 is phosphorylated by cAMP-dependant Protein Kinase A (PKA), whose activity is, in turn, regulated by the cAMP hydrolysis activity of phosphodiesterases, including PDE4. We propose that DISC1 acts as an assembly scaffold for all of these proteins and that the NDE1/NDEL1/LIS1/dynein complex is modulated by cAMP levels via PKA and PDE4.     

GAP-43，Netrin-1，Collapsin-1和Neuropilin-1在出生后小鼠小脑中的动态表达

GAP-43,netrin-1,collapsin-1和neuropilin-1被认为在成网络分布的神经联系中发挥重要的作用．在年幼的啮齿类动物中,小脑包含5种不同的集中分布层：白质、内颗粒细胞层(IGL)、浦肯野氏细胞层(PCL)、分子层(ML)和外颗粒细胞层(EGL)．与浦肯野氏神经元在出生前产生这一点不同的是,EGL中的细胞在出生后产生,它们接受从前脑olivary核团发出的攀援纤维的主要神经投射,以及从内颗粒细胞发出的平行纤维的神经投射．这些神经投射主要在出生后的前3个星期内建立,同时还有浦肯野氏细胞的发育和成熟．而GAP-43,netrin-1,collapsin-1和neuropilin-1在出生后小脑发育的潜在作用仍然不清楚．为了更加清楚地探讨上述问题,检验了GAP-43,netrin-1,collapsin-1和neuropilin-1的mRNA与蛋白质在出生后5,10,20天和成年小鼠小脑中的表达情况．研究结果显示,这4种分子在小鼠出生后的小脑中有不同的时间和空间表达形式,这些结果与出生后发育和成年期间的轴突发生、延伸以及突触形成都有关联．通过免疫组织化学双标染色,发现小鼠出生后10天的小脑中,GAP-43阳性的浦肯野氏细胞也显示netrin-1或collapsin-1阳性,并且collapsin-1阳性的细胞也对 netrin-1 阳性．上述研究结果证明这4种分子可能参与了小脑的出生后发育．     

Death domain complex of the TNFR-1, TRADD,and RIP1 proteins for death-inducing signaling

Apoptosis can be induced by an extrinsic pathway involving the ligand-mediated activation of death receptors such as tumor necrosis factor receptor-1 (TNFR-1). TNFR-1-associated death domain (TRADD) protein is an adapter molecule that bridges the interaction between TNFR-1 and receptor-interacting serine/threonine-protein kinase 1 (RIP1). However, the molecular mechanism of the complex formation of these proteins has not yet been identified. Here, the binding among TNFR-1, TRADD, and RIP1 was identified using a GST pull-down assay and Biacore biosensor experiment. This study showed that structural characterization and formation of the death-signaling complex could be predicted using TNFR-1, TRADD, and RIP1. In addition, we found that the structure-based mutations of TNFR-1 (P367A and P368A), TRADD (F266A), and RIP1 (M637A and R638A) disrupted formation of the death domain (DD) complex and prevented stable interactions among those DDs.     

Expression of KCNQ1OT1, CDKN1C,H19, and PLAGL1 and the methylation patterns at the KvDMR1 and H19/IGF2 imprinting control regions is conserved between human and bovine

Background

Beckwith-Wiedemann syndrome (BWS) is a loss-of-imprinting pediatric overgrowth syndrome. The primary features of BWS include macrosomia, macroglossia, and abdominal wall defects. Secondary features that are frequently observed in BWS patients are hypoglycemia, nevus flammeus, polyhydramnios, visceromegaly, hemihyperplasia, cardiac malformations, and difficulty breathing. BWS is speculated to occur primarily as the result of the misregulation of imprinted genes associated with two clusters on chromosome 11p15.5, namely the KvDMR1 and H19/IGF2. A similar overgrowth phenotype is observed in bovine and ovine as a result of embryo culture. In ruminants this syndrome is known as large offspring syndrome (LOS). The phenotypes associated with LOS are increased birth weight, visceromegaly, skeletal defects, hypoglycemia, polyhydramnios, and breathing difficulties. Even though phenotypic similarities exist between the two syndromes, whether the two syndromes are epigenetically similar is unknown. In this study we use control Bos taurus indicus X Bos taurus taurus F1 hybrid bovine concepti to characterize baseline imprinted gene expression and DNA methylation status of imprinted domains known to be misregulated in BWS. This work is intended to be the first step in a series of experiments aimed at determining if LOS will serve as an appropriate animal model to study BWS.

Results

The use of F1 B. t. indicus x B. t. taurus tissues provided us with a tool to unequivocally determine imprinted status of the regions of interest in our study. We found that imprinting is conserved between the bovine and human in imprinted genes known to be associated with BWS. KCNQ1OT1 and PLAGL1 were paternally-expressed while CDKN1C and H19 were maternally-expressed in B. t. indicus x B. t. taurus F1 concepti. We also show that in bovids, differential methylation exists at the KvDMR1 and H19/IGF2 ICRs.

Conclusions

Based on these findings we conclude that the imprinted gene expression of KCNQ1OT1, CDKN1C, H19, and PLAGL1 and the methylation patterns at the KvDMR1 and H19/IGF2 ICRs are conserved between human and bovine. Future work will determine if LOS is associated with misregulation at these imprinted loci, similarly to what has been observed for BWS.     

Integrity and Function of the Saccharomyces cerevisiae Spindle Pole Body Depends on Connections Between the Membrane Proteins Ndc1, Rtn1, and Yop1

The nuclear envelope in Saccharomyces cerevisiae harbors two essential macromolecular protein assemblies: the nuclear pore complexes (NPCs) that enable nucleocytoplasmic transport, and the spindle pole bodies (SPBs) that mediate chromosome segregation. Previously, based on metazoan and budding yeast studies, we reported that reticulons and Yop1/DP1 play a role in the early steps of de novo NPC assembly. Here, we examined if Rtn1 and Yop1 are required for SPB function in S. cerevisiae. Electron microscopy of rtn1Δ yop1Δ cells revealed lobular abnormalities in SPB structure. Using an assay that monitors lateral expansion of the SPB central layer, we found that rtn1Δ yop1Δ SPBs had decreased connections to the NE compared to wild type, suggesting that SPBs are less stable in the NE. Furthermore, large budded rtn1Δ yop1Δ cells exhibited a high incidence of short mitotic spindles, which were frequently misoriented with respect to the mother–daughter axis. This correlated with cytoplasmic microtubule defects. We found that overexpression of the SPB insertion factors NDC1, MPS2, or BBP1 rescued the SPB defects observed in rtn1Δ yop1Δ cells. However, only overexpression of NDC1, which is also required for NPC biogenesis, rescued both the SPB and NPC associated defects. Rtn1 and Yop1 also physically interacted with Ndc1 and other NPC membrane proteins. We propose that NPC and SPB biogenesis are altered in cells lacking Rtn1 and Yop1 due to competition between these complexes for Ndc1, an essential common component of both NPCs and SPBs.     

Gene expression of LOX-1, VCAM-1, and ICAM-1 in pre-atherosclerotic mice

To identify markers of the earliest stage of atherosclerosis, endothelial dysfunction, we evaluated the gene expression of lectin-like oxidized-low-density-lipoprotein receptor-1 (LOX-1), vascular cell adhesion molecule-1 (VCAM-1), and intercellular adhesion molecule-1 (ICAM-1) in very young pre-atherosclerotic mice. Furthermore, the plasma levels of the soluble VCAM-1 and ICAM-1 were compared to the gene expression profiles. Gene expressions of LOX-1 and VCAM-1 were up-regulated in young apoE^−/− mice, and thus, it seems probable that these genes play a role in pre-atherosclerosis. Contrarily, the gene expression profile of ICAM-1 did not show any apparent differences between the groups, questioning the involvement of this molecule in the early development of atherosclerosis. Plasma levels of sVCAM-1 and sICAM-1 were similar in all mice and did not correlate with the vascular gene expression of the corresponding genes. It therefore seems likely that these circulating markers are not suited to detect early atherosclerosis.     

Nod1, a CARD protein,enhances pro-interleukin-1beta processing through the interaction with pro-caspase-1

The production of bioactive interleukin-1beta (IL-1beta), a pro-inflammatory cytokine, is mediated by activated caspase-1. One of the known molecular mechanisms underlying pro-caspase-1 processing and activation involves interaction between the caspase recruit domains (CARDs) of caspase-1 and a serine/threonine kinase RIP2. While the association of Nod1 with both caspase-1 and RIP2 is already known, the consequences of these interactions are poorly understood. Because Nod1 also binds to RIP2, we hypothesized that Nod1 plays a role in pro-caspase-1 activation and IL-1beta processing. We show here that Nod1 binds to both RIP2 and caspase-1 by CARD interactions. Nod1 enhances pro-caspase-1 oligomerization and pro-caspase-1 processing. Nod1 enhances caspase-1-induced IL-1beta secretion, as well as lipopolysaccharide (LPS)-induced IL-1beta secretion in transfected cells. Moreover, HT1080 cells stably transfected with Nod1 showed higher LPS-induced IL-1beta secretion than non-transfected cells, suggesting a role of Nod1 in LPS-induced responses. Our data indicate that Nod1 can regulate IL-1beta secretion, implying that Nod1 may play a role in inflammatory responses to bacterial LPS.     

The Cytosolic J-protein, Jjj1, and Rei1 Function in the Removal of the Pre-60 S Subunit Factor Arx1

Although the biogenesis of ribosomal subunits occurs predominantly in the nucleus, final remodeling steps take place in the cytosol. One cytosolic step has two components: 1) the removal of the maturation factor Arx1, which transits from the nucleus to the cytosol with the pre-60 S subunit, and 2) its subsequent transport back into the nucleus. Two cytosolic proteins, Rei1 and Jjj1, are required, but their individual contributions to this step are not understood. Here we report that Rei1 and Jjj1 directly interact. This interaction is mediated by a C-terminal segment of Jjj1 encompassing a region rich in charged residues, flanked by C₂H₂-type zinc fingers. Deletion of the charged region results in defects in 60 S subunit biogenesis in vivo. In addition, we report resolution of an apparent contradiction in the literature regarding the association of Arx1 with the pre-60 S subunit in the absence of Rei1. The association of Arx1 with ribosomes is sensitive to the concentration of magnesium ions when Rei1 is absent. At near physiological concentrations, Arx1 remains associated with the pre-60 S particle, as it does in the absence of Jjj1; at higher concentrations, Arx1 dissociates in the absence of Rei1 but not in the absence of Jjj1. As both Rei1 and Jjj1 are required for dissociation of Arx1 from the pre-60 S subunit, and the region of Jjj1 that mediates interaction with Rei1 is required in vivo for 60 S subunit biogenesis, our data support the idea that the primary role of both Rei1 and Jjj1 is the first step of the Arx1 removal/recycling process.