雄性不育高粱线粒体与细胞核对热激的反应及其与育性关系的研究

由于我们发现热激(40℃)能诱导雄性不育高粱转变为雄性可育型的现象,故进一步探讨此现象的机理,研究了在热激条件下其细胞质(取其细胞质重要遗传物质——线粒体)中及细胞核中蛋白质的变化。试验结果,发现不育系的花粉母细胞期幼穗,在热激时细胞核中出现特异的80KD热激蛋白,且能被利福霉素、氯霉素所抑制。保持系在热激时细胞核中未出现此蛋白。80KD蛋白在结合部位存在的性质与其存在于细胞核中和线粒体中的现象是一致的。这种差异说明与雄性不育有关,且其原因是由细胞核与细胞质共同作用所致。     

雄性不育高粱四种酶在热激过程中的反应

热激时,雄性不育高梁中4种酶的同工酶的变化以细胞色素氧化酶最大,酸性磷酸酯酶次之,淀粉酶第三,酯酶最小,说明不同酶对热激的反应不同,故与育性的关系也有差异。苗期与穗期同工酶谱互不对应,表明酶随发育而改变。不育系的细胞色素氧化酶酶带C5,C8随温度上升而减弱,达40℃时与可育的保持系相似,此现象与不育系的雄蕊在40℃时呈现可育态同时发生,说明此酶带与育性相关。     

高粱热激蛋白(HSPs)的电泳分析与雄性不育性

本文以高粱雄性不育系为材料进行了热激蛋白的研究。苗期单向电泳表明,保持系在热激(40℃)过程中可溶性蛋白有24条带,对照(28℃)22条,出现2条带差异,不育系在热激后有33条带,比对照28条多5条,且有4条加强带,共9条带产生差异。苗期双向电泳,不育系在热激后比对照有19点产生差异,保持系热激后与对照比较有6点产生差异。表明双向电泳揭示了热激后蛋白质在分子量和电性方面有差异。花粉母细胞期幼穗的热激蛋白,不育系对照可溶性蛋白仅有3条带,热激后有11条,8条带有差异。保持系对照有10条带,热激后有15条,1条加强,共6条带有差异。幼穗期不育系蛋白质突出的缺少19kd以上的大分子量的带。从个体发育看,不育系由苗期可溶性蛋白比保持系带数多,发育到花粉母细胞期比保持系突出地减少,尤其是大分子量的蛋白带消失,热激后明显地增加了与可育的保持系相同的蛋白成份,表明不育系有其特殊的基因调控。     

高粱3197A雄性不育系热激处理后代的可育性遗传

对高梁细胞质雄性不育系3197A经热激处理后获得的可育结实后代进行了连续多代自交及回交观察,结果表明,由热激诱导的可育性变化不能稳定遗传。从H2代开始,其可育率逐代降低。到H5代,分离出的可育株数仅占观察植株总数的1．9％。H5代可育株与不育株花药总RNA与HSP70热激蛋白基因探针Northern杂交结果表明,H3A可育株均能与HSP70基因探针杂交,显示较强的杂交信号。而其不育株,有的完全不能与HSP70基因探针杂交,有的显示微弱的杂交信号。说明3197A雄性不育系热激可育后代的遗传稳定性与HSP70基因的转录状况有关。     

温度,热激蛋白与高粱育性的变化

高粱不育系3A在热激(43～45℃)诱导下结出了种子,由不育系转变为可育系。比较3A和3B线粒体在热激条件的热激蛋白得知,它们的热激蛋白是由核编码的,在细胞质中合成后才运到线粒体中,热激2h,3A出现70、31、24、18和16kDa 5条蛋白带,3B除出现上述5条蛋白带外还多出现96、94kDa 2条,而且70kDa含量比3A大。热激4h,3B的96、94kDa消失,两系趋于一致。此时,3A和3B线粒体总蛋白比热激前大量增加。此后HSPs急剧降低。热激8h,3B线粒体仅有70、31、24和16kDa 4条蛋白带,70kDa特别明显,而3A则全部消失。从而表明,HSPs在3B中是稳定的,在3A中是缺乏或不稳定的,这些差异可能与3B育性稳定性及3A不育性有关。     

高粱雄性不育系热激前后线粒体的变化与育性的关系

高粱雄性不育系3197A（3A）在花粉母细胞期进行热激处理后由不育变为可育。提取其处理小穗线粒体用流式细胞仪计数进行分析。结果发现经热激处理后,3A不育系的线粒体数目由1.55×106个／mg鲜穗增加至7.1×106／mg。同时对热激处理后3A的花粉育性及同工酶进行了研究,发现热激后3A的花粉粒变的饱满并可被I2朘I 溶液染色,其细胞色素氧化酶和过氧化物酶的电泳酶谱中出现与保持系3B一致的酶带。     

小麦T型细胞质雄性不育系、保持系蛋白质双向电泳比较研究

以小麦T型细胞质雄性不育系为材料,利用双向电泳技术,对苗期、分蘖期、拔节期和孕穗期叶片和花粉母细胞减数分裂期、单核小孢子期、二—三核小孢子期蛋白质变化作了分析。在细胞质雄性不育系小麦拔节期、孕穗期叶片中,有一个33KD/PI6.3蛋白组分存在,保持系中没有发现这个蛋白组分。在花粉败育的关键时期二—三核小孢子期,小麦细胞质雄性不育系有53KD/PI5.5、50KD/PI5.7、48KD/PI5.6和20KD/PI7.5四种蛋白组分存在,而保持系中也没有存在。小麦细胞质雄性不育系叶片和小孢子发育过程中存在的这五种特异蛋白可能参与育性调控,与细胞质雄性不育特性的形成有关。     

热激处理与水稻雄性不育系育性转变关系的初步研究

本实验对水稻雄性不育系珍汕97A及其保持系在幼苗(2叶或3叶)和孕穗期(抽穗前8天)分别进行一次热激处理,并比较了其同工酶组成、花粉育性以及自交结实性的变化。同工酶分析结果表明,热激处理能使不育系与保持系花药的POX和Est的表达发生不同的改变,导致两系在这两种同工酶组成上的原有差异缩小;育性观察结果显示,在正常温度条件下,热激处理对珍汕97A的育性表达几乎无影响,而在有高温作用(穗分化发育阶段的日平均温度高于30℃)的条件下,3叶期幼苗往后的热激处理似乎能不同程度地提高珍汕97A的育性稳定性。简要讨论了热激处理与不育系育性转变间的关系。     

高梁雄性不育系热激前后线粒体的变化与育性的关系

高梁雄性不育系3197A(3A)在花粉母细胞期进行热激处理后由不育变为可育。提取其处理小穗线粒体用流式细胞仪计数进行分析。结果发现经热激处理后,3A不育系的线粒体数目由1.55×106个/mg鲜穗增加至7.1×106/mg。同时对热激处理后3A的花粉育性及同工酶进行了研究,发现热激后3A的花粉粒变的饱满并可被I2-KI溶液染色,其细胞色素氧化酶和过氧化物酶的电泳酶谱中出现与保持系3B一致的酶带。     

高温处理对高粱雄性不育系育性的影响

高粱雄性不育系植株被高温激发产生可育的黄花药的现象表明,这些植株本身已含有产生可育黄花药的物质和雄性可育基因的存在,但可能被调节因子所抑制。对花药的同工酶电泳分析得到了证明。雄性不育系的黄花药和白花药的酶谱,在单核花粉粒时期有与雄性可育系植株(保持系)相同形式的区带,当发育到三核花粉时期,白花药的某些区带消失,而黄花药则保留了与雄性可育系植株相同的区带。试验还指明了出现黄花药的临界温度为39—40℃和花粉母细胞时期为其对温度的敏感期。     

金属离子诱导高粱雄性不育系、保持系热激反应的研究

本试验以高粱雄性不育系和保持系为材料进行金属离子诱导应激反应的研究。结果表明CuSO_4溶液诱导高粱雄性不育系产生的应激反应明显,所用CuSO_4溶液最适浓度为250pmol/L,最适培养时间为4小时。ZnSO_4溶液诱导反应不明显。电泳分析和自显影检测出CuSO_4诱导产生新蛋白区带在3197A为9条,3197B为20条。但各蛋白带产生所需浓度和时间不完全相同,表明各种蛋白的基因表达是相互独立的,且这些基因的表达随发育而有变化。从分析处理后的4种酶的反应可以看出,其中细胞色素氧化酶有5条带,过氧化物酶反应灵敏,但只有2条带。通过CuSO_4处理与热激处理比较,推测3197A、3197B有其特殊的基因调控系统。从产生的可溶性蛋白或酶来看,CuSO_4能使3197A产生的应激蛋白趋向于3197B,说明两者具有相似之处。     

Heat Shock Stress Causes Stage-specific Male Sterility in Arabidopsis thaliana

Arabidopsis during flower development, floral organs such as sepals, petals, stamens, and carpels developed normally. However, the development of pollen inside the anther was disrupted in a stage-specific manner, with floral stage 9 primordia failing to produce any pollen grains. Morphological analyses suggested that heat shock causes a failure of separation of pollen mother cells followed by microspore differentiation and/or inhibition of male meiotic processes. Heat shock also caused sterility in floral stage 12 flowers but the sterility was due to the failure of pollen release from the pollen sacs. Received 12 December 2000/ Accepted in revised form 4 April 2001     

Acute heat stress prior to downhill running may enhance skeletal muscle remodeling

Heat shock proteins (HSPs) are chaperones that are known to have important roles in facilitating protein synthesis, protein assembly and cellular protection. While HSPs are known to be induced by damaging exercise, little is known about how HSPs actually mediate skeletal muscle adaption to exercise. The purpose of this study was to determine the effects of a heat shock pretreatment and the ensuing increase in HSP expression on early remodeling and signaling (2 and 48 h) events of the soleus (Sol) muscle following a bout of downhill running. Male Wistar rats (10 weeks old) were randomly assigned to control, eccentric exercise (EE; downhill running) or heat shock + eccentric exercise (HS; 41°C for 20 min, 48 h prior to exercise) groups. Markers of muscle damage, muscle regeneration and intracellular signaling were assessed. The phosphorylation (p) of HSP25, Akt, p70s6k, ERK1/2 and JNK proteins was also performed. As expected, following exercise the EE group had increased creatine kinase (CK; 2 h) and mononuclear cell infiltration (48 h) compared to controls. The EE group had an increase in p-HSP25, but there was no change in HSP72 expression, total protein concentration, or neonatal MHC content. Additionally, the EE group had increased p-p70s6k, p-ERK1/2, and p-JNK (2 h) compared to controls; however no changes in p-Akt were seen. In contrast, the HS group had reduced CK (2 h) and mononuclear cell infiltration (48 h) compared to EE. Moreover, the HS group had increased HSP72 content (2 and 48 h), total protein concentration (48 h), neonatal MHC content (2 and 48 h), p-HSP25 and p-p70s6k (2 h). Lastly, the HS group had reduced p-Akt (48 h) and p-ERK1/2 (2 h). These data suggest that heat shock pretreatment and/or the ensuing HSP72 response may protect against muscle damage, and enhance increases in total protein and neonatal MHC content following exercise. These changes appear to be independent of Akt and MAPK signaling pathways.     

Response of Higher Plants to Heat Shock

When sorghum seedlings were rapidly shifted from the cultural temperature of 30℃ to 40℃ and 45℃, a set of abnormal proteins, generally referred to as heat shock proteins were induced. They are a group of high molecular weight proteins (about 66–117 kD), a few intermediate molecular weight proteins (33–66kD) and a low molecular weight protein of 18 kD. At the same time, the synthesis of normal proteins was relatively depressed. The res ponse of the shoot tissues of sorghum seedings to heat shock is similar to that of the root tissues, but there are some differences in more detail between the two tissues. The synthesis of heat shock proteins in sorghum seedlings was rapid. After one-hour exposure at 45℃ their synthesis in the roots was detectable. Maximum induction took place in the second hour of exposure, thereafter their synthesis began to decline markedly. Finally, there appear to be some proteins whose synthesis was not supressed during heat shock, It is not yet known why the synthesis of these proteins is so stable.