中红侧沟茧蜂滞育诱导和滞育茧的冷藏

中红侧沟茧蜂Microplitis mediator (Haliday)是夜蛾科害虫低龄幼虫的重要寄生蜂。田间实验表明,在冀中地区秋季田间条件下,当日平均气温为21.5℃、日平均光照时间为12 h 33 min时,少数个体进入滞育;当气温降至17.9℃以下、日光照时间缩短到11 h 45 min以下时,全部个体进入滞育。室内模拟实验结果表明,在17～26℃、光照时间10～14 h范围内,随着温度的降低和光照时间的缩短,滞育率明显提高。高温能抵消短光照对滞育诱导的影响,在26℃下,短光照不能诱导滞育。因此,低温和短光照是诱导该种天敌昆虫滞育的主要因子。中红侧沟茧蜂感受滞育信号的敏感期为低龄幼虫期,以预蛹（茧）进入滞育。低龄幼虫感受滞育信号以后,需要在滞育环境中发育到老熟幼虫才能全部进入滞育。将室内诱导的滞育茧在4℃左右环境条件下冷藏240天,成蜂的羽化率和寄生能力与没有冷藏的非滞育茧差异不显著;冷藏300天,滞育茧仍有81.4%可以正常羽化。本项研究结果为中红侧沟茧蜂的规模化、标准化生产提供了科学的依据。     

温度和光周期对管侧沟茧蜂滞育诱导及滞育茧的低温冷藏

【目的】为明确诱导管侧沟茧蜂Microplitis tuberculifer 滞育的主要因子,在田间和室内研究了不同温度和光周期下管侧沟茧蜂的滞育率和滞育茧的最佳冷藏温度。【方法】田间实验分别从8月31日到9月25日每隔5 d在室外罩笼内释放管侧沟茧蜂寄生的粘虫幼虫,待寄生蜂结茧后统计子代蜂的滞育率。室内实验共设5个不同温度（16℃, 18℃, 20℃, 22℃和24℃）和7个不同光周期（6L:18D, 8L:16D, 10L:14D, 12L:12D, 14L:10D, 16L:8D和18L:6D）,分别统计寄生蜂在各个处理条件下的滞育率。【结果】在河北中部地区秋季自然条件下,8月底当日平均气温为24.4℃,日平均光照时间为12 h 51 min 时,少数蛹（5.08%）开始进入滞育;9月25日,当日平均气温为17.2℃,日平均光照时间为11 h 36 min以下时,几乎所有蛹个体进入滞育,滞育率达到99.70%。在室内人工控制条件下,当温度为22℃以上,无论光周期如何变化,管侧沟茧蜂不能进入滞育,所结茧全部为非滞育茧。温度为22℃以下存在光周期反应,在温度16℃, 18℃和20℃,光周期10L:14D时形成滞育茧数量最多,滞育率分别为100%, 89.75% 和 29.58%。可见温度和光周期二者共同影响管侧沟茧蜂的滞育。滞育茧在0℃左右条件下冷藏 240 d 和5℃左右环境条件下冷藏180 d, 成虫的羽化率和寄生能力与发育茧差异不显著(P>0.05);0℃条件下冷藏300 d,滞育茧仍有79%可以正常羽化。【结论】该寄生蜂在秋季进入滞育主要是低温条件和短光照反应相互作用的结果,滞育茧在0℃和5℃温度下至少可以保存240 d。这些结果对管侧沟茧蜂的大规模繁殖和滞育茧的保存具有重要参考价值。     

低温对麦红吸浆虫滞育解除及蜕皮激素受体基因EcR和热激蛋白基因Hsp70和Hsp90表达水平的影响

【目的】本研究旨在明确破茧率和破茧所需时间作为典型的专性幼虫期滞育昆虫麦红吸浆虫Sitodiplosis mosellana滞育解除指标的可行性,探讨蜕皮激素受体基因EcR和热激蛋白基因Hsp70和Hsp90在低温解除滞育中的作用。【方法】9月上旬采自田间的麦红吸浆虫滞育幼虫在低温(4℃)和自然变温处理不同时间(0~90 d)后转移至24℃化蛹,调查幼虫的破茧率、化蛹率及破茧和化蛹所需时间;采用qPCR技术分析4℃低温终止滞育并在24℃下恢复发育的幼虫EcR, Hsp70和Hsp90的mRNA水平。【结果】不同时间低温(4℃)和自然变温处理的麦红吸浆虫滞育幼虫破茧率和化蛹率以及破茧和化蛹所需时间均存在显著差异。未经低温(4℃)处理的幼虫在水分满足的条件下破茧率为71.3%,但均不能化蛹;低温(4℃)处理60 d内,随着处理时间的延长破茧率和化蛹率逐渐升高,破茧和化蛹所需时间逐渐缩短;与4℃低温处理30~60 d相比,自然变温处理30~60 d的幼虫化蛹率显著较低;4℃低温处理60 d和自然变温处理90 d后幼虫的化蛹率均超过91%。低温(4℃)处理显著提高了麦红吸浆虫幼虫EcR, Hsp70和Hsp90的表达量,其中处理30 d时其表达量最高,随着低温(4℃)处理时间的延长,表达量逐渐降低,大部分幼虫滞育解除后表达量趋于恒定。【结论】低温能显著促进麦红吸浆虫的滞育解除,效果优于9-10月自然变温;破茧率及破茧所需时间可作为评判麦红吸浆虫幼虫滞育强度的参考,但不能独立作为滞育解除的指标;EcR,Hsp70和Hsp90的表达水平与滞育强度密切相关,在麦红吸浆虫滞育解除中发挥潜在作用。     

中红侧沟茧蜂滞育临界光周期和敏感光照虫态的测定

室内对中红侧沟茧蜂Microplitis mediator滞育诱导的光周期反应及敏感光照虫态进行了研究。结果表明：在16℃和18℃温度下,当光周期为10L∶14D时,中红侧沟茧蜂的滞育率最高为98%和87.8%;其次是光周期为8L∶16D时,滞育率为78.7%和73%;光周期为12L∶12D时,滞育率为55.8%和50.8%;而在全光照LL, 光周期22L∶2D,20L∶4D,18L∶6D,16L∶8D,14L∶10D ,2L∶22D和全黑暗DD时的滞育率均为0。中红侧沟茧蜂属于典型的长日照发育型,在16℃时诱导滞育的临界光周期是7.03L∶16.97D和12.21L∶11.79D;在18℃时是6.75L∶17.25D和12.03L∶11.97D。16℃时中红侧沟茧蜂对光照反应最敏感时期为2龄幼虫第5～8天,2龄幼虫第1～4,9～12和13～16天及3龄期次之,卵期和1龄幼虫不敏感。     

斑痣悬茧蜂在黏虫寄主体内与中红侧沟茧蜂竞争下的存活和发育表现

【目的】不同种寄生蜂在同一小生境中可能寄生同一共享寄主。本研究旨在为认识寄生蜂种间互作关系,并为科学评价寄生性天敌控害效应提供依据。【方法】在室内以单寄生性斑痣悬茧蜂Meteorus pulchricornis(Wesmael)和中红侧沟茧蜂Microplitis mediator(Halidy)及其寄主黏虫Mythimna separata(Walker)低龄幼虫为材料,以寄生顺序和间隔时间为因素进行种间寄生竞争效应实验,观察比较两种蜂的存活和斑痣悬茧蜂子代的发育历期、体型大小和寿命等发育参数。【结果】不同寄生顺序和间隔时间组合处理下斑痣悬茧蜂子代蜂存活率始终高于中红侧沟茧蜂,且先寄生的斑痣悬茧蜂子代存活率高于后寄生的蜂,提前24 h寄生的斑痣悬茧蜂子代存活率可高达96.7%。当斑痣悬茧蜂先于中红侧沟茧蜂寄生时,其子代幼虫发育历期比单独寄生(对照)时显著延长;当后于中红侧沟茧蜂寄生时,其子代幼虫发育历期与单独寄生时无显著差异。各竞争处理下,斑痣悬茧蜂子代成虫寿命均长于单独寄生的子代成虫;子代成虫后足胫节长度与单独寄生相比明显缩短。【结论】结果说明,斑痣悬茧蜂在黏虫低龄幼虫体内与中红侧沟茧蜂的竞争中占有优势,但竞争对斑痣悬茧蜂子代生长发育具有负面影响。     

中红侧沟茧蜂对烟草挥发物的触角电生理及行为反应

中红侧沟茧峰 Microplitis mediator是棉铃虫 H elicoverpa armigera的主要内寄生蜂 ,是优良的蜂种 ,在生物防治上具有广阔的应用前景。但迄今 ,有关中红侧沟茧蜂的化学生态的研究甚少。利用触角电位技术 ,测定了中红侧沟茧蜂对烟草叶片粗提物和烟草中 10种挥发物的触角电生理反应 ,并测定了中红侧沟茧蜂的触角电位反应随日龄的变化。以嗅觉仪测定了中红侧沟茧蜂对烟草粗提物、烟草中 4种挥发物的行为反应。结果表明中红侧沟茧蜂利用烟草释放的气味向烟草定向 ,进而找到其寄主——棉铃虫 ,也佐证了烟草同样是棉铃虫的重要寄主植物的论点。雌蜂对脂肪族化合物反应最强 ,对芳香族化合物和糠醛 (杂环化合物 )的反应居中 ,对萜类化合物反应较弱或者不反应。表明烟草挥发性物质在中红侧沟茧蜂寻找寄主的过程中起着重要的作用 ,但挥发物各组分的作用存在着差异。雌性中红侧沟茧蜂对顺 - 3-己烯醇的触角电位反应随日龄的增加而变化。 1日龄的雌蜂的反应较弱 ,2日龄、3日龄的雌蜂反应较强 ,4日龄、5日龄的雌蜂的反应又减弱。这与该蜂产卵习性相符 :羽化当天的雌蜂产卵较少 ,羽化 2～ 3d之后的雌蜂产卵量较大     

温度胁迫下棉花粉蚧热激蛋白基因的表达分析

【目的】为了研究热激蛋白 Hsp70, Hsp70-4和Hsp90在棉花粉蚧Phenacoccus solenopsis抵抗逆温中的作用。【方法】在测序棉花粉蚧转录组的基础上,分析了该虫热激蛋白Hsp70基因家族的2个序列［Pshsp70（GenBank登录号为KJ909505）和Pshsp70-4（GenBank登录号为KJ909506）］和Hsp90基因家族的1个序列,［Pshsp90(GenBank登录号为KJ909507)］,采用实时荧光定量 PCR（RT-qPCR）检测了在不同温度（18和32℃恒温, 37, 39, 41, 43和45℃热激1 h 后26℃恢复1 h)下棉花粉蚧不同发育阶段（2龄若虫、3龄若虫、雌成虫）3种热激蛋白基因的表达量。【结果】Pshsp70 cDNA序列包含1 923 bp的开放阅读框,编码641个氨基酸,理论分子量和等电点分别为70.9 kDa和5.65; Pshsp70-4 cDNA序列包含1 962 bp的开放阅读框,编码654个氨基酸,理论分子量和等电点分别为71.8 kDa和5.38;Pshsp90 cDNA序列包含2 172 bp的开放阅读框,编码724个氨基酸,理论分子量和等电点分别为83.5 kDa和4.93。Pshsp70 和Pshsp70-4均含有Hsp70基因家族高度保守的基序,Pshsp70编码的氨基酸序列与烟粉虱Bemisia tabaci和家蚕Bombyx mori等昆虫的Hsp70 的氨基酸序列一致性为 85%;Pshsp70-4编码的氨基酸序列与白蜡蚧Ericerus pela和点蜂缘蝽Riptortus pedestris等昆虫的Hsp70的氨基酸序列一致性高达95%;Pshsp90也含有Hsp90基因家族高度保守的基序,Pshsp90编码的氨基酸序列与赤拟谷盗Tribolium castaneum和东亚小花蝽Orius sauteri等昆虫的Hsp90 的氨基酸序列一致性为 87%。热激蛋白基因表达量分析结果表明,在18℃恒温条件下,粉蚧2龄若虫的3个PsHsps基因的mRNA相对表达量均比对照(26℃)低,在32℃恒温条件下,各龄期的Hsp70基因的相对表达量均显著高于对照。在37~45℃下热激1 h并在26℃下恢复1 h,棉花粉蚧3个龄期的3个热激蛋白PsHsps基因的相对表达量随温度的升高总体呈增加趋势,相关性分析表明,除Pshsp70-4在雌成虫中的表达量与热胁迫温度的相关系数为0.225外,各龄期中3个基因的表达量与温度的相关系数均大于0.6,显著相关;43℃和45℃胁迫下,各龄期的3个热激蛋白基因相对表达量均显著高于对照组（P<0.05）。【结论】棉花粉蚧热激蛋白基因的表达与温度呈正相关,在该虫应对高温中起着重要作用。     

清道夫受体MmSCRA-1调控中红侧沟茧蜂发育的作用研究

清道夫受体(scavenger receptors,SCRs)是细胞表面的一类跨膜糖蛋白,参与识别多种病原体。中红侧沟茧蜂Micropilits mediator有13个清道夫受体基因,为探究其功能,本文通过系统进化分析,确定与其他物种进化关系,并运用实时荧光定量PCR (qRT-PCR)技术检测中红侧沟茧蜂不同发育时期SCRs基因mRNA的表达水平。选取表达水平较高的A型清道夫受体MmSCRA-1基因进行RNA干扰。结果表明,中红侧沟茧蜂蛹期3日龄注射dsRNA后,dsSCRA-1注射组羽化率显著低于dsGFP注射组(P 0. 05),蛹期3. 5日龄注射dsRNA后,两组羽化率都升高,但dsSCRA-1注射组羽化率仍显著低于dsGFP注射组(P 0. 05)。干扰中红侧沟茧蜂MmSCRA-1基因后,其羽化率降低,说明MmSCRA-1基因在中红侧沟茧蜂蛹期发育过程中起到关键作用。     

中红侧沟茧蜂寄生对寄主粘虫血淋巴糖类、脂类和蛋白质含量的影响

为揭示寄生蜂寄生对其寄主的生理调控机制, 室内对中红侧沟茧蜂Microplitis mediator寄生与未被寄生寄主粘虫Mythimna separata幼虫血淋巴中糖类、脂类和蛋白含量变化进行了测定。结果显示: 在滞育与非滞育条件下, 被寄生的粘虫血淋巴中糖原浓度均比未被寄生的粘虫高。滞育条件下寄生后12 d差异显著（P<0.05）, 被寄生粘虫糖原含量为7.93 μg/mL, 未被寄生粘虫糖原含量为4.70 μg/mL; 非滞育条件下寄生后6 d差异显著（P<0.05）, 被寄生粘虫糖原含量为14.35 μg/mL, 未被寄生粘虫糖原含量为5.47 μg/mL。海藻糖含量测定结果显示, 在滞育条件下寄生蜂对被寄生粘虫无明显影响, 而非滞育条件下影响效果差异显著（P<0.05）, 寄生后4 d被寄生粘虫海藻糖含量为46.82 μg/mL, 未被寄生粘虫含量为26.72 μg/mL。在滞育与非滞育两种条件下, 寄生与未被寄生寄主脂类和蛋白含量没有显著性差异。结果说明: 寄生蜂的存在使寄主血淋巴中的糖原含量增高; 非滞育条件是影响被寄生粘虫海藻糖含量变化主要因素; 粘虫对中红侧沟茧蜂的寄生表现相当强的适应性和忍受力。     

中红侧沟茧蜂寄生对棉铃虫幼虫行为的影响

【目的】中红侧沟茧蜂Microplitis mediator是棉铃虫Helicoverpa armigera生物防控的重要因子,通过温室内对未被寄生和被中红侧沟茧蜂寄生棉铃虫幼虫在棉花植株上的行为观察,分析中红侧沟茧蜂寄生对棉铃虫在棉花植株上行为活动及分布位置的影响。【方法】采用"固定时间间隔扫描抽样"观察法,监测寄生后1-10 d被寄生与未被寄生棉铃虫幼虫在棉花植株上的取食和空间位移等行为的变化。【结果】中红侧沟茧蜂寄生可以极显著降低棉铃虫幼虫在棉花植株上的取食行为比例(P0.01);被寄生棉铃虫幼虫在棉株上的空间位移比例于寄生后的1-2 d极显著高于未被寄生棉铃虫(P0.01),而在发育后期极显著低于未被寄生棉铃虫幼虫(P0.01);被寄生棉铃虫幼虫在棉铃上的分布比例显著降低而在花上的分布比例显著升高(P0.05);棉铃虫发育后期,被寄生棉铃虫在棉叶背面的分布比例极显著高于未被寄生棉铃虫(P0.01)。【结论】中红侧沟茧蜂寄生对棉铃虫幼虫的取食、活动和栖境选择具有显著影响。     

Heat shock protein synthesis over time in infective Trichinella spiralis larvae raised in suboptimal culture conditions

Changes in the viability, infectivity and heat shock protein (Hsp) levels are reported in Trichinella spiralis first stage larvae (L1) stored in 199 medium for up to seven days at 37 degrees C. These conditions induce stress that the larvae, eventually, cannot overcome. After three days of storage, the infectivity and viability were unchanged, although higher Hsp70 levels were observed. After this time, larvae gradually lost viability and infectivity, coinciding with a decrease in Hsp70 and Hsp90 and an increase in actin (a housekeeping protein). In addition, a possibly inducible heat shock protein, Hsp90i, appeared as constitutive Hsp90 disappeared. No significant changes in Hsp60 levels were detected at any time. These results suggest that heat shock proteins initially try to maintain homeostasis, but on failing, may be involved in cell death.     

Antioxidant responses of Chilo suppressalis (Lepidoptera: Pyralidae) larvae exposed to thermal stress

High temperatures cause a variety of physiological stress responses in insects, including increased generation of reactive oxygen species (ROS), which can cause oxidative damage. This study investigated the effects of thermal stress on ROS generation, the expression of heat shock protein 70 (Hsp70) at the mRNA and protein levels, the activity of antioxidant enzymes (SOD, CAT), and apoptosis in hemocytes of Chilo suppressalis larvae. Results indicated that thermal stress significantly elevated the level of ROS and antioxidant enzyme activity in C. suppressalis larvae. Real-time quantitative PCR showed that hsp70 gene expression was induced by heat stress. Flow cytometric results revealed that the expression profile of Hsp70 at the protein level was in agreement with that at the mRNA level. The expression of Hsp70 at both the mRNA and protein levels reached a maximum at 36 °C in larval hemocytes. Exposure to tested temperatures did not cause any significant change in the rate of apoptosis in larval hemocytes. These results suggest that thermal stress leads to oxidative stress and that antioxidant enzymes and the Hsp70 play an important role in reducing oxidative damage in C. suppressalis larvae.     

Pim-1 kinase stability is regulated by heat shock proteins and the ubiquitin-proteasome pathway

Elevated expression of the serine/threonine kinase Pim-1 increases the incidence of lymphomas in Pim-1 transgenic mice and has also been found to occur in some human cancers. Pim-1 acts as a cell survival factor and may prevent apoptosis in malignant cells. It was therefore of interest to understand to what extent maintenance and degradation of Pim-1 protein is affected by heat shock proteins (Hsp) and the ubiquitin-proteasome pathway in K562 and BV173 human leukemic cells. The half-life of Pim-1 protein in these cells was found to increase from 1.7 to 3.1 hours when induced by heat shock or by treating the cells with the proteasome inhibitor PS-341 (bortezomib). The Hsp90 inhibitor geldanamycin prevented the stabilization of Pim-1 by heat shock. Using immunoprecipitation, it was determined that Pim-1 is targeted for degradation by ubiquitin and that Hsp70 is associated with Pim-1 under these circumstances. Conversely, Hsp90 was found to protect Pim-1 from proteasomal degradation. A luminescence-based kinase assay showed that Pim-1 kinase bound to Hsp70 or Hsp90 remains active, emphasizing the importance of its overall cellular levels. This study shows how Pim-1 levels can be modulated in cells through degradation and stabilization.     

Starvation reduces the heat shock protein responses in white sturgeon larvae

This study investigates the responses of white sturgeon larvae (Acipenser transmontanus) to starvation and thermal stress, through the measurement of nutritional status (i.e. growth performances) and cellular biomarkers: heat shock proteins (Hsp) 70 and 90. White sturgeon larvae (25 day post hatch; initial weight 179.0 ± 5.1 mg) were fed (20% body weight per day) or starved for 24, 48 or 72 hrs. Every 24 hrs, five larvae from each of the starved or fed treatment replicates were exposed to heat shock resulting from an increase in water temperature from 19°C to 26°C, at a rate of 1°C per 15 min, and maintained at 26°C for 4 hrs. No mortality was observed in this study. Starvation significantly (p < 0.05) decreased the body weight and body contents of energy, protein, and lipid of the experimental larvae, compared to the fed larvae. Heat shock induced the expressions of Hsp70 and Hsp90 in both the fed and starved group; however, starvation reduced the induction at all sampling points. The current study demonstrates that poor larval nutritional status, assessed by the aforementioned parameters, reduced heat shock responses to thermal stress, as measured by heat shock protein levels. Furthermore, Hsp70 and 90 are more sensitive to heat shock and starvation, respectively. This may be, in part, a result of the different functioning of the heat shock proteins in cellular stress response and warrants further study.     

Heat shock response and mammal adaptation to high elevation (hypoxia)

The mammal’s high elevation (hypoxia) adaptation was studied by using the immunological and the molecular biological methods to understand the significance of Hsp (hypoxia) adaptation in the organic high elevation, through the mammal heat shock response. (1) From high elevation to low elevation (natural hypoxia): Westem blot and conventional RT-PCR and real-time fluorescence quota PCR were adopted. Expression difference of heat shock protein of 70 (Hsp70) and natural expression of brain tissue of Hsp70 gene was determined in the cardiac muscle tissue among the different elevation mammals (yak). (2)From low elevation to high elevation (hypoxia induction): The mammals (domestic rabbits) from the low elevation were sent directly to the areas with different high elevations like 2300, 3300 and 5000 m above sea level to be raised for a period of 3 weeks before being slaughtered and the genetic inductive expression of the brain tissue of Hsp70 was determined with RT-PCR. The result indicated that all of the mammals at different elevations possessed their heat shock response gene. Hsp70 of the high elevation mammal rose abruptly under stress and might be induced to come into being by high elevation (hypoxia). The speedy synthesis of Hsp70 in the process of heat shock response is suitable to maintain the cells’ normal physiological functions under stress. The Hsp70 has its threshold value. The altitude of 5000 m above sea level is the best condition for the heat shock response, and it starts to reduce when the altitude is over 6000 m above sea level. The Hsp70 production quantity and the cell hypoxia bearing capacity have their direct ratio.     

Heat shock response and mammal adaptation to high elevation (hypoxia)

The mammal's high elevation(hypoxia) adaptation was studied by using the immu-nological and the molecular biological methods to understand the significance of Hsp(hypoxia) ad-aptation in the organic high elevation,through the mammal heat shock response.(1) From high ele-vation to low elevation(natural hypoxia) :Western blot and conventional RT-PCR and real-time fluo-rescence quota PCR were adopted.Expression difference of heat shock protein of 70(Hsp70) and natural expression of brain tissue of Hsp70 gene was determined in the cardiac muscle tissue among the different elevation mammals(yak) .(2) From low elevation to high elevation(hypoxia induction) :The mammals(domestic rabbits) from the low elevation were sent directly to the areas with different high elevations like 2300,3300 and 5000 m above sea level to be raised for a period of 3 weeks be-fore being slaughtered and the genetic inductive expression of the brain tissue of Hsp70 was deter-mined with RT-PCR.The result indicated that all of the mammals at different elevations possessed their heat shock response gene.Hsp70 of the high elevation mammal rose abruptly under stress and might be induced to come into being by high elevation(hypoxia) .The speedy synthesis of Hsp70 in the process of heat shock response is suitable to maintain the cells' normal physiological functions under stress.The Hsp70 has its threshold value.The altitude of 5000 m above sea level is the best condition for the heat shock response,and it starts to reduce when the altitude is over 6000 m above sea level.The Hsp70 production quantity and the cell hypoxia bearing capacity have their direct ratio.     

Evidence that the novobiocin-sensitive ATP-binding site of the heat shock protein 90 (hsp90) is necessary for its autophosphorylation

The 90kDa heat shock protein (Hsp90) is one of the most abundant protein and essential for all eukaryotic cells. Many proteins require the interaction with Hsp90 for proper function. Upon heat stress the expression level of Hsp90 is even enhanced. It is assumed, that under these conditions Hsp90 is required to protect other proteins from aggregation. One property of Hsp90 is its ability to undergo autophosphorylation. The N-terminal domain of Hsp90 has been shown to contain an unusual ATP-binding site. A well-known inhibitor of Hsp90 function is geldanamycin binding to the N-terminal ATP-binding site with high affinity. Recently it was shown that Hsp90 possesses a second ATP-binding site in the C-terminal region, which can be competed with novobiocin. Autophosphorylation of Hsp90 was analysed by incubation with gamma(32)P-ATP. Addition of geldanamycin did not interfere with the capability for autophosphorylation, while novobiocin indeed did. These results suggest that the C-terminal ATP-binding site is required for autophosphorylation of Hsp90.