LEXEN: Microbial Adaptation to High Temperature and Pressure

超嗜热微生物可能是祖先甚至外星生物系统的合适模型，因为它们非凡的适应能力使它们能够在地球上大范围的地下火山区域定居。深海热液喷口和陆地温泉在无法获得太阳能的情况下支持高度多样化的生态系统，在与火星或木卫二表面以下可能存在的地球化学条件类似的地球化学条件下，栖息着自养微生物及其依赖的异养微观和宏观群落。超嗜热菌还主导了通用系统发育树的大部分深层分支，这表明祖先微生物可能是嗜热的。将结合生物工程、微生物学和分子生物学，确定已知的超嗜热物种以及新分离的菌株的生长和生存的环境限制。该项目与加州大学伯克利分校的 Douglas S. Clark 博士（获奖号 9816490）合作，强调分子对高压和高温的适应，其目标如下： 1. 确定超适温度和压力对现有超嗜热微生物存活和生长率的影响； 2. 利用加压连续发酵系统分离和培养新的超嗜热微生物菌株； 3. 检查生理适应和基因表达的遗传调控，以应对高温和高压的短暂挑战。 正在测试的假设是静水压可能会大大延长超嗜热菌生长和存活的温度上限。这些研究人员正在进行的研究已经证实，许多来自嗜热菌的酶在压力下表现出增强的热稳定性。此外，在压力作用下，詹氏甲烷球菌的生长速度加快了五倍，其产甲烷的最高温度上升了6摄氏度，而新鉴定的深海超嗜热菌Pyrococcus horikoshii的生长速度和ATP产量在压力下也有所提高。用于在压力和热循环下连续培养培养超级嗜热菌的新型设备将与分子生物学相结合，以探索极端嗜热菌的适应性反应。将检查基因表达、膜脂组成和形态。在接近致死的条件下诱导的基因将通过消减克隆和转录测定来鉴定。 将使用来自浅层陆地采样点的分离物来提供压力响应的控制数据，确定深海超嗜热菌的生存上限作为压力的函数。来自西北太平洋弧后区域喷口系统（堪察加半岛的冲绳海槽和乌宗火山口）的超嗜热菌的富集培养物将在超过已知菌株耐受的温度和压力下进行培养，从而使用存活而不是快速生长作为选择新分离株的标准。新菌株的系统发育位置和生理要求将被确定。

Hyperthermophilic microorganisms may be suitible models for ancestral, and possibly extraterrestrial, biosystems because their extraordinary adaptive capabilities allow them to colonize a wide range of subsurface volcanic areas on Earth. Deep sea hydrothermal vents and terrestrial hot springs support highly diverse ecosystems without access to solar energy, harboring autotrophic microorganisms and their dependant heterotrophic micro- and macro communities in geochemical conditions similar to those that may exist below the surface of Mars or Europa. Hyperthermophiles also dominate most of the deeper branches of the universal phylogenetic tree, suggesting that ancestral microorganisms may have been thermophilic. The environmental limits for growth and survival of known hyperthermophilic species, as well as newly isolated strains will be established, using a combination of bioengineering, microbiology and molecular biology. This collaborative project with Dr. Douglas S. Clark of the University of California, Berkeley (Award 9816490) emphasizes molecular adaptations to high pressure and high temperature, with the following objectives: 1. To determine the effects of supraoptimal temperatures and pressures on the survival and growth rates of existing hyperthermophilic microorganisms;2.To utilize pressurized continuous fermentation systems for isolation and culture of new hyperthermophilic microbial strains and; 3. To examine physiological adaptations and genetic regulation of gene expression in response to transient challenges by heat and high pressure. The hypothesis being tested is that hydrostatic pressure may greatly extend the upper temperature limits of growth and survival of hyperthermophiles. Ongoing studies by these researchers have established that many enzymes from thermophiles display enhanced thermostability under pressure. Further, the growth rate of Methanococcus jannaschii accelerated five-fold and its maximum temperature for methane production rose by 6 degrees C in response to pressure, and the growth rate and ATP production of a newly characterized abyssal hyperthermophile, Pyrococcus horikoshii, were elevated under pressure. Novel equipment for incubating hyperthermophiles in continuous culture, under pressure and with thermal cycling, will be combined with molecular biology to explore the adaptive responses of hyperthermophiles in extremis. Gene expression, membrane lipid composition and morphology will be examined. The genes that are induced under conditions approaching lethality, will be identified by subtractive cloning and transcriptional assays. Upper survival limits of the abyssal hyperthermophiles as a function of pressure, using isolates from a shallow terrestrial sampling site to provide control data for pressure responses, will be determined. Enrichment cultures of hyperthermophiles from the vent systems of the back-arc region of the Northwest Pacific (Okinawa Trough and Uzzon Caldera on the Kamchatka Peninsula) will be incubated at temperatures and pressures exceeding those tolerated by known strains, thus using survival rather than rapid growth as the criterion for selection of new isolates. The phylogenetic positions and physiological requirements of the new strains will be determined.