LEXEN: Microbial Adapation to High Temperature and Pressure

超嗜热微生物可能是祖先，也可能是地外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广泛的地下火山区域定居。深海热液喷口和陆地温泉在没有太阳能的情况下支持高度多样化的生态系统，在类似于火星或木卫二表面下可能存在的地球化学条件下，庇护自养微生物及其依赖的异养微观和宏观群落。超嗜热微生物在普遍系统发育树的深层分支中也占主导地位，这表明祖先微生物可能是嗜热的。利用生物工程、微生物学和分子生物学的结合，将建立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极限。该项目与巴尔的摩马里兰大学的Frank T. Robb博士（奖项MCB-9809352）合作，强调分子对高压和高温的适应，目标如下：确定超优温度和压力对现有超嗜热微生物的生存和生长速度的影响；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. 研究高温和高压瞬时挑战下的生理适应和基因表达的遗传调控。正在测试的假设是静水压力可以大大延长超嗜热微生物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，jannaschii甲烷球菌的生长速度在压力作用下加快了5倍，产甲烷的最高温度提高了6℃；新发现的深海超嗜热细菌——博氏焦球菌（Pyrococcus horikoshii）的生长速度和ATP产量在压力作用下提高。在压力和热循环下连续培养超嗜热菌的新型设备将与分子生物学相结合，探索极端条件下超嗜热菌的适应反应。将检测基因表达、膜脂组成和形态。在接近致死的条件下诱导的基因，将通过减法克隆和转录测定来鉴定。将利用浅层陆地取样点的分离物为压力反应提供控制数据，确定深海超嗜热生物的生存上限作为压力的函数。来自西北太平洋弧后地区（冲绳海槽和堪察加半岛的Uzzon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培养，因此使用生存而不是快速生长作为选择新分离株的标准。将确定新菌株的系统发育位置和生理需求。＊＊＊

9816490 Clark Hyperthermophilic microorganisms may be suitable 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. Frank T. Robb of the University of Maryland, Baltimore (Award MCB-9809352) 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. ***