Transcriptional regulation of A. fumigatus virulence

描述(由申请人提供)：由烟曲霉引起的侵袭性感染正在增加，尽管有新的治疗方法，但死亡率仍高于50%。在侵袭性曲霉病的发病过程中，机体与多种宿主细胞相互作用，包括肺上皮细胞和内皮细胞。吸入后，烟曲霉分生孢子附着并被排列在肺泡内的肺上皮细胞内化。在敏感宿主中，这些分生孢子萌发形成菌丝，菌丝离开肺上皮细胞，侵入更深的组织，最明显的是肺血管。在这些血管中，菌丝以两种独特的方式与血管内皮细胞相互作用。首先，来自感染区的菌丝侵入血管的腔面，然后穿透内皮细胞进入血管腔。接下来，菌丝碎片被血流携带到远端部位，在那里它们附着并穿透内皮细胞的管腔表面，然后侵入深层器官。到目前为止，人们对烟曲霉毒力特征的遗传控制知之甚少，这些特征影响了有机体与这些寄主细胞的相互作用。在其他病原真菌中，控制真菌形态发生和生长的保守信号转导通路也是毒力性状的关键调节因子。来自我们小组和其他人的数据表明，这种模式适用于烟熏鱼。因此，我们推测，烟曲霉菌与肺上皮细胞和血管内皮细胞的相互作用既是发育调节的，也是侵袭性曲霉病发病的关键。为此，我们已经确定了一些预测控制真菌发育的A fumigatus转录因子。这些候选基因缺失的突变体已经构建完成。我们还开发了动物和体外模型系统，以生理相关的方式研究这些突变体与上皮细胞和内皮细胞的相互作用。我们假设这些突变体将有异常的发育，与宿主细胞的异常相互作用，并减弱毒力。我们将通过1)确定候选转录因子在烟曲霉菌的发育和形态发生中的作用；2)研究候选转录因子缺失的突变株在体外与肺上皮细胞和血管内皮细胞的相互作用；3)利用侵袭性曲霉病的小鼠模型表征这些候选转录因子的子集调节毒力和宿主反应的能力；以及4)确定其表达受我们发现的控制宿主细胞相互作用和毒力的候选转录因子调控的下游基因。这些研究将确定以前未知的烟曲霉转录因子在控制发育、宿主细胞相互作用和毒力方面的作用。他们还将识别新的基因，这些基因指定了明确定义的、生物学上相关的烟曲霉毒力特征。总的来说，这些结果将为侵袭性曲霉病的新治疗策略的未来发展奠定基础。

DESCRIPTION (provided by applicant): Invasive infections due to Aspergillus fumigatus are increasing, and despite new therapies are associated with greater than 50% mortality. During the pathogenesis of invasive aspergillosis, the organism interacts with several types of host cells, including pulmonary epithelial cells and endothelial cells. After being inhaled, A. fumigatus conidia adhere to and are internalized by the pulmonary epithelial cells that line the alveoli. In susceptible hosts, these conidia then germinate to form hyphae, which exit the pulmonary epithelial cells and invade the deeper tissues, most notably pulmonary blood vessels. Within these blood vessels, hyphae interact with vascular endothelial cells in two unique ways. First, hyphae from an infectious focus invade the abluminal surface of the blood vessel and then penetrate endothelial cells to gain access to the blood vessel lumen. Next, hyphal fragments are borne by the bloodstream to distal sites where they adhere to and penetrate the luminal surface of the endothelial cells and then invade the deep organs. To date, little is known about the genetic control of A. fumigatus virulence traits that influence the interactions of the organism with these host cells. In other pathogenic fungi, the conserved signal transduction pathways that control fungal morphogenesis and growth are also key regulators of virulence traits. Data from our group and others suggest that this paradigm holds true for A fumigatus. Thus, we hypothesize that the interactions of A fumigatus with pulmonary epithelial cells and the vascular endothelium are both developmental^ regulated and critical to the pathogenesis of invasive aspergillosis. Towards this end, we have identified a number of A fumigatus transcription factors predicted to govern fungal development. Mutants that are deficient in these candidate genes have been constructed. We have also developed animal and in vitro model systems to investigate the interaction of these mutants with epithelial and endothelial cells in a physiologically relevant manner. We hypothesize that these mutants will have abnormal development, aberrant interactions with host cells, and attenuated virulence. We will test this hypothesis by 1) determining the role of candidate transcription factors in the development and morpho- genesis of A fumigatus; 2) investigating the interactions of mutants deficient in candidate transcription factors with pulmonary epithelial cells and vascular endothelial cells in vitro; 3) characterizing the ability of a subset of these candidate transcription factors to modulate virulence and host response using murine models of invasive aspergillosis; and 4) identifying the downstream genes whose expression is regulated by candidate transcription factors that we find govern host cell interactions and virulence. These investigations will establish the roles of previously unexplored A. fumigatus transcription factors in governing development, host cell interactions, and virulence. They will also identify new genes that specify clearly defined, biologically relevant A fumigatus virulence traits. Collectively, these results will form the foundation for the future development of new therapeutic strategies for invasive aspergillosis.