Regulation of ß(1,3)-glucan exposure in Candida albicans

白色念珠菌及其相关种。每年造成约40万例侵入性感染，其中 死亡率约为50%。白色念珠菌和其他真菌的一个重要毒力特征是能够削弱它们的 被它们的宿主检测到。胞壁碳水化合物(1，3)-葡聚糖是免疫的重要表位 人类和其他哺乳动物的系统通过受体识别和响应真菌感染 如Dectin-1和补体受体3(CR3)。像白色念珠菌这样的真菌会减少它们对免疫系统的检测 通过将(1，3)-葡聚糖掩蔽在甘露糖苷糖蛋白(甘露聚糖)的外层。这个 白色念珠菌的毒力在？(1，3)-葡聚糖暴露更多(未被掩盖)的条件下受到损害。为 例如，棘球菌素类抗真菌药物，如卡泊芬净，可以抑制？(1，3)-葡聚糖合成酶，并导致细胞裂解。 即使在亚致死浓度下，也会导致(1，3)-葡聚糖的暴露。此外，还有一些 表现出更多的(1，3)-葡聚糖暴露的突变体降低了毒力。然而，一个主要的 研究挑战在于了解在卡泊芬净生产过程中，(1，3)-葡聚糖暴露对毒力的影响 治疗。很难区分这种药物的杀灭作用和(1，3)-葡聚糖的影响 曝光。与此密切相关的一个挑战是，卡泊芬净导致？(1，3)-葡聚糖的机制 暴露是未知的。我们已经发现，我们可以将卡泊芬净的杀戮作用与揭开面具脱钩， 这使我们能够应对这两个挑战。这可以通过激活卡泊芬净响应性来实现 使用遗传方法而不是药物的信号通路，我们发现至少有一个 这些途径导致了揭开面纱。卡泊芬净激活CEK1 MAPK通路 治疗，我们已经发现，这种级联反应的基因激活导致在以下情况下揭开面纱 即使在没有卡泊芬净的情况下也是如此。然而，与药物不同的是，激活这一途径会 而不是牺牲生存能力。因此，我们可以通过使用这条途径来解剖 揭开屏蔽发生的机制。此外，我们可以通过使用CEK1来迎接第一个挑战 途径作为工具探索免疫系统在小鼠全身感染过程中对揭幕的反应 因为，与卡泊芬净不同的是，它不是致命的。我们将通过三个具体目标来应对这些挑战。在目标1中 我们将阐明CEK1级联基因调控？(1，3)-葡聚糖暴露的机制。有两个 CEK1下游的主要转录因子，我们将确定该途径如何选择特定的 一种(Cph1)结合了遗传学、上位性和细胞生物学技术，将确定CEK1是如何- Cph1被激活以取消屏蔽。在目标2中，我们将确定CEK1-Cph1的转录靶标如何 改变细胞壁以揭开面罩。在目标3中，我们将阐明(1，3)-葡聚糖的暴露如何导致 降低小鼠的致病力。我们将使用转基因小鼠来定义中性粒细胞、巨噬细胞、Dectin-1 和/或CR3参与降低未遮盖白念珠菌的毒力。

Candida albicans and related Candida spp. are responsible for ~400,000 invasive infections/year, which have an ~50% mortality rate. A crucial virulence trait of C. albicans, and other fungi, is the ability to diminish their detection by their hosts. The cell wall carbohydrate ß(1,3)-glucan is an important epitope that the immune systems of humans and other mammals use to recognize and respond to fungal infections through receptors like Dectin-1 and complement receptor 3 (CR3). Fungi like C. albicans diminish their detection from immune cells through masking ß(1,3)-glucan under an outer layer of mannosylated glycoproteins (mannan). The virulence of C. albicans is compromised in conditions where ß(1,3)-glucan is more exposed (unmasked). For example, echinocandin antifungal drugs, like caspofungin, inhibit ß(1,3)-glucan synthase and cause cell lysis in vitro, but also induce exposure of ß(1,3)-glucan, even at sublethal concentrations. In addition, a number of mutants that exhibit increased exposure of ß(1,3)-glucan have decreased virulence. However, a major research challenge is to understand the impact of ß(1,3)-glucan exposure on virulence during caspofungin treatment. It has been difficult to differentiate between cidal effects of the drug and the impact of ß(1,3)-glucan exposure. A challenge closely related to this is that the mechanism by which caspofungin causes ß(1,3)-glucan exposure is unknown. We have found that we can decouple caspofungin's cidal effects from unmasking, which allows us to address both of these challenges. This can be done by activating caspofungin-responsive signaling pathways using a genetic approach rather than the drug, and we have discovered that at least one of these pathways causes unmasking. The Cek1 MAP kinase (MAPK) pathway is activated by caspofungin treatment, and we have discovered that genetic activation of this cascade causes unmasking when hyperactivated, even in the absence of caspofungin. However, unlike the drug, activation of this pathway does not compromise viability. Thus, we can meet the second challenge by using this pathway to dissect the mechanism through which unmasking occurs. Moreover, we can meet the first challenge by using the Cek1 pathway as tool to probe how the immune system responds to unmasking during mouse systemic infections because, unlike caspofungin, it is not cidal. We will address these challenges in three specific aims. In Aim 1 we will elucidate the mechanisms by which the Cek1 cascade regulates ß(1,3)-glucan exposure. There are two main transcription factors downstream of Cek1 and we will determine how the pathway chooses a particular one (Cph1) using a combination of genetic, epistasis, and cell biology techniques that will identify how Cek1- Cph1 is activated to cause unmasking. In Aim 2 we will determine how transcriptional targets of Cek1-Cph1 alter the cell wall to cause unmasking. In Aim 3, we will elucidate how exposure of ß(1,3)-glucan causes decreased virulence in mice. We will use transgenic mice to define how neutrophils, macrophages, Dectin-1 and/or CR3 participate to reduce the virulence of unmasked C. albicans.