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

白色念珠菌和相关念珠菌属。每年造成约40万例侵入性感染， 50%的死亡率C.白色念珠菌和其他真菌，是能够减少他们的能力， 被他们的主人发现。细胞壁碳水化合物β（1，3）-葡聚糖是机体免疫反应的重要表位 人类和其他哺乳动物的系统用于通过受体识别和响应真菌感染 如Dectin-1和补体受体3（CR 3）。真菌如C.白色念珠菌减少了免疫检测 通过在甘露糖基化糖蛋白（甘露聚糖）外层下掩蔽β（1，3）-葡聚糖来保护细胞。的 C.毒力白念珠菌在β（1，3）-葡聚糖更多暴露（未掩蔽）的条件下受损。为 例如，棘白菌素类抗真菌药物，如卡泊芬净，抑制β（1，3）-葡聚糖合酶，并导致细胞溶解， 体外，而且还诱导β（1，3）-葡聚糖的暴露，即使在亚致死浓度下。此外，一些 表现出增加的β（1，3）-葡聚糖暴露的突变体具有降低的毒性。然而一个主要 研究的挑战是了解卡泊芬净给药期间β（1，3）-葡聚糖暴露对毒力的影响 治疗很难区分药物的杀菌作用和β（1，3）-葡聚糖的影响 exposure.与此密切相关的一个挑战是，卡泊芬净引起β（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菌作用与暴露分开， 这让我们能够应对这两个挑战。这可以通过激活卡泊芬净应答来实现。 使用遗传方法而不是药物的信号通路，我们已经发现， 这些途径导致暴露。Cek 1 MAP激酶（MAPK）通路被卡泊芬净激活 治疗，我们已经发现，这种级联反应的遗传激活导致暴露， 即使在没有卡泊芬净的情况下也会过度活化。然而，与药物不同的是，这种途径的激活 不影响生存能力因此，我们可以通过使用这一途径来解剖 这是一种机制，通过这种机制，可以进行解密。此外，我们可以通过使用Cek 1来应对第一个挑战。 途径作为探测免疫系统在小鼠全身感染期间如何响应暴露的工具 因为与卡泊芬净不同，它不是杀菌剂。我们将在三个具体目标中应对这些挑战。目标1 我们将阐明Cek 1级联调节β（1，3）-葡聚糖暴露的机制。有两 Cek 1下游的主要转录因子，我们将确定该途径如何选择一个特定的 一个（Cph 1）使用遗传，上位性和细胞生物学技术的组合，将确定Cek 1- Cph 1被激活以引起暴露。在目标2中，我们将确定Cek 1-Cph 1的转录靶点如何 改变细胞壁导致暴露。在目标3中，我们将阐明β（1，3）-葡聚糖的暴露如何导致 降低小鼠的毒性。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CR 3参与降低未掩蔽的C.白色念珠菌

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.