Molecular Characterization Of Fungal Biofilm Formation

描述：本R21的目的是表征真菌生物膜形成的分子事件，从最初的白色念珠菌附着到非生物表面，通过成熟和连续的细胞网络的发展，牢固地附着在该表面。我们的假设是，在白色念珠菌生物膜发育过程中的转录分析将导致在这一过程中表达的关键阶段、途径和功能的鉴定。细菌和真菌的生物膜形成过程主要是通过描述事件来评估的，在微观和宏观水平上，导致在惰性表面上建立生物膜。在真菌生物膜的情况下，人们对与生物膜发育相关的潜在分子过程知之甚少。除了有限的细胞外基质的生化表征，生物膜中细胞的药物敏感性变化以及主要与菌丝形态发生调节相关的各种突变的影响外，与浮游细胞相比，贴壁细胞中可能表现出的分子变化知之甚少。随着贴壁细胞建立高度结构化的细胞网络，分子变化可能发生在贴壁细胞中，但对分子变化知之甚少。在这个高通量基因组分析的时代，我们有独特的能力来解决这个新出现的问题，通过使用的方法，允许在生物膜发育过程中基因表达的变化进行全面分析。在这项研究中，我们将使用DNA微阵列，这是我们与斯坦福基因组技术中心（STGC）的Ronald Davis博士及其同事共同开发的；这些阵列包括白色念珠菌的整个基因组。RNA样品将在高度控制的时间过程中从粘附细胞和浮游细胞中分离出来，该时间过程将跨越在应用程序中描述的条件下形成生物膜所需的24小时。从这样一项研究中产生的大量数据集将使用各种算法进行仔细分析，这些算法将允许定义特定基因和基因簇/途径的变化，这些变化可能在生物膜发育过程中表达。通过比较贴壁细胞和非贴壁细胞中的基因表达，我们将了解附着于惰性表面是否以及在多大程度上影响基因表达。

DESCRIPTION: The purpose of this R21 is to characterize the molecular events in fungal biofilm formation from the initial attachment of C. albicans to an abiotic surface through the development of a mature and continuous cellular network that is firmly adherent to that surface. It is our hypothesis that transcription profiling during the development of a C. albicans biofilm will result in the identification of key stages, pathways and functions that are expressed during this process. The process of biofilm formation in both bacteria and fungi has primarily been assessed through the description of events, at the microscopic and macroscopic level, which result in the establishment of biofilms on inert surfaces. In the case of fungal biofilms, little is known about the underlying molecular process associated with biofilm development. With the exception of limited biochemical characterization of the extracellular matrix, the changes in drug susceptibility in cells in biofilms and the effects of various mutations associated primarily with the regulation of hyphal morphogenesis, little is known about the molecular changes which may be manifest in adherent cells as compared with planktonic cells. Less is known about the molecular changes, which may occur in adherent cells as these proceed to establish highly-structured cellular networks. In this era of high throughput genome profiling, we have the unique ability to address this emerging problem by using methods that allow comprehensive analysis of the changes in gene expression during the course of biofilm development. For this study, we will use DNA microarrays, which we have developed with Dr. Ronald Davis and colleagues at the Stanford Genome Technology Center (STGC); these arrays are inclusive of the entire genome of C. albicans. RNA samples will be isolated from adherent and planktonic cells in a highly-controlled time course which will span the 24h period required for biofilm formation under the conditions described in the application. The large data set that will emerge from such a study will be carefully analyzed using a variety of algorithms that will allow definition of changes in specific genes and in gene clusters/pathways, which may be expressed during the course of biofilm development. By comparing gene expression in adherent and non-adherent cells, we will learn whether and to what degree, attachment to an inert surface can affect gene expression.