Contact-Dependent Expression of Met3 in Candida Biofilms

念珠菌生物膜中 Met3 的接触依赖性表达

基本信息

批准号：
7788871
负责人：
Luis A Murillo
金额：
$ 32.76万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
依托单位国家：
美国
项目类别：
财政年份：
2007
资助国家：
美国
起止时间：
2007-04-03 至 2012-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7788871
关键词：
Accounting Adherence Anabolism Animal Model Aspergillus Aspergillus Nuclease S1 Assimilations Biological Candida Candida albicans Catheters Cells Characteristics Code Communities Complex DNA DNA Footprint DNA Sequence Development Drug resistance Elderly Elements Epithelium Future Gene Expression Genes Genetic Transcription Genome Glutathione Growth Homologous Gene Immunocompromised Host Implant Indium Individual Infection Learning Maps Medical Device Methionine Microbial Biofilms Modeling Molecular Mucous Membrane Mutation Nail plate Neurospora Organism Oropharyngeal Pathogenesis Pathway interactions Pattern Phenotype Polystyrenes Polyvinyls Primer Extension Proteins Regulator Genes Regulatory Element Relative (related person)Research Personnel Ribonucleases Role Saccharomyces Signal Transduction Skin Stimulus Sulfate Adenylyltransferase Sulfur Surface System Transcription Initiation Site Up-Regulation Validation Virulence Work base candida biofilm cell behavior density in vitro Assay in vivo microbial community mutant neonate novel strategies novel therapeutics pathogen promoter research study trait transcription factor

项目摘要

DESCRIPTION (provided by applicant): The fungal pathogen, Candida albicans is able to form strongly adherent biofilms on inert surfaces such as those used for medical devices and on biological surfaces such as the skin, nails and mucous membranes. This attribute represents a key trait, which facilitates the persistence and dissemination of this opportunist, accounting in part for its increasing emergence as an important pathogen in immunocompromised individuals, neonates and the elderly. While it is widely known that microbial communities grow as biofilms and display distinct phenotypes [e.g. acquired drug resistance] as compared with their planktonic counterparts, the molecular basis through which these phenotypes are manifest remains unknown. Recent work shows that upon contact with an abiotic surface, C. albicans specifically and rapidly up regulates pathways for sulfur assimilation and glutathione biosynthesis and a number of other genes, many of which are of unknown function (103). These observations led us to ask how, at the molecular level, does a eukaryotic organism sense and respond to contact with a foreign (abiotic or biotic) surface and then transduce this information into altered patterns of gene expression and cell behavior. The immediate objective of this application is to identify key components, which regulate contact-dependent gene expression in C. albicans. To accomplish this we will use the MET3 gene, which encodes an ATP sulfurylase catalyzing the first step of the sulfur assimilation pathway. The MET3 gene is up-regulated over 20-fold within 50 minutes of cell contact to the abiotic surface and thus can be used to develop an experimental system to elucidate contact-dependent gene expression. In Specific Aim 1, we will define the c/s-acting regions, which regulate the differential expression of MET3 upon surface contact. Completion of this Aim will result in the identification of specific sequence elements in the DNA, which are required for contact- dependent expression of MET3 and will further produce a detailed map of the MET3 promoter in C. albicans, for which few have been characterized to this level. In Specific Aim 2, trans-acting elements required for contact-dependent gene expression will be identified. Completion of this Aim will result in the elucidation of the regulatory elements required for both planktonic and adherence-associated expression of MET3 and define transcription factors, which are uniquely responsive to the contact stimulus. Finally, in Specific Aim 3 we will assess the role of MET3 and the sulfur assimilation pathway in pathogenesis. The Candida sulfur assimilation pathway is qualitatively different from that of its mammalian host, providing promising targets for the development of new therapeutics. Understanding the underlying mechanism of contact sensing should provide novel approaches for interfering with cell adherence and thus controlling biofilm formation at the point of initial contact.

描述（由申请人提供）：真菌病原体白色念珠菌能够在惰性表面（例如用于医疗器械的表面）和生物表面（例如皮肤、指甲和粘膜）上形成牢固粘附的生物膜。这一属性代表了一个关键特征，有利于这种机会主义者的持续存在和传播，部分原因是它越来越多地成为免疫功能低下个体、新生儿和老年人的重要病原体。虽然众所周知，微生物群落以生物膜的形式生长并表现出不同的表型[例如，与浮游生物相比，这些表型表现出来的分子基础仍然未知。最近的研究表明，在接触非生物表面时，白色念珠菌特异性且快速地上调硫同化和谷胱甘肽生物合成的途径以及许多其他基因，其中许多基因的功能未知（103）。这些观察结果让我们想知道，在分子水平上，真核生物如何感知和响应与外来（非生物或生物）表面的接触，然后将这些信息转化为基因表达和细胞行为的改变模式。该应用的直接目标是确定调节白色念珠菌接触依赖性基因表达的关键成分。为了实现这一目标，我们将使用 MET3 基因，该基因编码催化硫同化途径第一步的 ATP 硫酸化酶。 MET3 基因在细胞接触非生物表面 50 分钟内上调超过 20 倍，因此可用于开发实验系统来阐明接触依赖性基因表达。在具体目标 1 中，我们将定义顺式/顺式作用区域，其调节表面接触时 MET3 的差异表达。该目标的完成将导致 DNA 中特定序列元件的鉴定，这些元件是 MET3 的接触依赖性表达所必需的，并将进一步产生白色念珠菌中 MET3 启动子的详细图谱，但很少有人对其进行表征。这个级别。在具体目标 2 中，将鉴定接触依赖性基因表达所需的反式作用元件。该目标的完成将阐明 MET3 的浮游和粘附相关表达所需的调控元件，并定义对接触刺激有独特反应的转录因子。最后，在具体目标 3 中，我们将评估 MET3 和硫同化途径在发病机制中的作用。念珠菌硫同化途径与其哺乳动物宿主有本质上的不同，为新疗法的开发提供了有希望的靶标。了解接触传感的基本机制应该提供干扰细胞粘附的新方法，从而控制初始接触点生物膜的形成。