MOLECULAR BASIS OF VIRULENCE FACTORS OF ORAL FUNGAL SP.

口腔真菌毒力因子的分子基础。

• 批准号: 6902145
• 负责人: MARY ANN Y JABRA-RIZK
• 金额: $ 0.09万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-09-01 至 2007-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6902145
• 关键词: Candida; Candida albicans; Fusobacterium nucleatum; candidiasis; cell adhesion; clinical research; fibronectins; fungal genetics; fungal proteins; gene expression; hydropathy; macrophage; mannans; microarray technology; molecular cloning; nucleic acid amplification techniques; nucleic acid hybridization; phagocytosis; polymerase chain reaction; postdoctoral investigator; respiratory epithelium; temperature sensitive mutant; virulence

DESCRIPTION (provided by applicant): This is a Revision of a K22 Scholar Development and Faculty Transition Award application, submitted on behalf of Mary Ann Jabra-Rizk who is currently a Post-Doctoral Fellow in the Department of Diagnostic Sciences and Pathology at the U of MD Dental School. A 2 year Scholar Development phase and three years of Faculty Transition win be centered at U of MD involving co-mentors and consultants at Johns Hopkins, the U of VA and the U of Wurzburg and will establish Dr. Jabra-Rizk as an independent expert in oral fungal diseases. An Advisory Committee at the U of MD will evaluate her progress and aid in the Faculty Transition. The hypothesis to be tested during the Scholar Development is that C. dubliniensis is a species that exhibits constant cell surface hydrophobicity, possessing a homologue to the C. albicans CSHl gene which encodes a hydrophobic cell wall protein involved in cell adherence but differs in the expression of the MNN mannosylation family of genes and other putative virulence genes. Two aims are planned: 1. (a) Determine the presence of the C. albicans CSHl and CaMNN9 genes or homologues of the genes in C. dubliniensis using PCR primers designed based on the C albicans CSIII and CaMNN9 gene sequences. These will be subsequently amplified and cloned using C. dubliniensis CDS6 genomic DNA and probe hybridization. A C. dubliniensis knockout of the CSH 1 homologue gene will be generated using the latest technique (ura3 amxotrophic C. dubliniensis mutants in the URA-blaster) in gene disruption technique (b) Determine the effect of the disruption of the CdCShl gene on adherence by assessing the C. dubliniensis knockout clone for cell surface hydrophobicity7 adhesion to fibronectin, adherence to Fusobacterium nucleatum and pooled human buccal epithelial cells, in comparison to the wild type (c) Determine differences in the ability of macrophages to phagocytize 37C-grown C. albicans, 37C grown CdCSH1 mutant and wild type C. dubliniensis and the C. albicans mnn9 mutant in order to determine the effect of structural changes in the side chains of cell wall mannan on the ability of C. dubliniensis and C. albicans to evade host cell phagocytosis. 2. Based on cDNA microarray sequences of genes in the C. albicans genome, we will determine levels of differential expression of the CSHl and CaMNN9 genes between hydrophilic (37C-grown C. albicans) and hydrophobic (250C-grown C. albicans, 25 and 37C-grown C. dubliniensis and C. albicans mutants, A9V10 and Camnn9) yeast cells, as well as the C. dubliniensis CdCSHl mutant generated in Aim 1. Dr. Jabra-Rizic will subsequently utilize the information and mutants generated to investigate whole genome differences between C. dubliniensis and C. albicans in the Faculty Transition phase using DNA microarrays. She will also analyze differences in the levels of expression of hsp90 gene and other glycosylation and heat shock proteins genes in C. dubliniensis. In addition, a hsp90 knock out mutant of C. albicans will be generated and assessed for thermotolerance and phagcytosis by macrophages. Long range plans include using the mutants generated to study Candida mannoprotein-specific host immunomodulation, such as stimulation of cytokines and chemokines.

描述（由申请人提供）： 这是代表玛丽·安·贾布拉·里兹克（Mary Ann Jabra-Rizk）提交的K22学者发展和教师过渡奖申请的修订，他目前是MD牙科学校U诊断科学和病理学系的博士后研究员。一个为期两年的学者发展阶段和三年的教师过渡胜利以MD为中心，涉及约翰·霍普金斯（Johns Hopkins），弗吉尼亚州U和尤尔茨堡（U）的联盟顾问和顾问，并将建立Jabra-Rizk博士作为口腔真菌疾病的独立专家。 MD U的咨询委员会将评估她的进步并帮助教师过渡。在学者发展期间要检验的假设是，都柏林梭菌是一种表现出恒定的细胞表面疏水性的物种，它具有与白色念珠菌CSHL基因的同源物，该物种编码与细胞粘附相关的疏水细胞壁蛋白，但在MNN mannosylation家族的表达中却有所不同。计划了两个目的：1。（a）使用基于基于C albicans cSIII和CAMNN9基因序列的PCR引物在dubliniensis中确定白色念珠菌CSHL和CAMNN9基因或基因的同源物。 这些将随后使用都柏林C. cds6基因组DNA和探针杂交对其进行扩增和克隆。使用最新技术（ura3氨基营养性弧菌C. c. dubliniensis突变体）中的CSH 1同源基因敲除基因破坏技术（b）确定CDCSHL基因对粘附的破坏对粘附的c. dublineysis c. c. dublinesis c.与野生型（c）相比，纤连蛋白，依从性核细菌和人类颊上皮细胞合并的人（C）确定了巨噬细胞吞噬37c成长的C. albicans的能力的差异，37c c.Cdcsh1突变体和野生型C. c. c. c. c. buttys cotty in Sustrant in and and cotty insunty in n and and cotty in n and nsunn typers in nsunn and nsunn cotty in n and cottyn99cottyn99。细胞壁曼南的链条对二柏林梭菌和白色念珠菌逃避宿主细胞吞噬作用的能力。 2。基于白色念珠菌基因组中基因的cDNA微阵列序列，我们将确定亲水性（37c生的白色念珠菌）和疏水性的CSHL和CAMNN9基因的差异表达水平AIM 1中产生的都柏林CDCSHL突变体。Jabra-Rizic博士随后将利用所产生的信息和突变体在教师过渡期使用DNA微阵列研究了都柏林梭菌和白色念珠菌之间的整个基因组差异。她还将分析都柏林梭菌中Hsp90基因表达水平以及其他糖基化和热休克蛋白基因的差异。此外，将产生HSP90敲除白色念珠菌的突变体，并评估巨噬细胞的耐热性和吞噬作用。远距离计划包括使用生成的突变体研究念珠菌特异性宿主免疫调节，例如刺激细胞因子和趋化因子。