Cell-fate determinants of yeast pseudohyphal growth

描述（由申请人提供）：缺氮导致酵母S的二倍体细胞。 酿酒酵母脱离营养生长并形成假菌丝。 这些是杆状细胞的细丝，从集落的外围辐射出来，并扩散到琼脂培养基中。 当酵母形式（YF）细胞分裂并产生与其母亲具有不同发育命运的PH女儿时，就会出现这种转变。 这种不对称转换与人类多能干细胞产生不同谱系过程中发生的转换基本相同。 在酵母中，就像在哺乳动物中一样，命运的改变需要协调由多种转录因子控制的程序。 一些是响应于细胞外的刺激和其他的细胞固有的机制，不对称分离的关键命运决定因素。 S.酿酒酵母Ash 1是一种GATA家族转录因子，其差异地分离至子代，并且对于PH转换是必需的。 虽然负责不对称本地化的阿什1的机制已被阐明，很少有人知道的女儿特定的程序，它介导建立PH的命运。 这个程序很难用标准方法研究，因为PH菌落是细胞类型的异质混合物。 申请人提出使用绿色荧光蛋白（GFP）标记来开发策略，其允许分析个体活细胞中的启动子功能以及蛋白质和mRNA动力学。 目前，细胞表面粘附蛋白Flo 11是PH生长所需的唯一已知基因产物，其转录依赖于Ash 1。 目的（1）研究Flo 11的转录调控。 研究将定义Ash 1依赖于该启动子的机制，并揭示Ash 1如何与Flo 11的信号依赖性转录调控因子合作。 目的（2）探讨Flo 11在PH生长中的子体特异性定位和功能。 目的（3）提出一种新的策略，以产生子体特异性互补脱氧核糖核酸（cDNA），用于阵列分析，以确定子体特异性基因受阿什1控制。 本申请人为Ash 1转录控制与信号依赖性转录因子的转录控制整合的机制建立的原理将广泛适用于GATA家族。 特别是，Ash 1是保守的，并且是致病真菌毒力所必需的YF-PH转换所必需的。 因此，Errede博士定义的分子机制将直接适用于那些潜在的真菌致病性。 此外，该项目有可能揭示真菌蛋白质，这些蛋白质将成为药物干预的绝佳候选目标。

DESCRIPTION (provided by applicant): Nitrogen deprivation causes diploid cells of the yeast S. cerevisiae to depart from vegetative growth and form pseudohyphae. These are filaments of rod-shaped cells that radiate from the periphery of a colony and spread over and into the agar medium. The transition arises when a yeast form (YF) cell divides and gives rise to a PH daughter that has a different developmental fate from its mother. This asymmetric switch is fundamentally the same as those that occur during the production of distinct lineages from pluripotent stem cells in humans. In yeast as in mammals, the fate change requires co-ordination of programs that are controlled by multiple transcription factors. Some are responsive to extracellular stimuli and others on cell inherent mechanisms for asymmetric segregation of the critical fate determining factors. S. cerevisiae Ash 1 is a GATA-family transcription factor that is differentially segregated to daughters and is essential for the PH transition. While the mechanism responsible for asymmetric localization of Ash1 has been elucidated, very little is known about the daughter-specific program that it mediates to establish the PH fate. This program has been difficult to study by standard methods because PH colonies are a heterogeneous mixture of cell-types. The applicant proposes to develop strategies using green fluorescent protein (GFP) labels that allow analyses of promoter function and protein and mRNA dynamics in individual living cells. Currently, the cell-surface adhesion protein, Flo11, is the only known gene product required for PH growth whose transcription is dependent on Ash1. Aim (1) focuses on Flo11 transcriptional regulation. Studies will define the Ash 1-dependent mechanism at this promoter and reveal how Ash1 collaborates with signal dependent transcriptional regulators of Flo11. Aim (2) addresses daughter-specific localization and functions of Flo11 in PH growth. Aim (3) proposes a novel strategy to generate daughter-specific complementary deoxyribonucleic acids (cDNAs) for use in array analyses to identify daughter-specific genes subject to Ash1 control. The principles the applicants establish for the mechanism by which Ash1 transcriptional control is integrated with that of signal dependent transcription factors will be broadly applicable to the GATA-family. In particular, Ash1 is conserved and required for the YF-PH transition that is essential for virulence of pathogenic fungi. Thus, the molecular mechanisms Dr. Errede defines will be directly applicable to those underlying fungal pathogenicity. Additionally, the project has the potential to reveal fungal proteins that would be excellent candidates as targets for drug intervention.