Osmotic responsiveness of the master immune regulator PU.1

描述（由申请人提供）：免疫细胞如巨噬细胞和活化淋巴细胞对暴露于高渗环境（如淋巴组织和局部炎症）或活跃的细胞增殖引起的渗透应激高度敏感。细胞通过积累相容的渗透液来适应渗透胁迫，渗透液允许细胞有效地保留细胞内水分，保持细胞质体积，稀释细胞离子和其他关键溶质。积累的渗透物显著地提高了细胞内溶液的渗透压，或相应地降低了水的活性。因此，与水合作用的大变化耦合的大分子相互作用对细胞内的渗透变化很敏感。PU.1是ets家族转录因子，调控巨噬细胞、淋巴细胞等免疫细胞的发育和功能。我们最近的研究表明，pu -1的dna结合亲和力和序列选择性对其渗透环境非常敏感，但对其接近的结构同源物Ets-1却不敏感。另外，基因组研究表明，pu .1靶基因的调控，而不是Ets-1靶基因的调控，强烈依赖于功能性渗透胁迫反应。基于这些证据，我们假设渗透敏感性是PU.1的生物反应性和靶标特异性的机制。为了验证我们的假设，我们将确定PU.1的渗透敏感性的物理化学性质，并确定赋予其渗透敏感性的结构元素。我们将使用ETS -1作为渗透不敏感的“标准”来解释渗透敏感性如何特异性地纳入PU.1，以及渗透敏感性如何影响位点选择性（基因激活潜力的主要决定因素）。这些拟议的研究补充了20多年来在培养和体内对PU.1和其他ETS转录因子的细胞和功能研究。它们与PU.1和Ets-1如何协调调节其活性以允许T淋巴细胞在高渗胸腺环境中正确成熟具有特定的相关性。在这项研究完成后，我们期望将渗透敏感性作为PU.1对细胞渗透应激反应程序的反应机制。更广泛地说，这项研究引起了人们对水合作用作为人体细胞对生理渗透应激反应的生物物理基础的关注。这一知识对我们理解与病理性细胞内渗透应激相关的“各向异性失调”（如糖尿病和炎症）具有潜在的意义。

DESCRIPTION (provided by applicant): Immune cells such as macrophages and activated lymphocytes are highly sensitive to osmotic stress from exposure to hyperosmotic environments (such as lymphoid tissues and local inflammation) or active cell proliferation. Cells adapt to osmotic stress by accumulating compatible osmolytes, which allow cells to efficiently retain intracellular water, maintain cytoplasmic volume, and dilute cellular ions and other critical solutes. Accumulating osmolytes significantly raise the intracellular solution osmolality, or equivalently reduce water activity. Macromolecular interactions coupled to large changes in hydration are therefore sensitive to osmotic changes inside the cell. PU.1 is an ETS-family transcription factor that regulates the development and function of immune cells such as macrophages and lymphocytes. Our recent studies have demonstrated that DNA-binding affinity and sequence selectivity for PU.1 are profoundly sensitive to their osmotic environment, but not for its close structural homolog, Ets-1. Independently, genomic studies show that regulation of PU.1-target genes, but not Ets-1 target genes, depend strongly on a functional osmotic stress response. Based on this evidence, we hypothesize that osmotic sensitivity is a mechanism for biological responsiveness and target specificity for PU.1. To test our hypothesis, we will determine the physicochemical nature of osmotic sensitivity by PU.1 and identify the structural elements that confer its osmotic sensitivity. We will use Ets-1 as an osmotically insensitive "standard" to interpret how osmotic sensitivity is specifically incorporated into PU.1, and how osmotic sensitivity affects site selectivity (the primary determinant of gene-activating potential) These proposed studies complement more than two decades of cellular and functional studies of PU.1 and other ETS transcription factors in culture and in vivo. They have specific relevance to how PU.1 and Ets-1 coordinately modulate their activities to permit correct maturation of T lymphocytes in the hyperosmotic thymus environment. Upon completion of this research, we expect to have established osmotic sensitivity as a mechanism for responsiveness by PU.1 to the cellular osmotic stress response program. More broadly, this research brings attention to hydration as a biophysical basis for responsiveness to physiologic osmotic stress in human cells. This knowledge has potential implication for our understanding of "anisotonic disorders" (such as diabetes mellitus and inflammation) associated with pathological intracellular osmotic stress.