REGULATION OF UTERINE SMOOTH MUSCLE EXCITABILITY

描述(申请人提供)：从子宫静止到收缩的转变对新生儿和母亲的健康至关重要，但这一事件的发生时间往往不正确；在美国，12%的婴儿早产，20%的婴儿在人工引产后分娩。因此，了解子宫肌层平滑肌细胞(MSMC)电活动的调节及其对收缩的影响，对于理解正常分娩和治疗功能障碍分娩都是至关重要的。维持子宫静止需要在促进宫缩的兴奋性去极化刺激和抑制子宫收缩的抑制性复极电流之间保持复杂的平衡。作为MSMC的主要通道之一，大电导钙激活的K+通道(KCa1.1)对兴奋性信号产生强烈的复极化电流，从而抑制MSMC的收缩，从而有助于静息。尽管有强有力的证据支持KCa1.1通道调节子宫兴奋性的概念，但其在怀孕期间参与生理调节的基本机制仍然很大程度上尚不清楚。我研究的长期目标是确定在怀孕期间从静止到收缩转变的离子机制。这项建议的目的是确定KCa1.1通道在怀孕期间被调节以控制子宫肌层兴奋性的机制。我们的中心假设是，这一通道既受固有属性的动态调节，也受其与调节蛋白的关联的调节。为了支持这一观点，我们在人骨髓基质细胞中的初步研究表明，KCa1.1受到选择性翻译起始的调控，导致KCa1.1亚型的细胞外N末端发生变化。这些N末端变异体的调节不同于附件1亚基。我们还产生了蛋白质组学数据，表明其他新的调节剂，包括最近描述的-亚基家族和蛋白酶抑制剂2巨球蛋白(A2M)，选择性地与子宫肌层KCa1.1相关，并可能改变通道活性。该项目的目标是：1)确定与KCa1.1相互作用的新蛋白在非劳动和劳动人类子宫肌层中的时空相互作用；2)确定KCa1.1的内在属性，改变其与相互作用伙伴的联系；以及3)确定KCa1.1在子宫肌层细胞系和非劳动和劳动人类肌层中的功能调节机制。这项研究将建立调节KCa1.1活性的分子通路，为调节子宫兴奋性的治疗提供生物学基础。

DESCRIPTION (provided by applicant): The transition from uterine quiescence to contraction is vital to the health of a newborn and mother, but timing of this event often fails to occur properly; in the U.S., 12% of babies are born preterm, and 20% are delivered following artificial induction of labor. Thus, understanding the regulation of myometrial smooth muscle cell (MSMC) electrical activity and its effect on contraction is essential for both comprehending normal labor and treating dysfunctional labor. Maintenance of uterine quiescence requires an intricate balance between excitatory depolarizing stimuli that promote contractions and inhibitory repolarizing currents that suppress uterine contraction. One predominant channel in MSMCs, the large conductance Ca2+-activated K+ channel (KCa1.1) contributes to quiescence by eliciting a potent repolarizing current in response to excitatory signals, thereby dampening MSMC contraction. In spite of strong evidence supporting the notion that the KCa1.1 channel modulates uterine excitability, the basic mechanisms involved in its physiological regulation during pregnancy remain largely uncharacterized. The long-term goal of my research is to identify the ionic mechanisms that regulate the transition from quiescence to contraction during pregnancy. The objective of this proposal is to define the mechanisms by which the KCa1.1 channel is modulated during pregnancy to control myometrial excitability. Our central hypothesis is that this channel is dynamically modulated by both intrinsic properties and by its association with modulatory proteins. In support of this idea, our preliminary studies in human MSMCs indicate that KCa1.1 is regulated by alternative translation initiation, resulting in KCa1.1 isoforms that vary in their extracellular N- termini. These N-terminal variants differ in their regulation by accessory 1-subunits. We also have generated proteomics data demonstrating that other novel modulators, including the recently described family of - subunits and the protease inhibitor 2macroglobulin (A2M), selectively associate with myometrial KCa1.1 and potentially modify channel activity. The goals of this project are to: 1) define the spatial and temporal interactions between novel proteins that interact with KCa1.1 in non-laboring and laboring human myometrium, 2) identify intrinsic properties of KCa1.1 that alter its association with interacting partners; and 3) determine the mechanism of functional regulation of KCa1.1 in both myometrial cell lines and non-laboring and laboring human myometrium. The research proposed here will establish the molecular pathways that regulate KCa1.1 activity, providing a biological basis for therapies designed to modulate uterine excitability.