CHOLINE TRANSPORTERS SPECIFICALLY TARGET NON-NEURONAL ACETYLCHOLINE SYNTHESIS

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 非神经元性乙酰胆碱(Ach)的合成和分泌可能在肺癌和哮喘等肺部疾病中发挥作用。为了确定非神经元信号在肺中的特征和靶点，有必要确定神经元和非神经元胆碱能信号之间的不同步骤。胆碱摄取是ACh合成的限速步骤，并可能提供这一靶点。在神经元中，钠依赖的高亲和力胆碱转运体(CHT1)绝对需要将胆碱转运到神经元中以合成ACh。我们的初步数据显示，一些肺细胞不需要CHT1来合成ACh，相反，一类新发现的胆碱转运体--胆碱转运蛋白样蛋白(CTL1-5)可以介导ACh的合成。因此，通过靶向正确的CTL，肺中非神经元ACh的合成可能是潜在的靶向，而不影响神经元ACh的合成。为了验证这一点，我们提出了以下具体目标：(1)鉴定哪些胆碱转运蛋白样蛋白与肺非神经细胞的胆碱摄取有关。(2)肺内CTL与非神经元性ACh合成的偶联。(3)确定下调与ACh合成有关的CTL是否能抑制肺细胞的增殖。这些目标将用特定的siRNAs敲除H82小细胞肺癌细胞中的CTL作为肺神经内分泌细胞的模型，并将H520鳞状细胞肺癌细胞作为呼吸道上皮细胞的模型。因此，这些研究将允许将非神经性ACh合成作为开发肺部疾病新疗法的第一步。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Non-neuronal acetylcholine (Ach) synthesis and secretion may play a role in lung diseases such as lung cancer and asthma. In order to characterize and target non-neuronal signaling in lung it is necessary to identify a step that differs between neuronal and non-neuronal cholinergic signaling. Choline uptake is a rate-limiting step for ACh synthesis and may provide this target. In neurons, the sodium-dependent, high-affinity choline transporter (CHT1) is absolutely required to transport choline into neurons for ACh synthesis. Our preliminary data shows that CHT1 is not required by some lung cells for ACh synthesis and that instead a newly described class of choline transporters, the choline transporter-like proteins (CTL1 - 5), can mediate ACh synthesis. Therefore non-neuronal ACh synthesis in lung may potentially be targeted without effecting neuronal ACh synthesis by targeting the correct CTL. To test this we propose the following specific aims: (1) Identification of which choline transporter-like proteins are involved in choline-uptake in lung non-neuronal cells. (2) Identification of which CTL is coupled to non-neuronal ACh synthesis in lung. (3) To determine if knockdown of the CTL linked to ACh synthesis can block lung cell proliferation. These aims will be implemented using specific siRNAs to knock down CTLs in H82 small cell lung carcinoma cells as a model for pulmonary neuroendocrine cells and H520 squamous cell lung carcinoma cells as a model for airway epithelial cells. These studies will thus allow the targeting of non-neuronal ACh synthesis as a first step in developing new therapies for lung diseases.