Fluid Secretion in Parotid Cells: Signal Transduction

腮腺细胞的液体分泌：信号转导

• 批准号: 6727422
• 负责人: Stephen Paul Soltoff
• 金额: $ 37.1万
• 依托单位: BETH ISRAEL DEACONESS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-05-01 至 2006-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6727422
• 关键词: acinar cell; calcium flux; cell population study; enzyme induction /repression; immunocytochemistry; immunoprecipitation; intermolecular interaction; laboratory rat; mass spectrometry; mitogen activated protein kinase; neurogenetics; neuroregulation; neurotransmitter receptor; neurotransmitters; phosphorylation; phosphotransferases; posttranslational modifications; protein kinase C; protein localization; protein structure function; protein tyrosine kinase; receptor coupling; salivary glands; secretion; sodium channel; western blottings

The broad long term objectives of this proposal are to understand on a molecular level intracellular signals that are involved in receptor-mediated alterations of salivary cells, particularly those signals that are initiated by secretagogues involved in saliva formation (fluid secretion) by parotid acinar cells. Fluid secretion is controlled by neurotransmitters that act on muscarinic, Substance P, and alpha-adrenergic receptors, which are linked to phospholipase C (PLC) by heterotrimeric GTP- binding (G) proteins. These receptors promote the PLC-mediated generation of DAG, which activates Protein Kinase C (PKC), and IP3, which mobilizes Ca2+. Parotid acinar cells also have multiple P2X ATP-binding receptors, including P2X7 receptors, that are nonselective cation channels, and these also elevate [Ca2+]i. ATP may be a neurotransmitter or act as an autocrine signal on salivary cells. P2X receptors are not coupled to G proteins, yet activation of the P2X7 receptor appears to stimulate PKC-dependent pathways in common with the muscarinic receptor in parotid acinar cells. These pathways may require the activation of PKCdelta, the one PKC family member that is activated by phosphorylation on tyrosine residues. PKCs and MAPKs play important roles in salivary and epithelial functions, and our data indicates that diverse receptor types, including the P2X7 receptor/ion channel and the muscarinic receptor, activate both of these proteins, which may be differentially regulated by these receptors. Aim 1 is to determine the effects of signaling molecules/events, including tyrosine phosphorylation, MAPK, and Src, on indices of fluid/electrolyte/protein secretion. Aim 2 is to define and compare the signal transduction cascades that activate MAPKs in secretagogue-treated and Na pump-inhibited rat parotid acinar cells, and to use MAPK activation as a paradigm to study receptor crosstalk and other aspects of cellular control. Na pump inhibition appears to activate a distinct signaling cascade that links pump activity to MAPK. Aim 3 is to study the activation of PKC by P2X7 and other receptors. Aim 4 is to determine downstream targets of PKCdelta and other proteins by identifying binding partners of these proteins in parotid acinar cells. PKCs and MAPKs have important functions in salivary cells and epithelia, although most studies have been performed using cell lines. Tyrosine phosphorylation, Src, PKCs, and MAPK may affect ion transport proteins, tight junction permeability, exocytosis, and other salivary functions. Understanding the signaling events that control their activation, as well as the effects of these proteins on secretion, is critical to providing a basis for understanding the regulation of salivary cells in health and in disease states.

该提案的广泛长期目标是了解涉及唾液细胞的受体介导的改变的细胞内信号，尤其是那些由参与唾液形成（流体分泌）的秘密累积的信号引起的信号。 流体分泌由作用于毒蕈碱，物质P和α-肾上腺素能受体的神经递质控制，这些受体与异物三聚体GTP-结合（G）蛋白与磷脂酶C（PLC）有关。 这些受体促进了PLC介导的DAG的产生，该DAG激活了动员Ca2+的蛋白激酶C（PKC）和IP3。 腮腺腺泡细胞还具有多个P2X ATP结合受体，包括P2X7受体，它们是非选择性阳离子通道，它们也升高[Ca2+] i。 ATP可以是神经递质，也可能是唾液细胞上的自分泌信号。 P2X受体不与G蛋白偶联，但P2X7受体的激活似乎刺激了羊梨腺泡细胞中的毒蕈碱受体共同的PKC依赖性途径。 这些途径可能需要激活PKCDELTA，PKCDELTA是通过酪氨酸残基上磷酸化激活的一种PKC家族成员。 PKC和MAPK在唾液和上皮功能中起着重要作用，我们的数据表明，包括P2X7受体/离子通道在内的各种受体类型和毒蕈碱受体都激活了这两种蛋白质，这些蛋白可能受这些受体差异调节。 AIM 1是确定信号分子/事件的影响，包括酪氨酸磷酸化，MAPK和SRC对流体/电解质/蛋白质分泌指标的影响。 AIM 2是定义和比较信号转导级联反应，该级联激活MAPK在经秘密处理和Na泵抑制的大鼠腮腺腺泡细胞中，并使用MAPK激活作为研究受体串扰和其他细胞对照方面的范式。 NA泵的抑制似乎激活了将泵活动与MAPK联系起来的不同信号级联。 AIM 3是研究P2X7和其他受体对PKC的激活。 AIM 4是通过鉴定这些蛋白质的结合伴侣在腮腺细胞中的结合伴侣来确定PKCDELTA和其他蛋白质的下游靶标。 尽管大多数研究是使用细胞系进行的，但PKC和MAPK在唾液细胞和上皮中具有重要功能。 酪氨酸磷酸化，SRC，PKCS和MAPK可能会影响离子转运蛋白，紧密连接性渗透性，胞吐作用和其他唾液功能。 了解控制其激活的信号传导事件以及这些蛋白质对分泌的影响，对于理解健康和疾病状态中唾液细胞的调节的基础至关重要。