Fraunhofer Institute for Cell Therapy and Immunology, 14476 Potsdam, Germany.
Institute of Experimental Anaesthesiology, Charité-Universitätsmedizin Berlin, 12203 Berlin, Germany.
Med Sci (Basel). 2022 Nov 17;10(4):63. doi: 10.3390/medsci10040063.
The sensory ion channel transient receptor potential vanilloid 1 (TRPV1) is mainly expressed in small to medium sized dorsal root ganglion neurons, which are involved in the transfer of acute noxious thermal and chemical stimuli. The Ankyrin-rich membrane spanning protein (ARMS) interaction with TRPV1 is modulated by protein kinase A (PKA) mediating sensitization. Here, we hypothesize that PKA phosphorylation sites of ARMS are crucial for the modulation of TRPV1 function, and that the phosphorylation of ARMS is facilitated by the A-kinase anchoring protein 79 (AKAP79). We used transfected HEK293 cells, immunoprecipitation, calcium flux, and patch clamp experiments to investigate potential PKA phosphorylation sites in ARMS and in ARMS-related peptides. Additionally, experiments were done to discriminate between PKA and protein kinase D (PKD) phosphorylation. We found different interaction ratios for TRPV1 and ARMS mutants lacking PKA phosphorylation sites. The degree of TRPV1 sensitization by ARMS mutants is independent on PKA phosphorylation. AKAP79 was also involved in the TRPV1/ARMS/PKA signaling complex. These data show that ARMS is a PKA substrate via AKAP79 in the TRPV1 signaling complex and that all four proteins interact physically, regulating TRPV1 sensitization in transfected HEK293 cells. To assess the physiological and/or therapeutic significance of these findings, similar investigations need to be performed in native neurons and/or in vivo.
感觉离子通道瞬时受体电位香草素 1（TRPV1）主要表达于中小直径背根神经节神经元，其参与急性伤害性热和化学刺激的传递。富含锚蛋白的跨膜蛋白（ARMS）与 TRPV1 的相互作用受蛋白激酶 A（PKA）介导的敏化作用调节。在这里，我们假设 ARMS 的 PKA 磷酸化位点对于 TRPV1 功能的调节至关重要，并且 ARMS 的磷酸化作用是由激酶锚定蛋白 79（AKAP79）促成的。我们使用转染的 HEK293 细胞、免疫沉淀、钙通量和膜片钳实验来研究 ARMS 及其相关肽中的潜在 PKA 磷酸化位点。此外，还进行了实验以区分 PKA 和蛋白激酶 D（PKD）的磷酸化作用。我们发现缺乏 PKA 磷酸化位点的 TRPV1 和 ARMS 突变体的相互作用比率不同。ARMS 突变体对 TRPV1 的敏化程度与 PKA 磷酸化无关。AKAP79 也参与了 TRPV1/ARMS/PKA 信号复合物。这些数据表明，ARMS 通过 AKAP79 成为 TRPV1 信号复合物中的 PKA 底物，并且这四种蛋白质相互物理作用，调节转染的 HEK293 细胞中 TRPV1 的敏化作用。为了评估这些发现的生理和/或治疗意义，需要在原代神经元和/或体内进行类似的研究。
