Schreiber Rainer, Ousingsawat Jiraporn, Kunzelmann Karl
Physiological Institute, University of Regensburg, Regensburg, Germany.
Front Immunol. 2025 Mar 26;16:1562871. doi: 10.3389/fimmu.2025.1562871. eCollection 2025.
Activation of T-cells is initiated by an increase in intracellular Ca, which underlies positive and negative regulation. Because the phospholipid scramblase and ion channel ANO9 (TMEM16J) was shown previously to regulated Ca signals in renal epithelial cells, we asked whether ANO9 demonstrates a similar regulation in T-cells.
We used measurements of the intracellular Ca concentration to examine the effects of ANO9 on intracellular Ca signaling and demonstrated expression of ANO9 and its effects on cellular and molecular parameters.
ANO9 was found to be expressed in human lymphocytes, including the Jurkat T-lymphocyte cell line and mouse lymphocytes. ANO9 has been shown to affect intracellular Ca signals in renal epithelial cells. Here we demonstrate the essential role of ANO9 during initiation of intracellular Ca signals in Jurkat T-cells and isolated mouse lymphocytes. ANO9 is essential for the initial rise in intracellular Ca due to influx of extracellular Ca through store-operated ORAI1 Ca entry channels. ANO9 is indispensable for T-cell function, independent on whether cells are activated by stimulation of the T-cell receptor with CD3-antibody or by PMA/phytohemagglutinin.
Upon activation of T-cells and formation of the immunological synapse, ANO9 recruits the Ca-ATPase (PMCA) to the plasma membrane, which is supported by the scaffolding protein discs large 1 (DLG1). PMCAs maintain low Ca levels near ORAI1 channels thereby suppressing Ca-inhibition of ORAI1 and thus retaining store-operated Ca entry (SOCE). It is suggested that ANO9 has a role in interorganelle communication and regulation of cellular protein trafficking, which probably requires its phospholipid scramblase function.
T 细胞的激活由细胞内 Ca 增加引发，这是正负调节的基础。由于先前已证明磷脂翻转酶和离子通道 ANO9（TMEM16J）可调节肾上皮细胞中的 Ca 信号，我们研究了 ANO9 在 T 细胞中是否表现出类似的调节作用。
我们通过测量细胞内 Ca 浓度来研究 ANO9 对细胞内 Ca 信号的影响，并证明了 ANO9 的表达及其对细胞和分子参数的影响。
发现 ANO9 在人类淋巴细胞中表达，包括 Jurkat T 淋巴细胞系和小鼠淋巴细胞。ANO9 已被证明会影响肾上皮细胞中的细胞内 Ca 信号。在此我们证明了 ANO9 在 Jurkat T 细胞和分离的小鼠淋巴细胞中细胞内 Ca 信号起始过程中的关键作用。ANO9 对于由于细胞外 Ca 通过储存操纵的 ORAI1 Ca 进入通道流入而导致的细胞内 Ca 初始升高至关重要。ANO9 对于 T 细胞功能是不可或缺的，无论细胞是通过用 CD3 抗体刺激 T 细胞受体还是通过佛波酯/植物血凝素激活。
在 T 细胞激活和免疫突触形成后，ANO9 将 Ca - ATP 酶（PMCA）募集到质膜，这得到支架蛋白盘大 1（DLG1）的支持。PMCA 在 ORAI1 通道附近维持低 Ca 水平，从而抑制 ORAI1 的 Ca 抑制作用，进而保留储存操纵的 Ca 内流（SOCE）。有人提出 ANO9 在细胞器间通讯和细胞蛋白运输调节中起作用，这可能需要其磷脂翻转酶功能。
