Kondo Naoyuki, Ueda Yoshihiro, Kita Toshiyuki, Ozawa Madoka, Tomiyama Takashi, Yasuda Kaneki, Lim Dae-Sik, Kinashi Tatsuo
Department of Molecular Genetics, Institute of Biomedical Science, Kansai Medical University, Hirakata, Osaka, Japan.
Division of Gastroenterology and Hepatology, Third Department of Internal Medicine, Kansai Medical University, Hirakata, Osaka, Japan.
Mol Cell Biol. 2017 Mar 31;37(8). doi: 10.1128/MCB.00424-16. Print 2017 Apr 15.
Antigen-specific adhesion between T cells and antigen-presenting cells (APC) during the formation of the immunological synapse (IS) is mediated by LFA-1 and ICAM-1. Here, LFA-1-ICAM-1 interactions were measured at the single-molecule level on supported lipid bilayers. High-affinity binding was detected at low frequencies in the inner peripheral supramolecular activation cluster (SMAC) zone that contained high levels of activated Rap1 and kindlin-3. Rap1 was essential for T cell attachment, whereas deficiencies of ste20-like kinases, Mst1/Mst2, diminished high-affinity binding and abrogated central SMAC (cSMAC) formation with mislocalized kindlin-3 and vesicle transport regulators involved in T cell receptor recycling/releasing machineries, resulting in impaired T cell-APC interactions. We found that NDR1 kinase, activated by the Rap1 signaling cascade through RAPL and Mst1/Mst2, associated with and recruited kindlin-3 to the IS, which was required for high-affinity LFA-1/ICAM-1 binding and cSMAC formation. Our findings reveal crucial roles for Rap1 signaling via NDR1 for recruitment of kindlin-3 and IS organization.
免疫突触（IS）形成过程中，T细胞与抗原呈递细胞（APC）之间的抗原特异性黏附由淋巴细胞功能相关抗原1（LFA-1）和细胞间黏附分子1（ICAM-1）介导。在此，我们在支持脂质双分子层上以单分子水平测量了LFA-1-ICAM-1相互作用。在包含高水平活化Rap1和踝蛋白3（kindlin-3）的内周超分子活化簇（SMAC）区域，以低频检测到高亲和力结合。Rap1对T细胞附着至关重要，而类丝裂原活化蛋白激酶激酶激酶（ste20样激酶）Mst1/Mst2的缺陷减少了高亲和力结合，并废除了中央SMAC（cSMAC）的形成，同时伴有踝蛋白3的定位错误以及参与T细胞受体循环/释放机制的囊泡转运调节因子异常，导致T细胞与APC的相互作用受损。我们发现，由Rap1信号级联通过RAPL和Mst1/Mst2激活的NDR1激酶与踝蛋白3相关联，并将其招募至免疫突触，这是高亲和力LFA-1/ICAM-1结合和cSMAC形成所必需的。我们的研究结果揭示了通过NDR1的Rap1信号在踝蛋白3招募和免疫突触组织中的关键作用。
