Analysis of molecular mechanisms on integrin-family specific adhesion

白细胞整合素LFA-1在白细胞迁移和免疫突触形成中起关键作用。在这些过程中，Rap 1参与了趋化因子和抗原刺激后LFA-1的亲合力调节。整合素的亲合力变化通过其胞质区域调节。为了确定参与这些过程的特定残基，我们重建了具有截短和点突变的αL和β2胞质区域的人LFA-1，并检查了它们对Rap 1依赖性粘附ICAM-1的影响。截断αL亚基胞质区而非β2亚基胞质区可消除Rap 1依赖性与ICAM-1的粘附。发现αL亚基中的两个赖氨酸残基（KK 1097/1099）在Rap-1依赖性粘附于ICAM-1中是关键的。KK 1097/1099 AA突变在SDF-1触发的ICAM-1结合中也有缺陷。另一方面，β2亚基的酪氨酸取代为丙氨酸（Y 735 A）抑制了LFA-1的内化，并损害了脱离，这导致了细长的细胞形状。我们的研究结果揭示了αL胞质区域在随后的粘附后事件中对Rap 1和β2亚基的由内而外信号传导的反应中的主要作用。

Leukocyte integrin LFA-1 plays a critical role in leukocyte migration and immunological synapse formation. In these processes, Rap1 are involved in avidity regulation of LFA-1 upon stimulation with chemokines and antigen. Avidity changes of integrins are regulated through their cytoplasmic regions. To identify specific residues that are involved in these processes, we have reconstituted human LFA-1 bearing truncated and point-mutated αL and β2 cytoplasmic regions and examined their effects on Rap1 -dependent adhesion to ICAM-1. Truncation of the αL, but not β2 subunit cytoplasmic region abolished Rap1 -dependent adhesion to ICAM-1. The two lysine residues (KK1097/1099) in the αL subunit were found to be critical in the Rap-1 dependent adhesion to ICAM-1. The KK1097/1099AA mutation was also defective in binding to ICAM-1 triggered by SDF-1. On the other hand, the substitution of tyrosine to alanine (Y735A) of the β2 subunit inhibited internalization of LFA-1, and impaired detachment, which resulted in elongated cell shape. Our findings have revealed the primary role of the αL cytoplasmic region in the response to the inside-out signaling of Rap1 and the β2 subunit in the subsequent post-adhesion events.