Determinants of phosphatidylinositol-4-phosphate 5-kinase type Iγ90 uropod location in T-lymphocytes and its role in uropod formation

We have previously identified phosphatidylinositol-4-phosphate 5-kinase type I (PIPKI)γ 90 as a T cell uropod component. However, the molecular determinants and functional consequences of its localization remain unknown. In this report, we seek to better understand the mechanisms involved in PIPKIγ 90 uropod targeting and the role that PIPKI 90 plays in T cell uropod formation. During T cell activation, PIPKIγ 90 cocaps with the membrane microdomain-associated proteins flotillin-1 and -2 and accumulates in the uropod. We report that the C-terminal 26 amino acid extension of PIPKIγ 90 is required for its localization to the uropod. We further use T cells fromPIPKI 90^-l- mice and human T cells expressing a kinase-dead PIPKIγ 90 mutant to examine the role of PIPKIγ 90 in a T cell uropod formation. We find that PIPKIγ 90 deficient T cells have elongated uropods on ICAM-1.Moreover, in human T cells overexpression of PIPKIγ 87, a naturally occurring isoformlacking the last 26 amino acids, suppresses uropod formation and impairs capping of uropod proteins such as flotillins. Transfection of human T cells with a dominant-negative mutant of flotillin-2 in turn attenuates capping of PIPKIγ 90. Our data contribute to the understanding of the molecular mechanisms that regulate T cell uropod formation.