AKAP350 PARTICIPATES IN LFA-1 CLUSTERING AT THE IMMUNE SYNAPSE IN EXVIVO NATURAL KILLER CELLS

PARIANI A; ALMADA E; RIVABELLA MAKNIS T; FAVRE C; LAROCCA MC

Congreso; Reunión Conjunta de la Sociedad Argentina de Investigaciones Clinicas y de la Sociedad Argentina de Fisiología; 2022

AKAP350 PARTICIPATES IN LFA-1 CLUSTERING AT THE IMMUNE SYNAPSE IN EXVIVO NATURAL KILLER CELLSPariani AP1, Almada E2, Rivabella Maknis T1, Favre C1, Larocca MC11. Instituto de Fisiología Experimental, Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Na-cional de Rosario (UNR), Rosario, Argentina.2. Instituto de Inmunología clínica y experimental de Rosario, Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Universidad Nacional de Rosario (UNR), Rosario, Argentina.Natural Killer Cells (NK) are cytotoxic cells from the innate immune system. They form a specialized junction with their target cells, the immune synapse (IS), which is highly dependent on NK receptorssuch as the integrin LFA-1. AKAP350 is an scaffold protein central for microtubule nucleation at the Golgi apparatus that, in NK derived YTS cells, participates in LFA-1 recruitment to the IS, thus condi-tioning NK cytolytic activity. The aim of this work was to analyze the participation of AKAP350 in LFA-1 clustering in ex vivo NK cultures. Methods: ex vivo NK (eNK) from volunteer blood donors were puri-fied by negative selection and cultured in IL-2 supplemented medium. eNK with reduced expression of AKAP350 (AKAP350KD) were prepared by transduction with shRNA expression lentiviral particles.eNK were exposed to erythroleukemia KT-86 cells (2:1 ratio) for 30 minutes. LFA-1 distribution was analyzed by immunofluorescence confocal microscopy. Relative distance to the IS (RD) was estimat-ed as the difference between the distance from each LFA-1 vesicle to the IS and the distance from the cell centroid to the IS, related to the latter. Results are expressed as media±standard error. Results and Conclusion: Our results revealed the presence of an intracellular pool of LFA-1 which partially colocalized with the Golgi apparatus in eNK cells. LFA-1 vesicles polarization to the IS was impaired in AKAP350KD cells ( CONTROL: -0.14±0.01, AKAP350KD: 0.05±0.02, n=30, p