title: Requirement and redundancy of the Src family kinases Fyn and Lyn in perforin-dependent killing of Cryptococcus neoformans by NK cells.
abstract: Natural killer (NK) cells directly recognize and kill fungi, such as the pathogenic fungus Cryptococcus neoformans, via cytolytic mechanisms. However, the precise signaling pathways governing this NK cell microbicidal activity and the implications for fungal recognition are still unknown. Previously, it was reported that NK cell anticryptococcal activity is mediated through a conserved phosphatidylinositol 3-kinase-extracellular signal-regulated kinase 1/2 (PI3K-ERK1/2) pathway. Using YT (a human NK-like cell line) and primary human NK cells, we sought to identify the upstream, receptor-proximal signaling elements that led to fungal cytolysis. We demonstrate that Src family kinases were activated in response to C. neoformans. Furthermore, pharmacologic inhibition with an Src kinase inhibitor blocked C. neoformans-induced downstream activation of PI3K and ERK1/2 and abrogated cryptococcal killing. At the same time, the inhibitor disrupted the polarization of perforin-containing granules toward the NK cell-cryptococcal synapse but had no effect on conjugate formation between the organism and the NK cell. Finally, small interfering RNA (siRNA) double (but not single) knockdown of two Src family kinases, Fyn and Lyn, blocked cryptococcal killing. Together these data demonstrate a mechanism whereby the Src family kinases, Fyn and Lyn, redundantly mediate anticryptococcal activity through the activation of PI3K and ERK1/2, which in turn facilitates killing by inducing the polarization of perforin-containing granules to the NK cell-cryptococcal synapse.
authors.affiliation: Departments of Microbiology, Immunology and Infectious Diseases.
authors.forename: Paul
authors.initials: P
authors.lastname: Oykhman
authors.forename: Martina
authors.initials: M
authors.lastname: Timm-McCann
authors.forename: Richard F
authors.initials: RF
authors.lastname: Xiang
authors.forename: Anowara
authors.initials: A
authors.lastname: Islam
authors.forename: Shu Shun
authors.initials: SS
authors.lastname: Li
authors.forename: Danuta
authors.initials: D
authors.lastname: Stack
authors.forename: Shaunna M
authors.initials: SM
authors.lastname: Huston
authors.forename: Ling Ling
authors.initials: LL
authors.lastname: Ma
authors.forename: Christopher H
authors.initials: CH
authors.lastname: Mody
mesh_terms: D045744:Cell Line, Tumor; D003455:Cryptococcus neoformans / Q000502:physiology*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation / Q000276:immunology; D006801:Humans; D007694:Killer Cells, Natural / Q000378:metabolism*; D021962:Membrane Microdomains; D054353:Perforin / Q000235:genetics / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D051076:Proto-Oncogene Proteins c-fyn / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D034741:RNA, Small Interfering; D014443:Tyrosine; D019061:src-Family Kinases / Q000235:genetics / Q000378:metabolism*
chemical_list: D048049:Extracellular Signal-Regulated MAP Kinases; D054353:Perforin; D019869:Phosphatidylinositol 3-Kinases; D051076:Proto-Oncogene Proteins c-fyn; D034741:RNA, Small Interfering; D014443:Tyrosine; D019061:src-Family Kinases; C494957:FYN protein, human; C060902:lyn protein-tyrosine kinase
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