Natural killer (NK) cells are cytotoxic lymphocytes that target tumor cells, virally infected cells, and microbes. The anti-tumor and anti-viral components of NK cells have been extensively studied, however much less is known about their anti-microbial activity. In this thesis I investigate NK cell anti-microbial signaling against the yeast pathogen Cryptococcus neoformans. Previous studies found that C. neoformans activates a SFK→ PI3K→ Erk cytotoxicity pathway. In order to expand our understanding I explored the role of Rac1, which is commonly an intermediate molecule linking PI3K and Erk. Surprisingly, I found that in cryptococcal killing Rac1 acts as an activator of PI3K. Additionally, I found that Rac1 and SFK were both necessary, but independent in activating PI3K.
Additionally, I explored adhesion molecules that are involved in NK cytotoxicity. I found that beta-1 integrins are needed for cryptococcal killing. Beta-1 integrins were found to stimulate an ILK→ Rac1 pathway. This pathway is independent of the NKp30 receptor, that is also required for cryptococcal killing. Although beta-1 integrins function as adhesion receptors in tumor killing, loss of function studies showed that beta-1 integrins were not required for NK cells to adhere to C. neoformans.
The lack of integrin involvement in adhesion caused me to investigate the immune synapse between C. neoformans and NK cells. I found that the adhesion force was lower compared to 721.221 tumor targets and NK cells. The adhesion force in tumor killing is dependent on actin remodeling proteins - Arp2/3 and formins. Interestingly cryptococcal killing required formins but not Arp2/3. Actin remodeling in a cryptococcal synapse was lower compared to a tumor synapse. However, inhibition of Arp2/3 reduced the level of actin remodeling in a tumor synapse to what is seen in a cryptococcal synapse.
Together, my data highlights the similarities and differences in NK cell mediated microbial killing compared to tumor killing. It reveals beta-1 integrins as possible anti-microbial receptors, a non-canonical role for Rac1 in cytotoxicity, and identifies the reduction in actin remodeling at the cryptococcal synapse as an explanation for the difference in kinetics between anti-fungal and anti-tumor killing.