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Phosphorylation plays a key role in regulating many signaling pathways. Although studies investigating the phosphorylated forms of signaling pathways are now commonplace, global analysis of protein phosphorylation and kinase activity has lagged behind genomics and proteomics. We have used a kinomics approach to study the effect of virus infection on host cell signaling in infected guinea pigs. Delineating the host responses which lead to clearance of a pathogen requires the use of a matched, comparative model system. We have used two passage variants of the arenavirus Pichinde, used as a biosafety level 2 model of Lassa fever virus as it produces similar pathologies in guinea pigs and humans, to compare the host cell responses between infections which lead to either a mild, self-limiting infection or lethal disease. Using this model, we can begin to understand the differences in signaling events which give rise to these markedly different outcomes. By contextualizing these data using pathway analysis, we have identified key differences in cellular signaling matrices. By comparing these differentially involved networks, we have identified a number of key signaling "nodes" which show differential phosphorylations between mild and lethal infections. We believe that these nodes provide potential targets for the development of antiviral therapies by acting at the level of the host response rather than by directly targeting viral proteins.

Arenaviridae is a family of enveloped viruses some of which are capable of causing hemorrhagic fever syndromes in humans. In this report, we demonstrate that treatment of host cells with the tyrosine kinase inhibitor genistein inhibits infection of cells with the New World arenavirus Pichindé (PICV). The greatest degree of inhibition was observed in pre-treated target cells, but modest inhibition of infection was also seen when drug was added to cultures up to 48h after infection. We show that PICV-induced phosphorylation of the activating transcription factor-2 protein (ATF-2) and cyclic adenosine monophosphate response element binding protein (CREB) is inhibited following genistein treatment. Lastly, genistein treatment also inhibited transduction of cells with pseudotyped retrovirus particles expressing envelope proteins of the Old World arenavirus Lassa virus. These results demonstrate that kinase activity is required for arenavirus infection and that therapeutics designed to inhibit kinase activity should be explored.

Immunosuppression following infection with HIV-1 predisposes patients to a myriad of opportunistic pathogens, one of the most important of which is Mtb. Granulysin, expressed by NK cells and CTL, exhibits potent antimicrobial activity against Mtb and several other opportunistic pathogens associated with HIV-1 infection. The immune signals that promote granulysin expression in human CTL are not fully understood. Using primary human CD8+ T cells, in this study, we identify IL-21 as a strong inducer of granulysin, demonstrate that IL-21 and IL-15 activate granulysin expression within CD8+ CD45RO+ T cells, and establish a role for Jak/STAT signaling in the regulation of granulysin within CD8+ T cells. We show that infection of PBMC from healthy donors in vitro with HIV-1 suppresses granulysin expression by CD8+ T cells, concomitant with reduced p-STAT3 and p-STAT5, following activation with IL-15 and IL-21. Of note, simultaneous signaling through IL-15 and IL-21 could partially overcome the immunosuppressive effects of HIV-1 on granulysin expression by CD8+ T cells. These results suggest that HIV-1 infection of PBMC may reduce the antimicrobial profile of activated CD8+ T cells by disrupting signaling events that are critical for the induction of granulysin. Understanding the effects of HIV-1 on CD8+ T cell activation is essential to understanding the physiological basis for inadequate cytotoxic lymphocyte activity in HIV+ patients and for informed guidance of cytokine-based therapy to restore T cell function.