NORMAL AND ABNORMAL INTERFERON ALPHA RECEPTORS

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Interferons (IFN), cytokines and growth factors play important roles as a first defense against viral infections, in the treatment of different types of cancers and in the regulation of the immune and hematopoietic system. Their effects involve almost every system in the organism underscoring their broad effects and the importance of elucidating their signaling mechanisms. Cytokine, growth factors and IFNs have similar signaling mechanism that involve receptor binding and activation of tyrosine kinases and transcription factors of the Jak and Stat (Signal Transducers and Activators of Transcription) families, that results in the transcriptional activation of genes responsible for the different cytokine responses. However, little is known about the mechanisms by which cytokine receptors activate the Jak kinases and Stat factors. This application attempts to characterize the mechanisms used by cytokine receptors to activate the Jak kinases. We will test the hypothesis that similar domains present in the cytoplasmic region of cytokine receptors may be utilized for the interaction and activation of Jak kinases. However, this cytoplasmic domain may have different roles in the activation of Jak1 and Jak2 by distinct receptors. In the first Specific Aims of this proposal is focused on the characterization of the Jak1 binding domain present in different cytokine receptors that activate this kinase. We will also characterize the domains in the Jak kinases that are required for the interaction with different cytokine receptors. The second Specific Aim proposed to test different models for the interaction/ activation between cytokine receptors and Jak kinases, stressing the interaction/activation of cytokine receptors with Jak1 and Jak2. To achieve these goals we propose in vivo and in vitro studies such as the production of GST fusion protein encoding mutation of the appropriate binding sites, two hybrid system, production of recombinant proteins using in vitro translation systems, transient and stable expression of recombinant proteins in different mammalian cells. The results obtained with this proposal will be important not only to understand the signaling pathway of IFNalpha, but also signaling by different cytokine receptors.