We have focused our efforts of understanding the role of SOCS family molecules into four members, namely SOCS1, SOCS3, SOCS4 and Cish. SOCS1 is highly expressed in immature thymocytes and necessary to suppress cytokine signaling in pre-selection thymocytes. During CD4/CD8 lineage commitment in the thymus, sustained and increasing amounts of SOCS1 expression is necessary for stringent CD4 lineage choice. However, it has not been known what controls SOCS1 expression during thymocyte differentiation. Interestingly, we found that SOCS1 is a direct downstream target of the transcription factor ThPOK. The zinc-finger nuclear factor ThPOK is specifically expressed in CD4 lineage cells and is induced by strong/persistent TCR signaling during CD4 lineage differentiation. ThPOK is considered to be a master regulator of CD4 lineage fate because its expression is both necessary and sufficient to impose CD4 lineage fate on positively selected thymocytes. However, why ThPOK is required for CD4 lineage choice has not been clear. Here we show that ThPOK is required for CD4 lineage choice because it upregulates transcription and expression of SOCS1, and other SOCS family molecules, including SOCS3 and Cish. To test the idea that ThPOK induces SOCS expression to direct CD4 lineage choice, we generated a series of ThPOK-transgenic mice with increasing levels of transgenic ThPOK expression. Transgenic ThPOK was functional because it redirected MHCI-selected thymocytes into CD4 lineage T cells. Interestingly, we found that increased ThPOK expression resulted in increased SOCS1, SOCS3 and Cish expression in CD4 T cells. We further identified that ThPOK directly induced SOCS1 transcription in luciferase reporter assays. ThPOK function was dependent on SOCS1 expression because CD4 lineage redirection by ThPOK was impaired when SOCS1 was absent. Reciprocally, enforced expression of SOCS1 was sufficient to generate CD4 T cells in the absence of ThPOK, indicating that a major role of ThPOK in CD4/CD8 lineage choice is to induce expression of SOCS genes. In sum, these data demonstrated a new circuitry of ThPOK and SOCS family molecules, and they revealed that SOCS1 and potentially other SOCS molecules are directly involved in CD4/CD8 lineage decision in the thymus. In addition to SOCS1, SOCS3 is also highly expressed in pre-selection thymocytes. SOCS3 expression was also upregulated by ThPOK during thymocyte differentiation. To further examine the role of SOCS3 in T cell development, we generated SOCS3 transgenic mice where the transgene is driven by a human CD2 mini-cassette. Overall T cell development in these mice was comparable to wildtype mice, except for a selective loss ( 50% reduction) of CD8SP thymocytes and peripheral CD8 T cells. These data are in agreement with a cytokine requirement for CD8 lineage differentiation, and they demonstrate that SOCS3 also plays a role in CD4/CD8 lineage choice. Interestingly, the inhibitory effect on CD8 lineage differentiation was much milder than in SOCS1 transgenic mice which have a near complete block in CD8 thymocyte generation. Indeed, SOCS3 overexpression potently suppressed IL-6-induced STAT3 activation in T cells, but only modestly suppressed IL-7-induced STAT5 phosphorylation. Of note, intrathymic IL-7 signaling is considered the most important cytokine signal for CD8 lineage choice. Collectively, these results suggest a cytokine-specific effect of SOCS3 which we are currently investigating with a broader panel of different cytokines. In addition to SOCS1 and SOCS3, we further identified SOCS4 as another SOCS family member that is highly expressed in immature thymocytes. To address its role in T cell development, we generated a T cell-specific SOCS4 transgenic mouse. We found that SOCS4 overexpression suppresses T cell development and impaired positive selection of HY-TCR transgenic thymocytes. We are currently in the process of further analyzing the molecular basis for this observation and how SOCS4 might interfere with T cell development and differentiation. Finally, we are also interrogating the role of Cish. In contrast to SOCS1 and SOCS3, Cish is expressed at low levels in thymocytes and in resting T cells. On the other hand, TCR stimulation induces expression of Cish, which is not the case for SOCS1 and SOCS3. Therefore, these results suggest distinct roles for Cish and SOCS1/SOCS3. In T cells, Cish has been shown to inhibit STAT5 phosphorylation by gc cytokines. However, why Cish expression is induced by TCR signaling rather than by cytokine signaling is unclear. Potentially, Cish could play a role as a mediator in the interplay between TCR and cytokine signaling, which in turn could be critical in T cell development and differentiation. To assess the role for Cish, we generated Cish transgenic mice by expressing a FLAG-tagged Cish cDNA under the control of the human CD2 mini-cassette. T cell development in these mice did not reveal a major difference compared to wildtype mice. We are currently analyzing the T cell function in these Cish transgenic mice.
Showing the most recent 10 out of 14 publications
