Dose -dependent transcriptional regulation by the calcineurin/NFAT signaling in developing myocardium transition

Abstract

Multiple transcription factors and signaling pathways are involved in the regulation and function of the myocardium in heart development. Disruption of certain transcription factors or critical components of signaling pathways frequently causes structural malformation in heart and persistence of "thin spongy myocardium". Thin spongy myocardium is critical at early embryonic stage [before embryonic day (E) 13.5 in mice] to allow diffusion of oxygen and nutrient to the developing cardiomyocytes. However, establishment of compact myocardium at later stage (-E16.5) during development is necessary to prepare for the increase in demand for blood circulation. Molecular targets of the spongy-compact myocardium transition at E13.5-E16.5 in heart development remain elusive. Previously, we have demonstrated calcineurin/NFAT activation at E14.5 in developing myocardium. Here, I report transcription targets, independently and dependently, regulated by the calcineurin/NFAT signaling during E13.5-E16.5 myocardium transition. I have uncovered that expression of one-third of the induced genes during myocardium transition is calcineurin/NFAT dependent. Among these calcineurin/NFAT dependent transcription targets, there is a dose dependent regulation. Formation of distinct NFAT:DNA complex may account for the dose-dependent regulation. Thus, in addition to temporal and spatial regulation, dose-dependent requirement provides another mechanism to modulate transcription response mediated by the calcineurin/NFAT signaling.;Among the identified genes that are dose-dependent regulated by calcineurin/NFAT signaling during myocardium transition, I focus on regenerating islet-derived 3 gamma (Reg3gamma). The expression of Reg3gamma in heart is developmentally regulated and coincident with changes of NFAT activity in heart, supporting the regulation of Reg3gamma by NFAT during heart development. Gel mobility shift assay confirmed the formation of NFAT:Reg3gamma DNA complex. Therefore, NFAT directly binds to and regulates the Reg3gamma gene promoter. In addition, I have shown that Reg3gamma is a secretory protein and its expression in intestine and heart increased upon inflammation and myocardial ischemia/reperfusion. Thus, Reg3gamma is directly regulated by NFAT and participates in heart development, inflammation and stress response.