Alternatively Spliced Variants of Murine CD247 Influence T Cell Development and Activation, Revealing the Importance of the CD3fC-Terminal Region

CD247, also known as CD3f, is a crucial signaling molecule that transduces signals delivered by TCR through its three ITAMs. CD3f is required for successful thymocyte development. Three additional alternatively spliced variants of murine CD247 have been described, that is, CD3i, CD3h, and CD3g, that differ from CD3f in the C terminus such that the third ITAM is lost. Previous studies demonstrated defects in T cell development in mice expressing CD3g, but the TCR signaling pathways affected by CD3g and the impacts of the CD3i and CD3h on T cell development were not explored. In this study, we used a retrovirus-mediated gene transfer technique to express these three isoforms individually and examined the roles of them on T cell development and activation. Rag1-/- mice reconstituted with CD3h-expressing bone marrow failed to develop mature T cells. CD3i-expressing T cells exhibited similar development and activation as cells expressing CD3f. In contrast, thymic development was severely impaired in CD3g-reconstituted mice. Single -positive but not double -positive CD3g-expressing thymocytes had reduced TCR expression, and CD5 expression was decreased at the double -positive stage, suggesting a defect in positive selection. Peripheral CD3g-expressing T cells had expanded CD44hi populations and upregulation of exhaustion markers seen by flow cytometry and RNA sequencing analysis. Analysis of early signaling events demonstrated significantly reduced activation of both the PLCg1 and Akt/mTOR signaling pathways. There was also a reduction in the frequency of activation of CD3g-expressing T cells. These studies reveal the importance of the CD3f C -terminal region in T cell development and activation.