Abstract
Development of T cells in the thymus is tightly controlled to continually produce functional, but not autoreactive, T cells. miRNAs provide a layer of post-transcriptional gene regulation to this process, but the role of many individual miRNAs in T-cell development remains unclear. miR-21 is prominently expressed in immature thymocytes followed by a steep decline in more mature cells. We hypothesized that such a dynamic expression was indicative of a regulatory function in intrathymic T-cell development. To test this hypothesis, we analyzed T-cell development in miR-21-deficient mice at steady state and under competitive conditions in mixed bone-marrow chimeras. We complemented analysis of knock-out animals by employing over-expression in vivo. Finally, we assessed miR-21 function in negative selection in vivo as well as differentiation in co-cultures. Together, these experiments revealed that miR-21 is largely dispensable for physiologic T-cell development. Given that miR-21 has been implicated in regulation of cellular stress responses, we assessed a potential role of miR-21 in endogenous regeneration of the thymus after sublethal irradiation. Again, miR-21 was completely dispensable in this process. We concluded that, despite prominent and highly dynamic expression in thymocytes, miR-21 expression was not required for physiologic T-cell development or endogenous regeneration.
Highlights
Development of T cells in the thymus is a tightly controlled process involving the commitment of progenitor cells to the T lineage, generation of a diverse T-cell antigen receptor (TCR) repertoire and selection of largely non-autoreactive T cell clones
For miR-21, thymic expression levels based on miRNA-sequencing were revealed to be high in DN thymocytes accompanied by an abrupt decrease toward the DP stage prior to re-expression in SP T cells
To validate global gene expression data, we sorted thymocyte populations of wildtype (WT) mice starting with ETPs (CD117hiCD25−CD44+), the most immature detectable thymocyte population, toward mature SP T cells and assessed relative expression of miR-21 (Figure 1 and Figure S1)
Summary
Development of T cells in the thymus is a tightly controlled process involving the commitment of progenitor cells to the T lineage, generation of a diverse T-cell antigen receptor (TCR) repertoire and selection of largely non-autoreactive T cell clones. In order to identify miRNAs with a putative function in T-cell development we hypothesized that such miRNAs should be expressed at high levels in at least some thymocyte populations and that such miRNAs should display a pattern of strong dynamic regulation at key developmental checkpoints. We hypothesized that high expression levels combined with strong dynamic changes in expression of miR-21 throughout different stages of T-cell development were indicative of a regulatory function in this process. To test this putative function, we analyzed the consequences of miR-21 deletion as well as overexpression in mice in vivo. We conclude that despite prominent and dynamic expression, miR-21 is dispensable for steady-state and stress-induced T-cell development in mice
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