T-cell receptor α enhancer is inactivated in αβ T lymphocytes

Abstract

The Tcra enhancer (Eα) is essential for Tcra locus germ-line transcription and primary Vα-to-Jα recombination during thymocyte development. We found that Eα is inhibited late during thymocyte differentiation and in αβ T lymphocytes, indicating that it is not required to drive transcription of rearranged Tcra genes. Eα inactivation resulted in the disruption of functional long-range enhancer-promoter interactions and was associated with loss of Eα-dependent histone modifications at promoter and enhancer regions, and reduced expression and recruitment of E2A to the Eα enhanceosome in T cells. Enhancer activity could not be recovered by T-cell activation, by forced expression of E2A or by the up-regulation of this and other transcription factors in the context of T helper differentiation. Our results argue that the major function of Eα is to coordinate the formation of a chromatin hub that drives Vα and Jα germ-line transcription and primary rearrangements in thymocytes and imply the existence of an Eα-independent mechanism to activate transcription of the rearranged Tcra locus in αβ T cells.

Insights into the Transcriptional Regulation of the Unrearranged and Rearranged Tcra and Tcrd Genes

“…The TFs are represented by colored ovals and their identity is indicated. The Eα enhanceosome assembled in αβ T lymphocytes lacks bound E2A and HEB TFs compared to those assembled in lDP thymocytes [62,76].No differences in the binding of CTCF to HS-1´ were detected between DP thymocytes and αβ T lymphocytes, indicating the binding of this factor is not involved in the inhibition of Eα function in αβ T cells [31,33,62]. These results suggest strong binding of E2A and HEB might be essential for Eα activity.…”

“…Furthermore, expression of reporter transgenes directed by the 7.4-kb LCR containing Eα is significantly inhibited in splenocytes and αβ T lymphocytes compared to thymocytes [30,63]. The transcriptional inhibition of the unrearranged Tcra locus by Eα in SP thymocytes and αβ T lymphocytes suggests this enhancer does not contribute to the transcription of the rearranged Tcra locus in these cells [62]. In support of this hypothesis, transgenic rearranged Tcra constructs containing the 3-kb region from the downstream Cα, including Eα and HS1´, are expressed at very low and variable levels, ranging from 1 to 20% in αβ T lymphocytes [29,64].…”

“…In support of this hypothesis, transgenic rearranged Tcra constructs containing the 3-kb region from the downstream Cα, including Eα and HS1´, are expressed at very low and variable levels, ranging from 1 to 20% in αβ T lymphocytes [29,64]. Two main questions rise from these findings: How is Eα inactivated and what is the molecular mechanism for transcribing the rearranged Tcra locus in SP thymocytes and αβ T cells?Recent experiments using chromatin immunoprecipitation to compare the active and inactive Eα enhanceosomes assembled in DP thymocytes and αβ T lymphocytes, respectively, have revealed that the [14,[57][58][59]62,76]. The TFs are represented by colored ovals and their identity is indicated.…”

“…These results suggest strong binding of E2A and HEB might be essential for Eα activity. The forced expression of E2A in αβ T lymphocytes through retroviral transduction cannot recover the enhancer activity of Eα, neither alone nor in combination with the upregulation of other TFs in the context of T-cell activation or T helper differentiation [62]. Future experiments are necessary to reveal the molecular mechanism of enhancer inactivation in mature αβ T-cells by evaluating the simultaneous functional effect of E2A and HEB on Eα activity, the analysis of the molecular consequences of different signaling pathways mediated by the pre-TCR and TCRα on Eα function, and a detailed comparison of the Eα enhanceosomes assembled in αβ and γδ T lymphocytes.…”

“…The forced expression of E2A in αβ T lymphocytes through retroviral transduction cannot recover the enhancer activity of Eα, neither alone nor in combination with the upregulation of other TFs in the context of T-cell activation or T helper differentiation [62]. Future experiments are necessary to reveal the molecular mechanism of enhancer inactivation in mature αβ T-cells by evaluating the simultaneous functional effect of E2A and HEB on Eα activity, the analysis of the molecular consequences of different signaling pathways mediated by the pre-TCR and TCRα on Eα function, and a detailed comparison of the Eα enhanceosomes assembled in αβ and γδ T lymphocytes.The inhibition of Eα in SP thymocytes and αβ T lymphocytes does not preclude the enhanced transcription of the rearranged Tcra locus in these cells compared to the unrearranged Tcra locus in preselected DP thymocytes in the presence of a fully active Eα [62]. The molecular basis for the transcription of the rearranged Tcra locus is currently unknown.…”