Reporter gene insertions reveal a strictly B lymphoid-specific expression pattern of Pax5 in support of its B cell identity function

The transcription factor Pax5 is essential for B cell commitment and development. Although the detailed Pax5 expression pattern within the hemopoietic system is still largely unknown, we previously reported that Pax5 is monoallelically transcribed in pro-B and mature B cells. In this study, we have investigated the expression of Pax5 at single-cell resolution by inserting a GFP or human cd2 indicator gene under the translational control of an internal ribosomal entry site element into the 3' untranslated region of Pax5. These insertions were noninvasive, as B cell development was normal in Pax5(ihCd2/ihCd2) and Pax5(iGFP/iGFP) mice. Transheterozygous Pax5(ihCd2/iGFP) mice coexpressed GFP and human CD2 at similar levels from pro-B to mature B cells, thus demonstrating biallelic expression of Pax5 at all stages of B cell development. No reporter gene expression could be detected in plasma cells and non-B cells of the hemopoietic system. Moreover, the vast majority of common lymphoid progenitors and pre-pro-B cells in the bone marrow Pax5(iGFP/iGFP) mice did not yet express GFP, indicating that Pax5 expression is fully switched on only during the transition from uncommitted pre-pro-B cells to committed pro-B cells. Hence, the transcriptional initiation and B cell-specific expression of Pax5 is entirely consistent with its B cell lineage commitment function.