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Meinrad Busslinger

Acquired immunity to foreign pathogens depends on the differentiation of B and T lymphocytes from hematopoietic stem cells, which is controlled by a multitude of transcription factors. We are interested in understanding the process by which transcription factors regulate the commitment of early hematopoietic progenitors to the B cell lineage and control their subsequent differentiation to mature B cells and plasma cells. We investigate the underlying transcriptional control mechanisms by using mouse transgenic, cell biological, and genome-wide molecular approaches.

Stem cell commitment in haematopoiesis

B cell commitment and development

A fundamental question in haematopoiesis is how stem cells and early progenitors become committed to a single developmental pathway, and then differentiate into mature cell types of the selected lineage. The entry of lymphoid progenitors into the B cell lineage depends on several transcription factors, including E2A, EBF1, and Pax5. E2A and EBF1 function as B cell specification factors while our work has identified Pax5 as the critical B cell commitment factor that restricts the developmental potential of lymphoid progenitors to the B cell pathway. More recently, we have shown that the transcription factor Ikaros controls signalling of the pre-B cell receptor by activating genes coding for signal transducers and repressing genes involved in the down-regulation of pre-BCR signalling. This year, we have demonstrated an essential role for the transcription factor Bhlhe41 in controlling the development and self-renewal of innate-like B cells.

B cell immunity and plasma cell function

Upon encounter of foreign antigens, mature B cells undergo affinity maturation of their B cell receptor and subsequently differentiate to plasma cells that provide acute and long-term protection against pathogens by generating and secreting high-affinity antibodies. Whereas the immune function of mature B cells strictly depends on Pax5, E2A, EBF1 and Ikaros (Figure 1), the transcriptional regulator Blimp1 controls terminal differentiation to plasma cells. By analysing the molecular mechanism of Blimp1 action, we recently showed that Blimp1 shuts down the B cell gene expression program by directly repressing B-cell-specific genes, while promoting plasma cell differentiation by activating genes involved in the unfolded protein response and in immunoglobulin secretion. 

Figure 1: Critical role of Pax5 in B cell immunity. The Cd23-Cre line deletes the floxed (fl) Pax5 allele in mature B cells of control Cd23-Cre Pax5fl/+ and experimental Cd23-Cre Pax5fl/– mice, which were immunized with the antigen NP-KLH. Immunostaining of spleen sections from control mice identified CD35+ follicular dendritic cells (red), mature IgD+ B cells (green), and CD3+ T cells (blue). However, in the absence of Pax5, germinal centers with follicular dendritic cells were not formed in experimental mice.

B cell leukemia

PAX5 mutations and translocations are frequently associated with B-cell acute lymphoblastic leukemia (B-ALL). By generating mouse models for the human PAX5-ETV6 and PAX5-JAK2 translocations, we could demonstrate that PAX5-ETV6 functions as a potent oncoprotein in combination with loss of the tumor suppressor CDKN2A,B. We are currently investigating whether the PAX5-JAK2 translocation may function as a dual-hit mutation by inactivating the PAX5 function of the translocated allele, while simultaneously activating JAK-STAT signaling.

Spatial control of V(D)J recombination

The development of B cells and αβ T cells depends on the functional rearrangement of Igh and Igk or Tcrb and Tcra loci, respectively. All four loci are large in size (0.7 to 3 megabases), have a complex structure, and undergo reversible contraction by looping in rearranging lymphocytes. Locus contraction is thus a general mechanism that juxtaposes distantly located V genes of the large V gene cluster next to D or J segments in the 3’ proximal domain, which facilitates V-(D)J recombination. We previously demonstrated an essential role of Pax5 in the control of Igh locus contraction, and identified Pax5-activated intergenic repeats (PAIRs) in the distal VH gene cluster as potential regulatory elements involved in this process.

By performing high-resolution mapping of long-range interactions, we recently showed that local interaction domains establish the three-dimensional structure of the extended Igh locus in lymphoid progenitors (Figure 2). In committed pro-B cells, these local domains engage in long-range interactions across the Igh locus (Figure 2), which depend on the regulators Pax5, YY1 and CTCF. The large VH gene cluster thereby undergoes flexible long-range interactions with the more rigidly structured 3’ proximal domain, which ensures similar participation of all VH genes in VH-DJH recombination to generate a diverse antibody repertoire.

Figure 2: Pax5-dependent long-range interactions across the Igh locus in committed pro-B cells. Uncommitted Pax5–/– Rag2–/– and committed Rag2–/– pro-B cells were analysed by 4C-sequencing using a viewpoint (HS3B in red) at the 3’ end of the Igh locus. The 4C-seq reads were plotted as reads per million mapped sequence reads (RPMs). The distinct VH gene families (different colors) in the distal, middle and proximal VH gene regions are shown together with the DH and CH elements in the 3’ proximal Igh domain.

Selected Publications

  • Kreslavsky, T., Vilagos, B., Tagoh, H., Poliakova, DK., Schwickert, TA., Wöhner, M., Jaritz, M., Weiss, S., Taneja, R., Rossner, MJ., Busslinger, M. (2017). Essential role for the transcription factor Bhlhe41 in regulating the development, self-renewal and BCR repertoire of B-1a cells. Nat Immunol. 18(4):442-455
  • Smeenk, L., Fischer, M., Jurado, S., Jaritz, M., Azaryan, A., Werner, B., Roth, M., Zuber, J., Stanulla, M., den Boer, ML., Mullighan, CG., Strehl, S., Busslinger, M. (2017). Molecular role of the PAX5-ETV6 oncoprotein in promoting B-cell acute lymphoblastic leukemia. EMBO J. 36(6):718-735
  • Minnich, M., Tagoh, H., Bönelt, P., Axelsson, E., Fischer, M., Cebolla, B., Tarakhovsky, A., Nutt, SL., Jaritz, M., Busslinger, M. (2016). Multifunctional role of the transcription factor Blimp-1 in coordinating plasma cell differentiation. Nat Immunol. 17(3):331-43
  • Wöhner, M., Tagoh, H., Bilic, I., Jaritz, M., Poliakova, DK., Fischer, M., Busslinger, M. (2016). Molecular functions of the transcription factors E2A and E2-2 in controlling germinal center B cell and plasma cell development. J Exp Med. 213(7):1201-21
  • Schwickert, TA., Tagoh, H., Gültekin, S., Dakic, A., Axelsson, E., Minnich, M., Ebert, A., Werner, B., Roth, M., Cimmino, L., Dickins, RA., Zuber, J., Jaritz, M., Busslinger, M. (2014). Stage-specific control of early B cell development by the transcription factor Ikaros. Nat Immunol. 15(3):283-93

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