A Hypomorphic Cbfb Allele Reveals Requirements for the Development of T Lymphocytes, Granulocytes, and Monocytes/Macrophages, and for Maintaining Proper Numbers of Hematopoietic Progenitors.

Abstract

CBFβ is the non-DNA binding subunit of the core binding factors, and is required for both the functions of Runx1 in hematopoiesis and Runx2 in osteogenesis. Homozygous disruption of Cbfb results in embryonic lethality at mid-gestation and a severe impairment of fetal liver hematopoiesis, a phenotype similar to Runx1 deficiency. To examine Cbfb requirements at later stages of hematopoiesis, we generated a hypomorphic Cbfb allele (Cbfbrss). Mice homozygous for the Cbfbrss allele have a 4- to 5-fold reduction in CBFβ protein levels, while Cbfbrss/− mice have an 8- to 10-fold reduction in CBFβ concentration. Cbfbrss/rss and Cbfbrss/− mice die at P0 for unknown reasons and display minor defects in bone ossification. Granulocyte and macrophage differentiation at E17.5 is impaired in both Cbfbrss/rss and Cbfbrss/−animals, as evidenced by a 2- to 3- fold decrease in GR1+ and Mac1+ cells. The percentages of Ter119+, B220+, and CD19+ cells are normal. Fetal liver hematopoietic progenitor numbers, as determined by cell surface marker and functional analyses, are increased approximately 2-fold. The most severe defects are in the T cell lineage. Cbfbrss/rss fetuses have reduced thymus cellularity and lower numbers of CD8 SP T cells, consistent with a role for Runx1 and Runx3 in epigenetic CD4 silencing. A further 2-fold reduction in CBFβ levels in Cbfbrss/− fetuses results in an absence of CD4/8 SP and DP cells. These hematopoietic defects were also seen upon transplantation of Cbfbrss/rss and Cbfbrss/− fetal liver cells into congenic Ly5.1/5.2 mice. Our data indicate that the threshold level of CBFβ required for normal hematopoiesis, bone development, and postnatal viability is somewhere between 25%-50% of its normal concentration. The data demonstrate that T cell development is particularly sensitive to CBFβ levels, and both granulocyte and macrophage differentiation require CBFβ. Since the conditional knockout of Runx1 in the adult mouse reported by Ichikawa et al. (Nat Med.10, 299, 2004) did not cause either granulocyte or monocyte/macrophage defects, we hypothesize that Runx2 and/or Runx3 are also required in these lineages. Hemizygosity for CBFB-MYH11, the product of the inv(16), is associated with a more severe phenotype than is seen in the Cbfb rss/− fetuses (Castilla et al. Cell87, 687, 1996). We therefore conclude that the hematopoietic consequences for cells expressing the inv(16) are equivalent to a greater than 10-fold reduction in the level of functional CBFβ.