KEY FUNCTIONAL-ROLE AND LINEAGE-SPECIFIC EXPRESSION OF SELECTED HOXB GENES IN PURIFIED HEMATOPOIETIC PROGENITOR DIFFERENTIATION

Although it is well established that homeobox (HOX) genes play a key r ole in normal human embryogenesis the expression and function of HOX g enes in normal hematopoiesis is largely unknown. We have investigated by reverse transcriptase-polymerase chain reaction the mRNA expression of HOXB cluster genes (3' to 5' position in the cluster: from HOXB2 t hrough B9) in 72% to 88% purified hematopoietic progenitor cells (HPCs ) from adult peripheral blood induced in liquid suspension culture to gradual erythroid or granulopoietic (largely eosinophilic) differentia tion and maturation by differential growth factor (GF) stimulus (ie, l ow-dose interleukin-3 [IL-3] and granulocyte-macrophage colony-stimula ting factor [GM-CSF] and high-dose erythropoietin, or saturating amoun ts of IL-3/GM-CSF, respectively). only B3 is expressed in quiescent HP Cs. After GF treatment B3 expression is enhanced in the initial 24 hou rs and then through differentiation and maturation in erythroid and gr anulopoietic cultures. HOXB4 and B5 are induced at slightly later time s and expressed through maturation in both lineages, whereas B6is sele ctively induced in granulocytic differentiation. B2 is transiently exp ressed at low level in the granulopoietic pathway, whereas it is detec ted only in advanced stages of erythropoiesis: B7, B8, and B9 are esse ntially not detected. Functional studies were performed with antisense phosphorothioate oligomers to HOX mRNAs and included control analysis of the targeted mRNA. The results are strictly coherent with the HOX mRNA expression pattern: (1) anti-B3 oligomer (alpha-B3) treatment of purified HPCs induces a striking blockade of both erythroid and granul o-monocytic colony formation (similarly, (alpha-B3 treatment of K562 c ell line causes a significant dose-related inhibition of cell prolifer ation); (2) alpha-B6 selectively and markedly inhibits granulomonocyti c colony formation; (3) alpha-B4 and alpha-B5 cause a significant, les s pronounced decrease of both colony types; (4) finally, alpha-B2 and alpha-B7, -B9 exert little and no effect, respectively. These studies provide novel evidence on the coordinate expression of selected HOXB c luster genes in erythropoiesis and granulopoiesis, particularly in the early stages of differentiation: B3 apparently functions as a master gene in early hematopoiesis, whereas B6 exerts a key selective functio n in the granulopoietic pathway. (C) 1994 by The American Society of H ematology.