ABSTRACT
To investigate the specificity of cytokine signals in hematopoietic differentiation, we generated mice with a targeted mutation of their G-CSF receptor (G-CSFR) such that the cytoplasmic (signaling) domain of the G-CSFR is replaced with the cytoplasmic domain of the erythropoietin receptor. In homozygous mutant mice, expression of this chimeric receptor had no apparent affect on lineage commitment and was able to support the production of morphologically mature neutrophils. However, mutant neutrophils displayed reduced chemotaxis, and G-CSF-stimulated mobilization of neutrophils and hematopoietic progenitors from the bone marrow to blood was markedly impaired. Thus, the G-CSFR is generating unique signals that are required for certain specialized hematopoietic cell functions but are not required for granulocytic differentiation or lineage commitment.
Subject(s)
Hematopoiesis , Hematopoietic Stem Cells/physiology , Receptors, Granulocyte Colony-Stimulating Factor/physiology , Animals , Chemotaxis, Leukocyte/drug effects , Female , Flow Cytometry , Granulocyte Colony-Stimulating Factor/pharmacology , Hematopoiesis/drug effects , Hematopoietic Stem Cell Mobilization , Interleukin-8/pharmacology , Mice , Mice, Inbred C57BL , Mice, Knockout , Neutrophils/drug effects , Neutrophils/immunology , Receptors, Erythropoietin/physiologyABSTRACT
Research concerning oligonucleotide-directed triple helix formation has mainly focused on the binding of DNA oligonucleotides to duplex DNA. The participation of RNA strands in triple helices is also of interest. For the pyrimidine motif (pyrimidine.purine.pyrimidine triplets), systematic substitution of RNA for DNA in one, two, or all three triplex strands has previously been reported. For the purine motif (purine.purine.pyrimidine triplets), studies have shown only that RNA cannot bind to duplex DNA. To extend this result, we created a DNA triple helix in the purine motif and systematically replaced one, two, or all three strands with RNA. In dramatic contrast to the general accommodation of RNA strands in the pyrimidine triple helix motif, a stable triplex forms in the purine motif only when all three of the substituent strands are DNA. The lack of triplex formation among any of the other seven possible strand combinations involving RNA suggests that: (i) duplex structures containing RNA cannot be targeted by DNA oligonucleotides in the purine motif; (ii) RNA strands cannot be employed to recognize duplex DNA in the purine motif; and (iii) RNA tertiary structures are likely to contain only isolated base triplets in the purine motif.
