Pluripotent hematopoietic stem cells contain high levels of mRNA for c-kit, GATA-2, p45 NF-E2, and c-myb and low levels or no mRNA for c-fms and the receptors for granulocyte colony-stimulating factor and interleukins 5 and 7.
Pluripotent hematopoietic stem cells (PHSCs) were highly enriched from mouse bone marrow by counterflow centrifugal elutriation, lineage subtraction, and fluorescence-activated cell sorting based on high c-kit receptor expression (c-kitBR). We used reverse transcriptase polymerase chain reaction to assay the c-kitBR subset and the subsets expressing low (c-kitDULL) and no (c-kitNEG) c-kit receptor for expression of mRNA encoding hematopoietic growth factor receptors and transcription factors. The c-kitBR cells had approximately 3.5-fold more c-kit mRNA than unfractionated bone marrow cells. The c-kitDULL cells had 47-58% of the c-kit mRNA found in c-kitBR cells and the c-kitNEG cells had 4-9% of the c-kit mRNA present in c-kitBR cells. By comparing mRNA levels in c-kitBR cells (enriched for PHSCs) with those of unfractionated bone marrow, we demonstrated that c-kitBR cells contained low or undetectable levels of mRNA for c-fms, granulocyte colony-stimulating factor receptor, interleukin 5 receptor (IL-5R), and IL-7R. These same cells had moderate levels of mRNA for erythropoietin receptor, IL-3R subunits IL-3R alpha (SUT-1), AIC-2A, and AIC-2B, IL-6R and its partner gp-130, and the transcription factor GATA-1 and high levels of mRNA for transcription factors GATA-2, p45 NF-E2, and c-myb. We conclude from these findings that PHSCs are programmed to interact with stem cell factor, IL-3, and IL-6 but not with granulocyte or macrophage colony-stimulating factor. These findings also indicate that GATA-2, p45 NF-E2, and c-myb activities may be involved in PHSC maintenance or proliferation.
Orlic
D
D
Hematopoiesis Section, National Center for Human Genome Research, National Institutes of Health, Bethesda, MD 20892, USA.
Anderson
S
S
Biesecker
L G
LG
Sorrentino
B P
BP
Bodine
D M
DM
D000818:Animals; D015703:Antigens, CD / Q000096:biosynthesis; D001483:Base Sequence; D001854:Bone Marrow Cells; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000096:biosynthesis*; D050981:Erythroid-Specific DNA-Binding Factors; D005260:Female; D050982:GATA1 Transcription Factor; D050989:GATA2 Transcription Factor; D015870:Gene Expression; D016236:Genes, fms; D016259:Genes, myc; D006412:Hematopoietic Stem Cells / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D050983:NF-E2 Transcription Factor; D050984:NF-E2 Transcription Factor, p45 Subunit; D009857:Oncogenes; D016133:Polymerase Chain Reaction; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis*; D019009:Proto-Oncogene Proteins c-kit; D020598:Proto-Oncogene Proteins c-myb; D012333:RNA, Messenger / Q000032:analysis* / Q000096:biosynthesis; D020794:Receptor Protein-Tyrosine Kinases / Q000096:biosynthesis*; D016186:Receptor, Macrophage Colony-Stimulating Factor / Q000096:biosynthesis; D016184:Receptors, Colony-Stimulating Factor / Q000096:biosynthesis*; D016188:Receptors, Granulocyte Colony-Stimulating Factor / Q000096:biosynthesis; D018123:Receptors, Interleukin / Q000096:biosynthesis; D053649:Receptors, Interleukin-5; D020395:Receptors, Interleukin-7; D014157:Transcription Factors / Q000096:biosynthesis*
D015703:Antigens, CD; D017931:DNA Primers; D004268:DNA-Binding Proteins; D050981:Erythroid-Specific DNA-Binding Factors; D050982:GATA1 Transcription Factor; D050989:GATA2 Transcription Factor; C494689:Gata1 protein, mouse; C494712:Gata2 protein, mouse; D050983:NF-E2 Transcription Factor; D050984:NF-E2 Transcription Factor, p45 Subunit; C494693:Nfe2 protein, mouse; D011518:Proto-Oncogene Proteins; D020598:Proto-Oncogene Proteins c-myb; D012333:RNA, Messenger; D016184:Receptors, Colony-Stimulating Factor; D016188:Receptors, Granulocyte Colony-Stimulating Factor; D018123:Receptors, Interleukin; D053649:Receptors, Interleukin-5; D020395:Receptors, Interleukin-7; D014157:Transcription Factors; D019009:Proto-Oncogene Proteins c-kit; D020794:Receptor Protein-Tyrosine Kinases; D016186:Receptor, Macrophage Colony-Stimulating Factor