Blood, Vol. 93 No. 2 (January 15), 1999:
pp. 527-536
D3: A Gene Induced During Myeloid Cell Differentiation of
Linlo c-Kit+ Sca-1+ Progenitor
Cells
Sarah R. Weiler,
John M. Gooya,
Mariaestela Ortiz,
Schickwann Tsai,
Steven J. Collins, and
Jonathan R. Keller
From the Laboratory of Molecular Immunoregulation, Division of Basic
Sciences, and the Intramural Research and Support Program, Science
Applications International Corp-Frederick, National Cancer
Institute-Frederick Cancer Research and Development Center, Frederick,
MD; and the Fred Hutchinson Cancer Research Center, Seattle, WA.
In an effort to characterize molecular events contributing to
lineage commitment and terminal differentiation of stem/progenitor cells, we have used differential display reverse
transcription polymerase chain reaction (DDRT-PCR) and
cell lines blocked at two distinct stages of differentiation. The cell
lines used were EML, which is representative of normal multipotential
primitive progenitors (Sca-1+, CD34+,
c-Kit+, Thy-1+) able to differentiate into
erythroid, myeloid, and B-lymphoid cells in vitro, and MPRO, which is a
more committed progenitor cell line, with characteristics of
promyelocytes able to differentiate into granulocytes. One clone
isolated by this approach was expressed in MPRO but not in EML cells
and contained sequence identical to the 3
untranslated region of
D3, a gene cloned from activated peritoneal macrophages of unknown
function. We have observed a novel pattern of D3 gene expression and
found that D3 is induced in EML cells under conditions that promote
myeloid cell differentiation (interleukin-3 [IL-3], stem cell factor
[SCF], and all-trans-retinoic acid [atRA]) starting at 2 days,
corresponding to the appearance of promyelocytes. D3 RNA
expression reached a maximum after 5 days, corresponding to the
appearance of neutrophilic granulocytes and macrophages, and decreased
by day 6 with increased numbers of differentiated neutrophils and
macrophages in vitro. Induction of D3 RNA in EML was dependent on IL-3
and was not induced in response to SCF or atRA alone or SCF in
combination with 15 other hematopoietic growth factors (HGF) tested.
Similarly, D3 was not expressed in the normal bone marrow cell (BMC)
counterpart of EML cells, Linlo c-Kit+
Sca-1+ progenitor cells. D3 RNA expression was induced in
these cells when cultured for 7 days in IL-3 plus SCF. A comparison of
the expression of D3 RNA in cell lines and normal BMC populations demonstrated that D3 is induced during macrophage and granulocyte differentiation and suggests a potential physiological role for D3 in
normal myeloid differentiation.
