Angiopoietin-1 (Ang-1) is required for developing vessels, and its absence
leads to defects in vessel remodeling. Ang-1 has been identified as the lig
and for the tyrosine kinase receptor Tie-a, which is expressed specifically
on endothelial cells and early hematopoietic cells. In studying the role o
f Tie-2 and Ang-1 in megakaryocytopoiesis, 3 alternatively spliced species
of Ang-1 mRNA(Ang-1.3 kb, Ang-0.9 kb, and Ang-0.7 kb) were identified in ad
dition to the full-length Ang-1 (Ang-1.5 kb), in the megakaryocyte cell lin
e CHRF by reverse transcription-polymerase chain reaction (RT-PCR), and the
n cloned and sequenced. The expression of 3 alternatively spliced isoforms
of Ang-l was confirmed by RT-PCR using specific primer pairs derived from j
unction sites and the 3' end of Ang-1 cDNA, and it was further demonstrated
by nuclease protection assay, Northern blotting, and immunoblotting in CHR
F cells. Expression of the Ang-1,3 kb isoform was also detected in human pr
imary fibroblast cell line FS4, breast cancer cell line MDAMB-463, and CD34
(+)CD41(+) cells of fetal liver and platelets. The function of the 1.5-kb,
1.3-kb, and 0.9-kb isoforms was examined. Recombinant proteins Ang-1.5 and
0.9 kb bind strongly to the recombinant Tie-2 receptor (Tie-2-Fc), whereas
the 1,3-kb isoform does not the Ang-1.3 kb isoform binds to the 1.5-kb isof
orm, Ang-1.5 kb, but not the 1,3-kb and 0.9-kb isoforms, induces tyrosine p
hosphorylation of Tie-5 in human umbilical vein endothelial cells. These da
ta suggest that isoforms 1.3 kb and 0.9 kb could serve as dominant negative
molecules for the full-length Ang-1, The possible involvement of the newly
identified Ang-1 Isoforms in angiogenesis and in growth and differentiatio
n of hematopoietic progenitor cells provides a greater complexity to these
processes. (C) 2000 by The American Society of Hematology.