Abstract. Endoglin is a homodimeric membrane glycoprotein which can bind the 131 and 133 isoforms of transforming growth factor-13 (TGF-13). We reported previously that endoglin is upregulated during monocyte differentiation. We have now observed that TGF-13 itself can stimulate the expression of endoglin in cultured human monocytes and in the U-937 monocytic line. To study the functional role of endoglin, stable transfectants of U-937 cells were generated which overexpress L-or S-endoglin isoforms, differing in their cytoplasmic domain. Inhibition of cellular proliferation and downregulation of c-myc mRNA which are normally induced by TGF-t31 in U-937 cells were totally abrogated in L-endoglin transfectants and much reduced in the S-endoglin transfectants. Inhibition of proliferation by TGF-132 was not altered in the transfectants, in agreement with the isoform specificity of endoglin. Additional responses of U-937 cells to TGF-131, including stimulation of fibronectin synthesis, cellular adhesion, platelet/endothelial cell adhesion molecule i (PECAM-1) phosphorylation, and homotypic aggregation were also inhibited in the endoglin transfectants. However, modulation of integrin and PECAM-1 levels and stimulation of mRNA levels for TGF-131 and its receptors R-I, R-II, and betaglycan occurred normally in the endoglin transfectants. No changes in total ligand binding were observed in L-endoglin transfectants relative to mock, while a 1.5-fold increase was seen in S-endoglin transfectants. The degradation rate of the ligand was the same in all transfectants. Elucidating the mechanism by which endoglin modulates several cellular responses to TGF-131 without interfering with ligand binding or degradation should increase our understanding of the complex pathways which mediate the effects of this factor.

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Endoglin is an homodimeric membrane antigen with capacity to bind transforming growth factor-beta (TGF-beta) and whose expression is up-regulated on myeloid cells upon differentiation to macrophages. We have isolated full-length cDNA clones from a lambda gt 10 library, prepared from phorbol 12-myristate 13-acetate-differentiated HL60 cells by screening with an endoglin-specific cDNA probe from endothelial cells. Sequencing of the largest clone (3073 bp), revealed that the leader sequence contains 25 residues and that the 586 amino acids of the extracellular and transmembrane domains were identical to those described for endothelial endoglin. However, the cytoplasmic tail encoded by this cDNA clone contains only 14 amino acids as opposed to the 47 residues previously reported, suggesting the existence of two alternative endoglin variants. The expression of these isoforms was demonstrated by polymerase chain reaction analyses on endothelial cells, myelomonocytic cell lines HL-60 and U-937, and placenta. Independent cDNA constructs corresponding to both forms were transfected into mouse fibroblasts leading to the expression of two distinct endoglin molecules. Both forms were shown to bind TGF-beta 1 and, when overexpressed in transfected mouse fibroblasts, to form disulfide-linked homodimers, indicating that the cysteine residues present in the extracellular domain are responsible for the dimerization.