The platelet-endothelial cell adhesion molecule-1 (PE-CAM-1), defined by the CD31 monoclonal antibody (MoAb), was initially described as a cell-cell adhesion molecule mediating both homotypic and heterotypic adhesion. In this report, we show that enriched CD34+ human hematopoietic progenitor cell populations, containing early myeloid, erythroid, and multipotential progenitor cells, are CD31+. Analyses of CD34+ cell lines representing early myeloid, multipotential, and pre- pre-B-lymphoid progenitors indicate that precursors of both myeloid and B-lymphoid cells express PECAM-1 at high levels. Three-color flow- cytometric analyses also show that normal human bone marrow CD31+ CD34+ subsets coexpress myeloid (CD33) or B-lymphoid (CD19, CD10) markers. Except for the monocytic cell line, U937, all CD34- cell lines tested, which represent more mature stages of the myeloid, erythroid, and lymphoid lineages, expressed substantially lower or negligible levels of PECAM-1. Western blotting studies indicated that the CD31 MoAb, JC/70A, detected molecules in the 120- to 140-kD molecular weight range on the monocytic CD34- CD33+ CD31+ cell line, U937; on the CD34+ CD31+ CD33+ CD19- multipotential/lymphomyeloid precursor cell lines, KG1 and KG1B; on the CD34+ CD31+ CD19+ CD10+ CD33- precursor pre-pre-B-cell line, MIK-ALL; and on a CD34(+)-enriched precursor cell population from normal human bone marrow. A single molecular weight species was generally observed with enriched membrane preparations, whereas two PECAM-1 molecules were present in whole-cell lysates of cell lines and the CD34+ bone marrow cell subset. Preliminary studies show that a proportion of the PECAM-1 molecules on the lymphomyeloid/multipotential progenitor cell line, KG1, and on the monocytic cell line, U937, binds to heparin-sepharose. A soluble form of PECAM-1 also binds heparin- sepharose. The high level of expression of PECAM-1 on CD34+ cells suggests that this glycoprotein may function as a heterotypic adhesion molecule, possibly mediating multipotential, myeloid, and early-B- lymphoid precursor cell interactions with stromal cells and extracellular matrix molecules via heparan sulfate proteoglycans. It may also act as a homotypic adhesion molecule by interacting with PECAM- 1 on bone marrow stromal macrophage-like cells and endothelial cells or on endothelial cells during stem/progenitor cell migration. Thus, this molecule has the potential importance of directing both lineage commitment and trafficking of early hematopoietic progenitor cells.