Uncoupling of stem cell inhibition from monocyte chemoattraction in MIP-1α by mutagenesis of the proteoglycan binding site

Abstract

We have studied the role of proteoglycans in the function of Macrophage Inflammatory Protein-1 alpha (MIP-1α), a member of the proteoglycan binding chemokine family. Sequence and peptide analysis has identified a basic region within MIP-1α which appears to be the major determinant of proteoglycan binding and we have now produced a mutant of MIP-1α lacking the basic charges on two of the amino acids within this proteoglycan binding site. This mutant (Hep Mut) appears to have lost the ability to bind to proteoglycans. Bioassay of Hep Mut indicates that it has retained stem cell inhibitory properties but has a compromised activity as a monocyte chemoattractant, thus suggesting uncoupling of these two properties of MIP-1α. Receptor studies have indicated that the inactivity of Hep Mut on human monocytes correlates with its inability to bind to CCR1, a cloned human MIP-1α receptor. In addition, studies using proteoglycan deficient cells transfected with CCR1 have indicated that the proteoglycan binding site in MIP-1α is a site that is also involved in the docking of MIP-1α to the monocyte receptor. The site for interaction with the stem cell receptor must therefore be distinct, suggesting that MIP-1α utilizes different receptors for these two different biological processes.