Expression of a previously cloned secretory protein named adrenocortical zonation factor 1 (AZ-1, also called Tin-ag-RP or lipocalin 7) is tightly linked with the zonal differentiation of adrenocortical cells. It is also present in vascular smooth muscle (VSM), although its function has remained unknown. In this study, the location of AZ-1 was specified to the basal laminae along adrenocortical sinusoidal capillaries and surrounding VSM cells in the arterial system, consistent with the fact that AZ-1 was extractable under denaturing conditions as a 52 kDa polypeptide. Purified recombinant AZ-1 exhibited abilities to bind to fibronectins via the first type III repeat (anastellin) and to collagens with affinities in submicromolar ranges. AZ-1 immobilized on substratum or bound to collagens or anastellin promoted adhesion and spreading of adrenocortical cells. Although VSM cells spread on AZ-1 slowly, AZ-1 bound to anastellin facilitated the spreading. The adhesion activity of AZ-1 was mediated by a subset of integrins, including alpha(1)beta(1), alpha(2)beta(1), and alpha(5)beta(1), in a cell type-specific manner. Collectively with the putative role of AZ-1 in the adrenocortical zonation, we propose that AZ-1 potentially regulates functions of adrenocortical and VSM cells by modulating cell-matrix interactions.