The fibulins represent a novel family of extracellular matrix proteins. We report the temporo-spatial expression of fibulin-2 in skin regenerating from keratinocyte autografts. In normal dermis, fibulin-2 was associated with the fibrillin-containing microfibrillar apparatus, except for the portion immediately adjacent to the dermo-epidermal junction. In contrast, early regenerating dermis showed numerous fusiform fibrillin-microfibrils along the basement membrane, whereas fibulin-2 was present in a distinct and separate layer below. Both proteins formed independent fibrillar systems also in the reticular dermis without significant colocalization; however, over time both fibril systems became congruent: after 4 mo there was extensive colocalization of fibulin-2/fibrillin in the reticular dermis, after 17 and 24 mo this also occurred in the papillary dermis. Simultaneous visualization of fibulin-2 and fibronectin revealed an inverse pattern: complete colocalization at 7 d and discordant distribution 17-24 mo after grafting. In particular, the fibrillar fibronectin pattern at early time points changed into a faint granular distribution throughout the dermis and along the subbasement membrane region as in normal skin. Dermal fibroblast cultures showed that fibrillin and fibronectin participated in distinct fibrillar systems; however, fibulin-2 colocalized with either protein. We propose that, in regenerating skin, fibulin-2 is a late component of the cutaneous microfibrillar apparatus with an earlier existence in a fibrillar matrix mediated by fibronectin. This suggests interaction of fibulin-2 with both fibronectin fibrils and fibrillin microfibrils, and is consistent with in vitro binding data.