BASEMENT MEMBRANE BIOSYNTHESIS

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Basement membranes are thin condensations of specialized extracellular matrix that underlie epithelia, surround muscle cells, and serve various physiological functions. Their pathobiological chemistry is important in several diseases, as well as in both aging and development. Their durability and replacement are not understood. We have shown that several major constituents of basement membranes: procollagen IV, laminin and entactin, are closely similar in man and Drosophila melanogaster, and have established methodology for studying basement membrane formation in Drosophila. We have cDNA clones for these materials and for some others and have obtained the complete nucleic acid sequence of Drosophila procollagen IV. In the lethal mutant, myospheroid, in which muscle basement membranes detach, we found that this gene codes for part of an extracellular cell surface receptor. We shall investigate the molecular functions of those parts of basement membrane components and their receptors which are highly similar in man and Drosophila, for example, the carboxyl (NC1) junctional domain of procollagen IV. We shall use the special advantages of the Drosophila system to reintroduce into the gene line genes that have been changed by specific, site-directed mutagenesis at evolutionary conserved 1 parts of basement membrane proteins. We shall also introduce these modified genes into Drosophila mass cell cultures that secrete basement membrane components, isolate the modified proteins, and study their interactions in vitro. We will complete our ongoing characterization of several high molecular weight glycoproteins that are secreted by Drosophila cells and are putative additional components of basement membranes. The Drosophila receptor chain that we found has a very similar vertebrate homolog, the integrin- beta chain, which is the common component of several important receptors for extracellular matrix molecules. We plan to isolate corresponding Drosophila receptors and characterize them. We have demonstrated the utility of specific antibodies and nucleic acid probes for defining the time and site of appearance of basement membrane collagen IV, and its mRNA, during Drosophila development. We will use similar probes for Drosophila laminin and entactin. We shall fuse the promoter regions of basement membrane genes to the marker beta-galactosidase, to study the action of the promoters, and will search for common controlling elements of basement membrane proteins and their receptors.