Search

By means of immunocytochemistry with a battery of specific antibodies, we describe the expression of the collagen IV chain produced by the gene DCg1 during the two phases of Drosophila neurogenesis. DgC1 was not expressed in neuronal tissues as shown by in situ hybridization, but the onset of its expression in cells of mesodermal origin was concomitant with the appearance of collagen IV on early axon pathways and peripheral nerve roots in the embryonic developing CNS. A similar situation was found during imaginal CNS development at metamorphosis, when collagen IV immunoreactivity was detected on centrifugal and centripetal nerve pathways, and specially on retinula axons that develop from the eye imaginal disc towards the lamina anlage in the brain optic lobe. Our results strongly suggest that collagen IV could be involved, together with other informative molecules of basement membranes, in a dynamic process of cell-matrix interactions during the establishment of initial axon pathways and neurite outgrowth in vivo.

We have developed and characterized a battery of specific polyclonal antibodies directed against specific portions of the alpha-chain of collagen type IV synthesized in Drosophila by the gene DCg1. Here, we describe the use of these antibodies together with in situ hybridization experiments in an attempt to study the expression and localization of collagen type IV during Drosophila oogenesis and early embryogenesis. The results clearly demonstrate that DCg1 is maternally expressed by follicle cells and that the collagen type IV chain produced is stockpiled in the growing oocyte. During the gastrulation stages, this component of Drosophila basement membranes concentrated on cells involved in the gradual invaginations leading to morphogenetic furrows. The presence of collagen type IV, which is an RGD-bearing molecule, during early stages of Drosophila development is discussed in comparison with the crucial, active role its vertebrate counterpart is supposed to play in morphogenetic processes.

We have used a cloned chicken collagen cDNA sequence to help identify hypothetic members of the collagen gene family from Drosophila melanogaster. Several experimental evidences have been obtained which indicate that the Drosophila genome contains numerous collagen-like sequences. We have characterized in more detail ten distinct DNA sequences that hybridized strongly to the heterologous collagen probe. By in situ hybridization we have shown that these sequences are dispersed throughout the Drosophila genome. Two of them are shown to originate from the previously described DCg 1 and DCg 2 collagen genes. In other respects, we show that in addition to DCg 1 and DCg 2, at least five putative collagen genes are expressed during the Drosophila lifetime. These genes are unique, and some of them are seen to be transcribed into different size classes of mRNAs. Additionally, the data presented so far demonstrate that the expression of these genes is regulated temporally and/or quantitatively during the Drosophila life cycle.

We have examined directly the expression of one collagen gene (DCg1) during Drosophila melanogaster embryogenesis by means of in situ hybridization. Transcripts of this gene, which were demonstrated to encode a basement membrane type IV collagen chain, began to accumulate specifically in mesodermal derivatives at stages 12-13 of embryogenesis, and not before. Cells expressing this gene overlap, or are closely intermingled with, somatic and visceral mesoderm in stages 12-14. In stages 15-17, in addition to the strongly positive fat bodies, highly labelled cell spots are found scattered around all the parts of the gut and symmetrically on each side of the ventral nerve cord. They correspond to circulating mesodermal cells which we consider to be haemocytes or mesoblasts.

Type IV collagen, a major structural component of basement membrane, has been characterized only in vertebrates. It is unique among the collagenous proteins in that it forms specific lattice networks by end-to-end interactions. In particular, in mammals the C-terminal noncollagenous domain (NCl) of collagen IV was shown to be one of the major cross-linking sites in the network assembly. Here, we give the first direct evidence of type-IV-related collagen in invertebrates by sequence analysis of cDNA and genomic DNA clones for the 3'-end of a previously characterized Drosophila collagen gene. The data describe the C-terminal 190 amino acid residues of the triple helix and the entire noncollagenous domain (231 amino acids) of the chain encoded for by this gene. Comparison with data reported for human and mouse alpha 1(IV) chains reveals that triple-helix regions are quite different, while NC1 structures are very similar. This suggests different constraints on triple-helix and NC1 domains during evolution. Present data support the assumption that the NC1 structure originated from duplication of an ancestral sequence; the extent of both interspecies and intramolecular homologies suggests the maintenance in vertebrates and invertebrates of an ancestral specific function.

Based on data from developmental RNA profiles and in situ hybridization, we report a direct examination of the expression of one collagen gene (Dcg1) during drosophila melanogaster life cycle. These studies show, for the first time, that the expression of a collagen gene is both differential and tissue-specific during the course of development. Moreover, they demonstrate that the connective tissues in Drosophila do contain a collagen type synthesized by mesodermal tissues. Indeed the accumulation of Dcg1 transcripts was located mainly within the second instar fat bodies, the third instar lymph glands, and over adepithelial cells associated with third instar imaginal discs. In addition, these results seem to confirm the interpretation that wandering hemocytes released by the lymph glands could contribute in extracellular matrix composition in some tissues in the larva.