Chapter 7. Future perspectives

We now have knowledge of the complete genomic organizations of type XV collagen genes in man and mouse, the functional properties of their promoter regions, the developmental dynamics during murine development, and the consequences of the use of two different promoter fragments in generating transgenic mice expressing truncated type XV collagen protein. The elucidation of the exon-intron organization of the human COL15A1 gene provides the necessary database for future studies for screening for mutations in patient samples for candidate diseases caused by mutations in this collagen. When choosing the candidate diseases, several features, such as the tissue distribution of type XV collagen and the phenotypic consequences in the genetically engineered mice, will be considered. Based on the consequences of the lack of this collagen, diseases affecting skeletal muscle and the cardiovascular system will be a top priority. The genomic clones obtained during both studies are valuable tools in e.g. studies of gene regulation and expression, as well as in preparing gene constructs for generating genetically engineered mice in lacking this collagen or expressing mutant α1(XV) chains. This study also initiated the analysis of the functional properties of the type XV collagen promoters in two species, providing putative conserved regions implicated in the gene regulation, and pinpointing to potential regions involved in the positive and negative regulation of the mouse Col15a1 gene. Future studies on the issues of gene regulation will be easier to tailor with this knowledge in hand. Further, an antibody against mouse type XV collagen was generated, providing an essential tool for the analysis of the tissue distribution of collagen XV in mature and developing mouse tissues, as well as in the analysis of transgenic mice. The knowledge of the developmental dynamics of type XV collagen is of great value, as it helps to understand the physiological consequences that the as yet unidentified mutations in type XV collagen may cause in humans. In the study of transgenic mice expressing truncated type XV collagen, two different promoter fragments used to drive the expression of the transgene resulted in clear differences in tissue specificities, indicating a better complement of regulatory elements in the longer fragment. The lack of an obvious phenotype in the lines driven by the shorter promoter, in contrast to apparent prenatal lethality in one line driven by the longer promoter fragment, needs to be further studied before detailed conclusions on the consequences of the generated mutation can be drawn. If the hypothesis of the prenatal lethality as a result of a mutation in type XV collagen gains further evidence, these mice will provide new evidence concerning the critical function of this collagen during development. Although recent developments in the field of collagen XV research have provided a wealth of information regarding its biological significance and function, there are still quite a few unresolved issues to be addressed. The biosynthesis, protein structure, and macromolecular aggregates formed by this collagen are still obscure, as well as the interactions with other matrix molecules. In addition, the role of the proteolytic fragments of type XV collagen, such as the endostatin homology region and the parent NC domain, both of which have been implicated in morphogenetic processes in type XVIII collagen, requires further evaluation. Lastly, the production of conditional knockout mice targeted to endothelial cells may also clarify the role of type XV collagen in capillary endothelia. Nonetheless, the data presented here have opened the door for these future studies by providing a number of important tools and basic knowledge needed.