Biochemical and genetical analysis of
extracellular matrix proteins

Our research interest is focused on the function of extracellular matrix proteins and their receptors in relation to human diseases and to animal models. Extracellular matrices are complex structures formed by proteoglycans, collagens, and noncollagenous proteins and providing the structural integrity needed in organs and tissues. The importance of the different components has been deduced from mutant phenotypes in humans or knockout mice. Far from just having a structural function, the extracellular matrix has been linked to multiple additional roles, including regulation of cell growth, differentiation, growth factor sequestration, apoptosis, and cell migration. Further, the extracellular matrix and their receptors are key players in guiding neuronal migration and axonal outgrowth in the central and peripheral nervous system. Among these are proteins such as semaphorins, netrins, and slits.

The generous funding obtained from the Alexander von Humboldt foundation as well as the DFG allowed us to quickly establish the research group and in our research we are pursuing two main areas of interest. We are investigating the role of matrix molecules on the nervous system, and we are studying the function of novel collagens. These two projects are outlined below.

1. Nervous system
Netrins comprise a family of secreted molecules structurally related to the laminins and like them, involved in axon guidance, neuronal migration and in some cases neuronal survival. Axons sense netrins as either attractants or repellents, depending upon which netrin receptors are expressed on their growth cones, or upon differences in the cellular signal transduction machinery. To date, three netrins and their receptors (DCC, and neogenin) have been described. Netrins are structurally related to the short arms of laminin g chains with a laminin VI domain, and three epidermal growth factor-like (EGF-like) repeats as well as a positively charged heparin-binding C-terminal domain.
In a database screen, we identified a new netrin, termed netrin-4/b-netrin. The protein was detected in the basement membrane of kidney, testis, and ovary. Additional staining was observed in the smooth muscle layer surrounding arteries, heart muscle, and in the vascular system of the brain. Further, mRNA was detected in the olfactory bulb and in the retina. Netrin-4 has the ability to promote neurite outgrowth of tissue fragments from the olfactory bulb and of primary retinal ganglion cells in cell culture (in collaboration with D. Hunter and W. Brunken, Tufts). Since cell culture experiments are limited and might even be misleading, netrin-4 knockout mice were generated. The analysis of the netrin-4 knockout mice are ongoing.

2. Collagens
The collagen family of proteins plays a dominant role in maintaining the integrity of various tissues. The 19 different mammalian collagens known up to date can be subdivided into several different classes based on their polymeric assemblies and other features: fibrillar collagens (types I, II, III, V, and X), network forming collagens (types IV, VIII, and X), beaded filament forming collagens (type VI), anchoring fibril forming collagens (type VII), type XV and XVIII collagen, fibril associated collagens (types IX, XII, XIV, XVI), and transmembrane collagens (types XIII and XVII). The critical roles of collagens have been clearly illustrated by the wide spectrum of disease found in human. More than 1000 mutations in 12 out of the 19 types of collagen have been described. These diseases include osteogenesis imperfecta, many chondrodysplasias, Ehlers-Danlos syndrome, Alport syndrome, sub-types of epidermolysis bullosa etc. The latest addition of collagen induced pathology is a mutation in the COL18A1 gene causing the Knoblauch syndrome. The characterization of mutations in additional collagen genes will probably add further diseases to the list. It also has become evident that the globular domains found in many collagens also have biological functions. Major interest has been focused on endostatin, which is a proteolytic released fragment from type XVIII collagen. Endostatin as well as the noncollagenous domain of type IV collagen are potent angiogenesis inhibitor. In the lab we focus on the biochemical and genetical analysis of novel collagens. During the last years, we identified four novel collagens. Beside a novel fibrillar collagen and a transmembrane collagen, we also identified a myotendinous junction specific collagen.