Mice Lacking the Extracellular Matrix Protein WARP Develop Normally but Have Compromised Peripheral Nerve Structure and Function

Abstract

WARP is a recently identified extracellular matrix molecule with restricted expression in permanent cartilages and a distinct subset of basement membranes in peripheral nerves, muscle, and the central nervous system vasculature. WARP interacts with perlecan, and we also demonstrate here that WARP binds type VI collagen, suggesting a function in bridging connective tissue structures. To understand the in vivo function of WARP, we generated a WARPdeficient mouse strain. WARP-null mice were healthy, viable, and fertile with no overt abnormalities. Motor function and behavioral testing demonstrated that WARP-null mice exhibited a significantly delayed response to acute painful stimulus and impaired fine motor coordination, although general motor function was not affected, suggesting compromised peripheral nerve function. Immunostaining of WARP-interacting ligands demonstrated that the collagen VI microfibrillar matrix was severely reduced and mislocalized in peripheral nerves of WARP-null mice. Further ultrastructural analysis revealed reduced fibrillar collagen deposition within the peripheral nerve extracellular matrix and abnormal partial fusing of adjacent Schwann cell basement membranes, suggesting an important function for WARP in stabilizing the association of the collagenous interstitial matrix with the Schwann cell basement membrane. In contrast, other WARP-deficient tissues such as articular cartilage, intervertebral discs, and skeletal muscle showed no detectable abnormalities, and basement membranes formed normally. Our data demonstrate that although WARP is not essential for basement membrane formation or musculoskeletal development, it has critical roles in the structure and function of peripheral nerves. (3), and in the present study we identify type VI collagen as a ligand for WARP. WARP has a restricted distribution in developing cartilage tissues, where it is expressed at sites of joint cavitation and articular cartilage formation rather than cartilage structures that will undergo endochondral ossification (3). In adult tissues, WARP is highly restricted to the chondrocyte pericellular matrix in articular cartilage and fibrocartilages, where it colocalizes with perlecan and collagen VI (3). Several of the major basement membrane components have been found in the chondrocyte pericellular matrix, suggesting that this structure may be the functional equivalent of a basement membrane in cartilage tissues (4). Consistent with this hypothesis, recent data from our laboratory have demonstrated that WARP is a component of the basement membrane in a limited subset of tissues including the apical ectodermal ridge, the endomysium surrounding muscle fibers, the vasculature of the central nervous system, and the endoneurium of peripheral nerves (5). The principal components of basement membranes are type IV collagen, laminins, nidogens, and proteoglycans including perlecan; however, the composition, structure, and biological properties of basement membranes can differ considerably between different tissues (6,...

Immunocytochemical distribution of WARP (von Willebrand A domain-related protein) in the inner ear

“…Allen et al (2008) uses a sheep antibody raised against full length recombinant WARP purified from the media of transfected 293-EBNA cells. In that study, WARP was located in the mice vasculature nervous system and in perineural basement membranes, and developing muscle tissue and articular cartilage (Allen 2009). In another study Allen et al used a rabbit polyclonal antisera against VWA domain and C-terminal domains of WARP, using the GST-VWA domain (amino acids 21-212) and the maltose-binding protein-C-terminal domain (amino acids 389-415) fusion proteins, expressed in bacteria as antigens.…”

“…Allen et al (2008) have suggested that WARP may have an important role in maintaining the blood-brain barrier throughout life. There are only a handful of studies of WARP expression in mice, initially discovered in mouse chondrocytes (Fitzgerald et al, 2002; Allen et al, 2006, 2008, 2009). Further studies on WARP expression showed that WARP is present in the vasculature of neural tissues and in basement membrane structures and cardiac muscle (Allen et al, 2008).…”

“…Further studies on WARP expression showed that WARP is present in the vasculature of neural tissues and in basement membrane structures and cardiac muscle (Allen et al, 2008). A recent study shows that WARP-null mice are healthy, viable and no overt abnormalities (Allen et al, 2009). However, the null mice demonstrated diminished pain sensitivity and motor abnormalities.…”

“…Many of these matrix proteins are modular in structure, composed of protein domains (Allen et al, 2008; 2009). The identification of BM and ECM proteins in the inner ear may be clinically relevant given the identification of DFNA9 deafness and association with mutations in the secreted ECM protein cochlin (Robertson et al, 2006).…”

“…WARP is a multimeric component of the chondrocyte pericellular matrix in articular cartilage and intervertebral disc, where it interacts with the BM heparan sulfate proteoglycan perlecan (Allen et al, 2006). WARP is expressed in the vasculature of neural tissues, in the BMs of the peripheral nervous system, and in the apical ectodermal ridge of developing limb buds, and in skeletal and cardiac muscle (Allen et al, 2008, 2009). Among the suggested functions of WARP is its role in maintaining the blood-brain barrier (Allen et al, 2008).…”