The Combined Extracellular Matrix Cross - linking Activity of the Peroxidase MLT - 7 and the
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چکیده
The nematode cuticle is a protective collagenous extracellular matrix (ECM) that is modified, cross-linked and processed by a number of key enzymes. This Ecdyzoanspecific structure is synthesized repeatedly and allows growth and development in a linked degradative and biosynthetic process known as molting. A targeted RNAi screen using a cuticle collagen marker has been employed to identify components of the cuticle biosynthetic pathway. We have characterized an essential peroxidase, MoLT-7 (MLT-7) that is responsible for proper cuticle molting and re-synthesis. MLT-7 is an active, inhibitable peroxidase that is expressed in the cuticle synthesizing hypodermis coincident with each larval molt. mlt-7 mutants show a range of body morphology defects, most notably molt, dumpy and early larval stage arrest phenotypes that can all be complemented with a wild type copy of mlt-7. The cuticle of these mutants lack di-tyrosine cross-links, become permeable to dye, accessible to tyrosine iodination and have aberrant collagen protein expression patterns. Overexpression of MLT-7 causes mutant phenotypes further supporting its proposed enzymatic role. In combination with BLI-3, a H2O2 generating NADPH Dual oxidase (Duox), MLT-7 is essential for post-embryonic development. Disruption of mlt -7 , and particularly bli-3, via RNA interference also causes dramatic changes to the in vivo crosslinking patterns of the cuticle collagens DPY-13 and COL-12. This points towards a functionally cooperative relationship for these two hypodermally expressed proteins that is essential for collagen cross-linking and proper ECM formation. INTRODUCTION Collagenous extracellular matrices (ECMs) serve numerous critical roles and are found throughout the animal kingdom. The cuticle is an ECM that makes up the most external surface of nematodes and its roles are diverse. This tough but flexible exoskeleton maintains body shape, provides a protective barrier to the environment, and permits motility via its attachments to muscles (1). There are two distinct sets of discernible cuticular structures on the surface of the nematode Caenorhabditis elegans: circumferential indentations termed annulae, and longitudinal ridges termed alae. The latter are present only in the first and alternative third stage (dauer) larvae, and in adult C. elegans (2). At the
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