title: Combined extracellular matrix cross-linking activity of the peroxidase MLT-7 and the dual oxidase BLI-3 is critical for post-embryonic viability in Caenorhabditis elegans.
abstract: The nematode cuticle is a protective collagenous extracellular matrix that is modified, cross-linked, and processed by a number of key enzymes. This Ecdysozoan-specific structure is synthesized repeatedly and allows growth and development in a linked degradative and biosynthetic process known as molting. A targeted RNA interference 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 cuticles of these mutants lacks di-tyrosine cross-links, becomes permeable to dye and 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, an H2O2-generating NADPH dual oxidase, 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 cross-linking patterns of the cuticle collagens DPY-13 and COL-12. This points toward a functionally cooperative relationship for these two hypodermally expressed proteins that is essential for collagen cross-linking and proper extracellular matrix formation.
authors.affiliation: Institute of Comparative Medicine, Veterinary Faculty, University of Glasgow, Glasgow, Scotland, UK.
authors.forename: Melanie C
authors.initials: MC
authors.lastname: Thein
authors.forename: Alan D
authors.initials: AD
authors.lastname: Winter
authors.forename: Gillian
authors.initials: G
authors.lastname: Stepek
authors.forename: Gillian
authors.initials: G
authors.lastname: McCormack
authors.forename: Genevieve
authors.initials: G
authors.lastname: Stapleton
authors.forename: Iain L
authors.initials: IL
authors.lastname: Johnstone
authors.forename: Antony P
authors.initials: AP
authors.lastname: Page
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D030801:Animals, Genetically Modified; D017173:Caenorhabditis elegans / Q000196:embryology*; D029742:Caenorhabditis elegans Proteins / Q000378:metabolism*; D003094:Collagen / Q000378:metabolism*; D003432:Cross-Linking Reagents / Q000378:metabolism; D004625:Embryo, Nonmammalian / Q000378:metabolism*; D005109:Extracellular Matrix / Q000378:metabolism*; D005260:Female; D018507:Gene Expression Regulation, Developmental; D008297:Male; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D019255:NADPH Oxidases / Q000378:metabolism*; D010544:Peroxidases / Q000378:metabolism*; D010641:Phenotype; D034622:RNA Interference; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
chemical_list: D029742:Caenorhabditis elegans Proteins; D003432:Cross-Linking Reagents; D011994:Recombinant Proteins; D003094:Collagen; D010544:Peroxidases; D019255:NADPH Oxidases