title: Membrane-associated collagens with interrupted triple-helices (MACITs): evolution from a bilaterian common ancestor and functional conservation in C. elegans.
abstract: BACKGROUND
  Collagens provide structural support and guidance cues within the extracellular matrix of metazoans. Mammalian collagens XIII, XXIII and XXV form a unique subgroup of type II transmembrane proteins, each comprising a short N-terminal cytosolic domain, a transmembrane domain and a largely collagenous ectodomain. We name these collagens as MACITs (Membrane-Associated Collagens with Interrupted Triple-helices), and here investigate their evolution and conserved properties. To date, these collagens have been studied only in mammals. Knowledge of the representation of MACITs in other extant metazoans is lacking. This question is of interest for understanding structural/functional relationships in the MACIT family and also for insight into the evolution of MACITs in relation to the secreted, fibrillar collagens that are present throughout the metazoa.
  
  
  RESULTS
  MACITs are restricted to bilaterians and are represented in the Ecdysozoa, Hemichordata, Urochordata and Vertebrata (Gnathostomata). They were not identified in available early-diverging metazoans, Lophotrochozoa, Echinodermata, Cephalochordata or Vertebrata (Cyclostomata). Whereas invertebrates encode a single MACIT, collagens XIII/XXIII/XXV of jawed vertebrates are paralogues that originated from the two rounds of en-bloc genome duplication occurring early in vertebrate evolution. MACITs have conserved domain architecture in which a juxta-membrane furin-cleavage site and the C-terminal 34 residues are especially highly conserved, whereas the cytoplasmic domains are weakly conserved. To study protein expression and function in a metazoan with a single MACIT gene, we focused on Caenorhabditis elegans and its col-99 gene. A col-99 cDNA was cloned and expressed as protein in mammalian CHO cells, two antibodies against COL-99 protein were generated, and a col-99-bearing fosmid gene construct col-99::egfp::flag was used to generate transgenic C. elegans lines. The encoded COL-99 polypeptide is 85 kDa in size and forms a trimeric protein. COL-99 is plasma membrane-associated and undergoes furin-dependent ectodomain cleavage and shedding. COL-99 is detected in mouth, pharynx, body wall and the tail, mostly in motor neurons and muscle systems and is enriched at neuromuscular junctions.
  
  
  CONCLUSIONS
  Through identification of MACITs in multiple metazoan phyla we developed a model for the evolution of MACITs. The experimental data demonstrate conservation of MACIT molecular and cellular properties and tissue localisations in the invertebrate, C. elegans.
authors.affiliation: Centre of Excellence in Cell-Extracellular Matrix Research, Faculty of Biochemistry and Molecular Medicine, Biocenter Oulu, University of Oulu, Aapistie 5, Oulu, FIN 90014, Finland. hongmin.tu@oulu.fi.
authors.forename: Hongmin
authors.initials: H
authors.lastname: Tu
authors.affiliation: Centre of Excellence in Cell-Extracellular Matrix Research, Faculty of Biochemistry and Molecular Medicine, Biocenter Oulu, University of Oulu, Aapistie 5, Oulu, FIN 90014, Finland. pirkko.huhtala@oulu.fi.
authors.forename: Pirkko
authors.initials: P
authors.lastname: Huhtala
authors.affiliation: Centre of Excellence in Cell-Extracellular Matrix Research, Faculty of Biochemistry and Molecular Medicine, Biocenter Oulu, University of Oulu, Aapistie 5, Oulu, FIN 90014, Finland. elitemao@googlemail.com.
authors.forename: Hang-Mao
authors.initials: HM
authors.lastname: Lee
authors.affiliation: School of Biochemistry, University of Bristol, Biomedical Sciences Building, University Walk, Bristol, BS8 1TD, UK. jo.adams@bristol.ac.uk.
authors.forename: Josephine C
authors.initials: JC
authors.lastname: Adams
authors.affiliation: Centre of Excellence in Cell-Extracellular Matrix Research, Faculty of Biochemistry and Molecular Medicine, Biocenter Oulu, University of Oulu, Aapistie 5, Oulu, FIN 90014, Finland. taina.pihlajaniemi@oulu.fi.
authors.forename: Taina
authors.initials: T
authors.lastname: Pihlajaniemi
mesh_terms: D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D017173:Caenorhabditis elegans / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D029742:Caenorhabditis elegans Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D003094:Collagen / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D006224:Cricetinae; D003412:Cricetulus; D019143:Evolution, Molecular; D007814:Larva / Q000378:metabolism; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment
chemical_list: C000607485:COL-99 protein, C elegans; D029742:Caenorhabditis elegans Proteins; D003094:Collagen