title: Identification and functional analysis of two Golgi-localized UDP-galactofuranose transporters with overlapping functions in Aspergillus niger.
abstract: BACKGROUND
  Galactofuranose (Galf)-containing glycoconjugates are present in numerous microbes, including filamentous fungi where they are important for morphology, virulence and maintaining cell wall integrity. The incorporation of Galf-residues into galactomannan, galactomannoproteins and glycolipids is carried out by Golgi-localized Galf transferases. The nucleotide sugar donor used by these transferases (UDP-Galf) is produced in the cytoplasm and has to be transported to the lumen of the Golgi by a dedicated nucleotide sugar transporter.
  
  
  METHODS
  Based on homology with recently identified UDP-Galf-transporters in A. fumigatus and A. nidulans, two putative UDP-Galf-transporters in A. niger were found. Their function and localization was determined by gene deletions and GFP-tagging studies, respectively.
  
  
  RESULTS
  The two putative UDP-Galf-transporters in A. niger are homologous to each other and are predicted to contain eleven transmembrane domains (UgtA) or ten transmembrane domains (UgtB) due to a reduced length of the C-terminal part of the UgtB protein. The presence of two putative UDP-Galf-transporters in the genome was not unique for A. niger. From the twenty Aspergillus species analysed, nine species contained two additional putative UDP-Galf-transporters. Three of the nine species were outside the Aspergillus section nigri, indication an early duplication of UDP-Galf-transporters and subsequent loss of the UgtB copy in several aspergilli. Deletion analysis of the single and double mutants in A. niger indicated that the two putative UDP-Galf-transporters (named UgtA and UgtB) have a redundant function in UDP-Galf-transport as only the double mutant displayed a Galf-negative phenotype. The Galf-negative phenotype of the double mutant could be complemented by expressing either CFP-UgtA or CFP-UgtB fusion proteins from their endogenous promoters, indicating that both CFP-tagged proteins are functional. Both Ugt proteins co-localize with each other as well as with the GDP-mannose nucleotide transporter, as was demonstrated by fluorescence microscopy, thereby confirming their predicted localization in the Golgi.
  
  
  CONCLUSIONS
  A. niger contains two genes encoding UDP-Galf-transporters. Deletion and localization studies indicate that UgtA and UgtB have redundant functions in the biosynthesis of Galf-containing glycoconjugates.
authors.affiliation: Leiden University, Institute of Biology Leiden, Molecular Microbiology and Biotechnology, Sylviusweg 72, 2333 BE, Leiden, The Netherlands. joohae@gmail.com.
authors.forename: Joohae
authors.initials: J
authors.lastname: Park
authors.affiliation: Department of Molecular Cell Biology and Immunology, VU University Medical Center, van den Boechorststraat 7, 1081 BT, Amsterdam, The Netherlands. Boris.Tefsen@xjtlu.edu.cn.|Department of Biological Sciences, Xi'an Jiaotong-Liverpool University, 111 Ren Ai Road, Dushu Lake Higher Education Town, Suzhou Industrial Park, Suzhou, Jiangsu, 215123, China. Boris.Tefsen@xjtlu.edu.cn.
authors.forename: Boris
authors.initials: B
authors.lastname: Tefsen
authors.affiliation: Leiden University, Institute of Biology Leiden, Molecular Microbiology and Biotechnology, Sylviusweg 72, 2333 BE, Leiden, The Netherlands. marcheemskerk1989@hotmail.com.
authors.forename: Marc J
authors.initials: MJ
authors.lastname: Heemskerk
authors.affiliation: Leiden University, Institute of Biology Leiden, Molecular Microbiology and Biotechnology, Sylviusweg 72, 2333 BE, Leiden, The Netherlands. E.L.Lagendijk@biology.leidenuniv.nl.
authors.forename: Ellen L
authors.initials: EL
authors.lastname: Lagendijk
authors.affiliation: Leiden University, Institute of Biology Leiden, Molecular Microbiology and Biotechnology, Sylviusweg 72, 2333 BE, Leiden, The Netherlands. c.a.m.van.den.Hondel@biology.leidenuniv.nl.
authors.forename: Cees A M J J
authors.initials: CA
authors.lastname: van den Hondel
authors.affiliation: Department of Molecular Cell Biology and Immunology, VU University Medical Center, van den Boechorststraat 7, 1081 BT, Amsterdam, The Netherlands. im.vandie@vumc.nl.
authors.forename: Irma
authors.initials: I
authors.lastname: van Die
authors.affiliation: Leiden University, Institute of Biology Leiden, Molecular Microbiology and Biotechnology, Sylviusweg 72, 2333 BE, Leiden, The Netherlands. a.f.j.ram@biology.leidenuniv.nl.
authors.forename: Arthur F J
authors.initials: AF
authors.lastname: Ram
mesh_terms: D001234:Aspergillus niger / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002473:Cell Wall / Q000378:metabolism; D019143:Evolution, Molecular; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D005690:Galactose / Q000031:analogs & derivatives* / Q000378:metabolism; D017353:Gene Deletion; D020440:Gene Duplication; D006056:Golgi Apparatus / Q000378:metabolism*; D012689:Sequence Homology, Nucleic Acid; D014166:Transferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014530:Uridine Diphosphate / Q000031:analogs & derivatives* / Q000378:metabolism
chemical_list: D005656:Fungal Proteins; C104136:uridine diphosphate galactofuranose; D014530:Uridine Diphosphate; D014166:Transferases; D005690:Galactose