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Gene annotation: Laccases and multicopper oxidases

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Presentation on theme: "Gene annotation: Laccases and multicopper oxidases"— Presentation transcript:

1 Gene annotation: Laccases and multicopper oxidases
4th Workshop of the Laccaria Genome Consortium Georg-August-Universität, Göttingen - November 16-17, 2006 P.J. Hoegger1, S. Kilaru1, P.E. Courty2, F. Martin2, U. Kües1 1Georg-August-University Göttingen, Germany 2INRA Nancy, France

2 Multi-copper oxidase family
main members: - laccases - fungi - plants - insects - bacteria - ascorbate oxidases - ferroxidases - ceruloplasmins - mammals

3 Laccase function and application
Biological functions: - lignin degradation - lignification - pigment synthesis - fruiting body formation - melanin polymerization - detoxification - cuticle sclerotization - soil litter polymer degrad. Applications: - bleaching processes - bioremediation - composite wood production

4 1 type-1 Cu 1 type-2 Cu 2 type-3 Cu
Trametes versicolor laccase Lcc1 Piontek et al. 2002, JBC 277,

5 Fungal laccase structure
Signal sequence Substrate binding loops I II III IV H H H H H H H HCH H L1 L2 L3 L4 signature sequences, including copper ligands

6 MCOs in the genome of L. bicolor
Scaffold 39 lcc11 lcc1 262 kb 222 kb 25 kb Scaffold 69 Scaffold 2 lcc2 73 kb 175 kb lcc3 112 kb 3166 kb Species MCO members C. cinerea 17 C. neoformans 6 U. maydis P. chrysosporium 5 F. graminearum 13 M. grisea 11 N. crassa 10 A. nidulans 7* C. globosum 7 C. immitis 2 R. oryzae yeasts 3-5 S. pombe 1 Scaffold 13 Scaffold 47 lcc4 194 kb 904 kb lcc5 116 kb 312 kb Scaffold 14 lcc6 429 kb lcc7 lcc8 0.53 kb 569 kb Scaffold 67 Scaffold 27 lcc9 143 kb 110 kb lcc10 647 kb 65 kb

7 Multi-copper oxidase phylogenetic tree
basidiomycete laccases ascomycete insect laccases Cel NP fungal pigment MCOs fungal ferroxidases Mgr Mco7 Sce AAB64948 Cgl XP Kla XP ascorbate oxidases "ferroxidases / laccases" Pch AAO42609 MCO1 Pch AAS21669 MCO4 Pch AAS21659 MCO2 Pch AAS21662 MCO3 Mgr Mco1 Fgr Mco1 Uma Mco1 Ego NP Uma Mco3 Cne Mco5 Cne A36962 Cne Mco6 Cim Mco2 Fgr Mco10 0.1 bacterial MCOs plant LMCOs sensu stricto Multi-copper oxidase phylogenetic tree group nr. sequences basidiomycetes 146 ascomycetes 108 zygomycetes 5 plants 62 animals 12 bacteria 29 total 362

8 L. bicolor MCOs in phylogenetic tree
LbiLcc1 LbiLcc2 LbiLcc3 LbiLcc4 LbiLcc5 LbiLcc6 LbiLcc7 LbiLcc8 LbiLcc9 basidiomycete laccases ascomycete insect laccases Cel NP fungal pigment MCOs fungal ferroxidases Mgr Mco7 Sce AAB64948 Cgl XP Kla XP ascorbate oxidases "ferroxidases / laccases" Pch AAO42609 MCO1 Pch AAS21669 MCO4 Pch AAS21659 MCO2 Pch AAS21662 MCO3 Mgr Mco1 Fgr Mco1 Uma Mco1 Ego NP Uma Mco3 Cne Mco5 Cne A36962 Cne Mco6 Cim Mco2 Fgr Mco10 0.1 bacterial MCOs plant LMCOs sensu stricto L. bicolor MCOs in phylogenetic tree LbiLcc11 LbiLcc10

9 (Aphyllophorales, Agaricales) wood and litter decomposers
wood decaying species (Aphyllophorales, Agaricales) wood and litter decomposers (Agaricales)

10 Intron position conservation in basidiomycete laccase genes

11 Tree based on intron positions

12 L. bicolor MCO gene family
Scaffold 39 lcc11 lcc1 262 kb 222 kb 25 kb Scaffold 69 Scaffold 2 lcc2 73 kb 175 kb lcc3 112 kb 3166 kb Scaffold 13 Scaffold 47 lcc4 194 kb 904 kb lcc5 116 kb 312 kb Scaffold 14 lcc6 429 kb lcc7 lcc8 0.53 kb 569 kb Scaffold 67 Scaffold 27 lcc9 143 kb 110 kb lcc10 647 kb 65 kb

13 Syn. / non-syn. substitution rates

14 L. bicolor MCOs in phylogenetic tree
LbiLcc1 LbiLcc2 LbiLcc3 LbiLcc4 LbiLcc5 LbiLcc6 LbiLcc7 LbiLcc8 LbiLcc9 basidiomycete laccases ascomycete insect laccases Cel NP fungal pigment MCOs fungal ferroxidases Mgr Mco7 Sce AAB64948 Cgl XP Kla XP ascorbate oxidases "ferroxidases / laccases" Pch AAO42609 MCO1 Pch AAS21669 MCO4 Pch AAS21659 MCO2 Pch AAS21662 MCO3 Mgr Mco1 Fgr Mco1 Uma Mco1 Ego NP Uma Mco3 Cne Mco5 Cne A36962 Cne Mco6 Cim Mco2 Fgr Mco10 0.1 bacterial MCOs plant LMCOs sensu stricto L. bicolor MCOs in phylogenetic tree LbiLcc11 LbiLcc10

15 Kosman 2003, Mol Microbiol, 47:1185-1197
Iron uptake in fungi Kosman 2003, Mol Microbiol, 47:

16 lcc10 and lcc11 loci ftr1a lcc10 ftr1b lcc11 nucleotide id. 89% 86%
179 bp iron permease 331 bp ferroxidase 115 bp ftr1a lcc10 nucleotide id. 89% 86% aa id. / sim. 91% / 95% 84% / 88% ftr1b lcc11 205 bp

17 Syn. / non-syn. substitution rates of lcc10/lcc11 and ftr1a/ftr1b
observed syn. sub. potential syn. sub. ratio observed non-syn. sub. potential non-syn. sub. ftr1a / ftr1b 52 280 0.19 38 890 0.04 lcc10 / lcc11 81 410 0.20 53 1318

18 Intron position conservation in basidiomycete ferroxidase genes

19 MCO arsenal in basidiomycete species

20 MCO arsenal in basidiomycete species

21 MCO expression profile
MCO gene L 0.5 % L 2 % W 0.1 % W 0.4 % Fb M lcc1 1.60 ± 0.23 4.97 ± 0.26 2.59 ± 0.43 2.68 ± 0.03 1.60 ± 0.20 5.68 ± 0.88 lcc2 1.97 ± 0.26 1.01 ± 0.16 1.08 ± 0.14 3.07 ± 0.56 1.50 ± 0.17 7.56 ± 0.81 lcc3 1 lcc4 0.94 ± 0.13 0.64 ± 0.25 0.80 ± 0.18 0.91 ± 0.05 0.65 ± 0.15 0.33 ± 0.08 lcc5 0.87 ± 0.13 0.86 ± 0.13 0.44 ± 0.09 2.38 ± 0.34 1.63 ± 0.22 0.59 ± 0.24 lcc6 1.09 ± 0.31 0.98 ± 0.33 1.34 ± 0.13 1.13 ± 0.35 1.21 ± 0.38 0.46 ± 0.03 lcc7 1.29 ± 0.20 0.67 ± 0.36 1.19 ± 0.19 1.17 ± 0.34 14.49 ± 1.18 0.69 ± 0.09 lcc8 1.04 ± 0.28 0.69 ± 0.11 0.32 ± 0.10 0.95 ± 0.19 0.76 ± 0.23 0.56 ± 0.04 lcc9 1.30 ± 0.44 0.86 ± 0.22 1.02 ± 0.24 0.79 ± 0.12 1.72 ± 0.24 0.21 ± 0.12 lcc10 3.83 ± 0.16 1.79 ± 0.51 3.10 ± 0.45 1.27 ± 0.23 1.26 ± 0.26 0.54 ± 0.09 lcc11 1.27 ± 0.50 0.32 ± 0.19 0.36 ± 0.08 3.45 ± 0.78 0.22 ± 0.03 1.49 ± 0.28

22 Acknowledgements Molecular Wood Biotechnology, Institute of Forest Botany, Georg-August-University Göttingen, Germany Ursula Kües Patrik Hoegger Sreedhar Kilaru UMR 1136 Interactions Arbres Micro-organismes, Centre INRA de Nancy, Champenoux, France Francis Martin Marc Buée Pierre-Emmanuel Courty

23 filamentous ascomycetes
MCO arsenal in fungal species basidiomycetes filamentous ascomycetes ascomycetous yeasts z

24 lcc10 and lcc11 loci ftr1a lcc10 ftr1b lcc11 nucleotide id. 89% 86%
179 bp 331 bp 115 bp ftr1a lcc10 nucleotide id. 89% 86% aa id. / sim. 91% / 95% 84% / 88% ftr1b lcc11 205 bp frame- shift? A>G T/C>G G>A A/C>T


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