Additional file 3.
Figure S1. Pictures of P. carnosa and P. chrysosporium grown on various carbon sources. P. carnosa grew significantly slower than P. chrysosporium; as a result, pictures were taken after 13 and 3 days of cultivation for P. carnosa and P. chrysosporium, respectively. Cultivations were performed in duplicate and no significant differences in colony diameter or thickness were observed between the duplicates on any of the carbon sources. Monomeric and oligomeric carbon sources were used at a final concentration of 25 mM, while pure polymers were used at a final concentration of 1%. Crude plant biomass was used at a final concentration of 3%. Relative growth was determined by comparing the radius and density of the mycelia on a particular carbon source to that on D-glucose. The extent of growth relative to plates containing glucose are summarized as follows, from high to non-detectable: +++, ++, +, ±, -. This semi-quantitative, consistent assessment of both colony diameter and thickness allowed comparisons to include a broad range of substrates, including those that yield too little mycelia for accurate weight measurement, or that could interfere with protein measurements or ergosterol production [84,85]. Complete growth profiles of P. carnosa and P. chrysosporium, and other fungi can be found at . Figure S2. Phylogenetic tree of GH61 enzymes from P. carnosa and P. chrysosporium. Proteins are labeled with protein IDs from the JGI databases for Phanerochaete carnosa v1.0 (Phaca) and Phanerochaete chrysosporium v2.0 (Phchr). The sequences were aligned using MAFFT, and the tree was drawn by FigTree. In the heatmap bar, abbreviations are; Y, YMPG; F, balsam fir; P, lodgepole pine; S, white spruce; M, sugar maple . Heat map represents the number of sequence reads per million kb as shown in this figure and described in , 0 (black) to 50,000 (pink). Figure S3. Phylogenetic tree of GH5 enzymes in P. carnosa and P. chrysosporium. The tree was generated as described in Figure S2. Of the 34 upregulated wood-degrading CAZymes, that were at least four times more abundant in P. carnosa grown on at least one wood substrate compared to nutrient medium, 5 (15%) were GH5 enzymes . Figure S4. Phylogenetic tree of predicted sugar transporters and permeases from genomes of P. carnosa and P. chrysosporium. Proteins are labeled with protein IDs from the JGI database for P. carnosa v1.0 (Phaca) and P. chrysosporium v2.0 (Phchr). Protein sequences of transporters from other yeast and fungal species were used for pylogenetic comparison, including; An Aspergillus nidulans, Ao Aspergillus oryzae, Ca Candida albicans, Ci Candida intermedia, Gz Gibberella zeae, Hp Hansenula polymorpha, Kl Kluyveromyces lactis, Lb Laccaria bicolor, Nc Neurospora crassa, Pp Postia placenta, Ps Pichia stipitis, Sc Saccharomyces cerevisiae, Sp Schizosaccharomyces pombe, Tm Tuber melanosporum, and Tr Trichoderma reesei. The GenBank accession numbers of corresponding sequences are: AnHyp1 (XP_682442.1), AnHyp2 (XP_660070.1), AnMstA (CAC80843), Ao_BAE58341.1 (BAE58341.1), CaHgt1 (CAA76406), CaHgt4 (XP_723173), CaHgt11 (XP_719597), CiGxf1 (AJ937350), CiGxs1 (AJ875406), GzHyp1 (EAA74528), HpGcr1 (AAR88143), KlHgt1 (XP_451484), KlRag1 (XP_453656), KlRag4 (CAA75114), Lb_EDR07962 (EDR07962), NcHyp1 (XP_328858), NcHxt3 (CAD21508), NcNCU00801(EAA34565.1), NcNCU08114 (XP_963873.1), NcRco3 (CAE76420), Pp_115604 (EED81359), Ps_ABN65648.2 (ABN65648.2), PsSut1 (AAD00266), ScHxt1 (M82963), ScHxt7 (NP_010629), ScSnf3 (P10870), SpGht1 (Q9P3U6), Tm-CAZ81962.1 (CAZ81962.1), TrHxt1 (AAR23147), TrXlt1 (shown enlarged; AY818402), TrHxt2 (DQ852622; Ruohonen and Margolles-Clark, unpublished). The sequences were aligned using MAFFT, and the tree was drawn by FigTree. In the heatmap bar, abbreviations are: Y YMPG, F balsam fir, P lodge pole pine, S white spruce, M sugar maple . Group I contains a predicted monosaccharide transporter (ID 100265) and hypothetical proteins with high similarity to known sugar transporters. Group IV also contains known glucose transporters found in yeast species and cellulolytic fungi, including Trichoderma reesei and Neurospora crassa. Group V consists of high affinity glucose transporters in yeast and hexose transporters in fungal species, including the xylose transporter found in T. reesei. Group VI and VII contains predicted sugar transporters and putative sucrose transporters (ID 89844 and 254080). Group VIII consists of predicted cellobiose transporters found in yeasts and filamentous fungi, and biochemically characterized cellodextrin transporters from N. crassa (NcNCU00801 and 08114) . Figure S5. Phylogeny, genome position, and intron distribution of genes encoding manganese peroxidases and lignin peroxidases. Protein IDs of manganese peroxidases (A) and lignin peroxidases (B) of P. carnosa and P. chrysosporium are obtained from P. carnosa v1.0 and P. chrysosporium v2.0. Alternative names are from MacDonald et al.  for P. carnosa and from Vanden Wymelenberg et al.  for P. chrysosporium.Figure S6. Mycelial growth of P. carnosa and P. chrysosporium on heartwood and sapwood samples isolated from different hardwood and softwood species. Colony diameter was measured for P. carnosa grown on heartwood (A) and sapwood (B), and for P. chrysosporium grown on heartwood (C) and sapwood (D). Filled square (red), sugar maple; filled circle (green), yellow birch; filled triangle (blue), trembling aspen; filled diamond (pink), red spruce; open square (purple), white spruce; open circle (orange), balsam fir; open triangle (gray), red pine. Error bars show the standard deviation in biological triplicates. Since P. chrysosporium mycelia was no longer visible after day 8 of cultivation on heartwood of sugar maple, yellow birch, white spruce and balsam fir, those data were not obtained. Figure S7. Different modes of wood decay described by FT-IR analysis. (A) Grouping on Principal Components (PCs) 1 and 2 for normalized FT-IR data obtained from heartwood and sapwood samples of trembling aspen and red pine after cultivation of P. carnosa and P. chrysosporium. Circle; P. carnosa, triangle; P. chrysosporium, square; control (untreated wood samples). (B) PC loadings that distinguish wood samples treated with P. carnosa from corresponding control samples. For example, high positive loadings describe components in control samples that were lost in the decayed samples. Loadings for PC1 are shown for trembling aspen heartwood and red pine sapwood, while loadings for PC2 are shown for trembling aspen sapwood and red pine heartwood. (C) PC loadings that distinguish wood treated with P. carnosa from corresponding wood samples treated with P. chrysosporium. Horizontal dotted lines at magnitude |0.05| represent thresholds for loading significance. Corresponding wavenumbers (cm-1) are indicated for peaks with significant loadings; identities of significant wavenumbers are summarized in Additional file: Table. S17. Percent values given in y-axes denote the percent of total sample variance described by the PC.
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Suzuki et al. BMC Genomics 2012 13:444 doi:10.1186/1471-2164-13-444