Bioinformatics annotation of Pycnoporus sanguineus BAFC 2126 transcriptome

ROHR CRISTIAN; LEVIN LAURA; MENTABERRY ALEJANDRO; WIRTH SONIA

Rosario

Congreso; IV Argentinean Conference on Computational Biology and Bioinformatics & the IV Conference of the Iberoamerican Society for Bioinformatics (SOIBIO); 2013

CIFASIS-CONICET, UNR, AMU

Background Plant cell walls are mainly composed of cellulose, hemicellulose and lignin and they constitute the most abundant source of organic carbon on Earth. Fungi of the genus Pycnoporus are white-rot basidiomycetes widely studied because they are particularly effective in using lignocellulose of plant cell walls as carbon source through the synthesis of a considerable number of hydrolytic enzymes, including cellulases, hemicellulases, pectinases, lignin-modifying enzymes and other accessory enzymes which can be employed in a wide range of industrial processes [1]. Materials and methods Poly(A) RNA was purified from total RNA extracted from Pycnoporus sanguineus BAFC2126 mycelia grown on GA medium supplemented with copper sulfate as laccase inductor. cDNA libraries were prepared and sequenced using the Roche 454 GS FLX Titanium pyrosequencing platform (INDEAR, Rosario, Argentina). Reads were assembled using Newbler v2.6 (Roche). Similarities BLAST search for the assembled transcripts were done against the NCBI non- redundant (nr) protein database and Uniprot using BLASTx algorithm. Blast2GO suite was used to annotate the transcripts with Gene ontology information. KEGG pathways were annotated using KEGG Automatic Annotation Server. Enzyme commission numbers were assigned from the blast top hits. Best open reading frames were predicted using OrfPredictor and blast vs. NCBI nr, then analyzed using SignalP and TargetP. HMMSEARCH from HMMER package was used to scan the transcripts against the PFAM and TIGRFAM domain databases. Carbohydrate Active Enzymes family prediction was done using CAZYmes Analysis Toolkit and manually curated. Results We identified 7304 transcripts and putative functions for 4736 were manually assigned and curated. We found 178 putative carbohydrate active enzymes, including representatives of 15 families with roles in lignocellulose degradation. Furthermore, we identified many enzymes related to lignin hydrolysis and modification, including laccases and peroxidases, GMC-oxidoreductases, copper radical oxidases and other enzymes involved in the generation of extracellular hydrogen peroxide and iron homeostasis in non-enzymatic lignin degradation processes. We also, identified all of the enzymes involved in terpenoid backbone biosynthesis pathway and various terpene synthases related to the biosynthesis of sesquiterpenoids and triterpenoids precursors, and also cytochrome P450 monooxygenases, glutathione S-transferases and epoxide hydrolases with potential functions in the biodegradation of xenobiotics and the enantioselective biosynthesis of biologically active drugs. Conclusions The availability of this first reference transcriptome for the genus Pycnoporus could facilitate the analysis and annotation of additional sequencing projects and provide a tool for the study of metabolic pathways and for the cloning of enzymes with biotechnological interest.