title: Genome-wide analysis of pectate-induced gene expression in Botrytis cinerea: identification and functional analysis of putative d-galacturonate transporters.
abstract: The fungal plant pathogen Botrytis cinerea produces a spectrum of cell wall degrading enzymes for the decomposition of host cell wall polysaccharides and the consumption of the monosaccharides that are released. Especially pectin is an abundant cell wall component, and the decomposition of pectin by B. cinerea has been extensively studied. An effective concerted action of the appropriate pectin depolymerising enzymes, monosaccharide transporters and catabolic enzymes is important for complete d-galacturonic acid utilization by B. cinerea. In this study, we performed RNA sequencing to compare genome-wide transcriptional profiles between B. cinerea cultures grown in media containing pectate or glucose as sole carbon source. Transcript levels of 32 genes that are induced by pectate were further examined in cultures grown on six different monosaccharides, by means of quantitative RT-PCR, leading to the identification of 8 genes that are exclusively induced by d-galacturonic acid. Among these, the hexose transporter encoding genes Bchxt15 and Bchxt19 were functionally characterised. The subcellular location was studied of BcHXT15-GFP and BcHXT19-GFP fusion proteins expressed under control of their native promoter, in a B. cinerea wild-type strain. Both genes are expressed during growth on d-galacturonic acid and the fusion proteins are localized in plasma membranes and intracellular vesicles. Target gene knockout analysis revealed that BcHXT15 contributes to d-galacturonic acid uptake at pH 5∼5.6. The virulence of all B. cinerea hexose transporter mutants tested was unaltered on tomato and Nicotiana benthamiana leaves.
authors.affiliation: Laboratory of Phytopathology, Wageningen University, Droevendaalsesteeg 1, 6708 PB Wageningen, The Netherlands; Department of Biology, University of Kaiserslautern, Kaiserslautern, Germany. Electronic address: lzhang@rhrk.uni-kl.de.
authors.forename: Lisha
authors.initials: L
authors.lastname: Zhang
authors.affiliation: Laboratory of Phytopathology, Wageningen University, Droevendaalsesteeg 1, 6708 PB Wageningen, The Netherlands.
authors.forename: Chenlei
authors.initials: C
authors.lastname: Hua
authors.affiliation: Laboratory of Phytopathology, Wageningen University, Droevendaalsesteeg 1, 6708 PB Wageningen, The Netherlands; Department of Animal and Plant Sciences, University of Sheffield, United Kingdom.
authors.forename: Joost H M
authors.initials: JHM
authors.lastname: Stassen
authors.affiliation: Laboratory of Phytopathology, Wageningen University, Droevendaalsesteeg 1, 6708 PB Wageningen, The Netherlands.
authors.forename: Sayantani
authors.initials: S
authors.lastname: Chatterjee
authors.affiliation: Laboratory of Phytopathology, Wageningen University, Droevendaalsesteeg 1, 6708 PB Wageningen, The Netherlands.
authors.forename: Maxim
authors.initials: M
authors.lastname: Cornelissen
authors.affiliation: Laboratory of Phytopathology, Wageningen University, Droevendaalsesteeg 1, 6708 PB Wageningen, The Netherlands.
authors.forename: Jan A L
authors.initials: JAL
authors.lastname: van Kan
mesh_terms: D020171:Botrytis / Q000187:drug effects* / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D002462:Cell Membrane / Q000201:enzymology; D003470:Culture Media / Q000737:chemistry; D022162:Cytoplasmic Vesicles / Q000201:enzymology; D015870:Gene Expression / Q000187:drug effects*; D020869:Gene Expression Profiling; D055786:Gene Knockout Techniques; D016681:Genome, Fungal; D006603:Hexuronic Acids / Q000378:metabolism*; D018551:Solanum lycopersicum / Q000382:microbiology; D026901:Membrane Transport Proteins / Q000096:biosynthesis* / Q000235:genetics; D010368:Pectins / Q000378:metabolism*; D060888:Real-Time Polymerase Chain Reaction; D014026:Nicotiana / Q000382:microbiology; D014774:Virulence
chemical_list: D003470:Culture Media; D006603:Hexuronic Acids; D026901:Membrane Transport Proteins; C007819:galacturonic acid; D010368:Pectins; C003181:polygalacturonic acid
keywords: Botrytis cinerea; Transcriptome; Transporter; d-Galacturonic acid