PCR-mediated analysis of lignocellulolytic gene transcription by Phanerochaete chrysosporium: substrate-dependent differential expression within gene families.
We compare the kinetics of appearance of supernatant enzyme activities (lignin peroxidase, manganese peroxidase, and cellulase) and gene expression (LIG, mnp, and cbhI gene families and the unique cbhII gene) in Phanerochaete chrysosporium ME446 when grown on four different carbon sources: ball-milled straw, representing the natural substrate lignocellulose; Avicel as a crystalline cellulose; and high and low concentrations of glucose, in all cases with limiting nitrogen. PCR-based technology utilizing pairs of primers specific for particular genes showed that there is differential expression between and within the families. There were a number of instances of mRNA species being present only on a single day, implying tight regulation of lignocellulose degradation at the mRNA level. The patterns of extracellular enzyme activities and mnp and cbh gene expression are similar whereas LIG gene expression can be detected when no corresponding enzyme activity is observed in the extracellular supernatant. The enzyme produced under these conditions is presumably sequestered by the mycelium and is likely to be functionally significant. Another striking result is that cellulose, in the form of Avicel, elicits the expression of three LIG gene for which there is no expression under the same conditions with the other carbon sources.
Broda
P
P
Department of Biochemistry and Applied Molecular Biology, University of Manchester Institute of Science and Technology, United Kingdom.
Birch
P R
PR
Brooks
P R
PR
Sims
P F
PF
D001483:Base Sequence; D002480:Cellulase / Q000235:genetics* / Q000378:metabolism; D005658:Fungi / Q000201:enzymology* / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic; D008969:Molecular Sequence Data; D010544:Peroxidases / Q000235:genetics* / Q000378:metabolism; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000032:analysis; D013379:Substrate Specificity; D014158:Transcription, Genetic
D012333:RNA, Messenger; D010544:Peroxidases; C042858:lignin peroxidase; C051129:manganese peroxidase; D002480:Cellulase