Figure 5

Expression cellobiohydrolases and activation of the UPR. Time course of CBH1 activity on MULac (grey bars), relative plasmid copy number (black bars), cbh1 mRNA (triangles), cbh2 mRNA (open circles) and HAC1i mRNA (open squares). (a) The T.e.cbh1 probe corresponding to the catalytic domain of T.e.CBH1, and the T.r.CBM probe, corresponding to the T. reesei cbh1 CBM were used for cbh1 mRNA detection on two identical Northern blots; hybridized separately with the two radioactively labeled probes that had the same specific activity. The signals were detected using a Typhoon scanner and quantified. The cbh1 hybridization signals were first normalized to ACT1 and then to T.e.cbh1-CBM signal at 41 hours. RNA was isolated after 9, 17, 41 and 65 hour cultivation, and enzyme activities on MULac in the culture supernatants were determined at 17, 41, and 65 hours. Quantification of C.l.cbh2b mRNA and T.r.cbh2 mRNA were done as explained above for cbh1. (b) Relative plasmid copy number (black bars) in yeast cells grown overnight in YPD. The hybridization signals were normalized to T.r.cbh1 signal set as 1. (c) HAC1 hybridization signal was first normalized to ACT1 and then to T.e.cbh1 signal at 9 hours and expressed as relative units.