Conditional cell-wall mutants of Saccharomyces cerevisiae as delivery vehicles for therapeutic agents in vivo to the GI tract.
Strains of Saccharomyces cerevisiae capable of lysis upon conditional down-regulation of cell-wall biogenesis genes (SRB1 and PKC1) have been reported. Here, we show that they lyse and release recombinant protein not only under laboratory conditions, but (more importantly) under conditions found in the human stomach and duodenum. These findings provide proof that, in principle, such conditional lysis strains could be used as an integral part of a system for the oral delivery of therapeutic proteins. However, the current mechanism of conditional lysis is based on the use of the MET3 promoter which requires addition of methionine and cysteine for down-regulation of SRB1 and PKC1. This requirement makes it difficult to apply in vivo. We reasoned that promoters, suitable for in vivo down-regulation of lysis-inducing genes, could be identified amongst yeast genes whose transcript abundance is reduced under conditions found in the human gut. A microarray experiment identified a number of candidate genes with significantly reduced transcript levels under simulated human gut conditions. The greatest effects were seen with ANB1, TIR1, and MF(ALPHA)2), and we propose that their promoters have the potential to be used in vivo to achieve yeast lysis in the gut.
Omara
Walid A M
WA
Faculty of Life Sciences, The University of Manchester, Michael Smith Building, Oxford Road, Manchester M139PT, UK.
Rash
Bharat M
BM
Hayes
Andrew
A
Wickham
Martin S J
MS
Oliver
Stephen G
SG
Stateva
Lubomira I
LI
D049109:Cell Proliferation; D002473:Cell Wall / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D003545:Cysteine / Q000378:metabolism; D004386:Duodenum / Q000737:chemistry* / Q000378:metabolism; D005753:Gastric Mucosa / Q000378:metabolism; D020869:Gene Expression Profiling; D005800:Genes, Fungal; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008715:Methionine / Q000378:metabolism; D009154:Mutation; D009713:Nucleotidyltransferases / Q000235:genetics / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods; D014677:Pharmaceutical Vehicles / Q000737:chemistry*; D011401:Promoter Regions, Genetic; D011493:Protein Kinase C / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012441:Saccharomyces cerevisiae / Q000737:chemistry* / Q000166:cytology / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D013270:Stomach / Q000737:chemistry*
D014677:Pharmaceutical Vehicles; D029701:Saccharomyces cerevisiae Proteins; D049452:Green Fluorescent Proteins; D008715:Methionine; C490702:PKC1 protein, S cerevisiae; D011493:Protein Kinase C; D009713:Nucleotidyltransferases; C026917:mannose 1-phosphate guanylyltransferase; D003545:Cysteine