Abstract Expression of the STA1-3 glucoamylase genes, responsible for starch degradation in Saccharomyces ce- revisiae, is down regulated by the presence of STA10. In order to elucidate the role of STA10 in the regulation of the glucoamylase system, a multicopy genomic library was constructed and screened for genes that enhanced growth of a STA2-STA10 S. cerevisiae strain on starch media. This screen allowed us to clone and characterize a novel acti- vator gene of STA2 (and by extrapolation, STA1 and STA3), designated MSS11. A strain transformed with multiple cop- ies of MSS11 exhibits increased levels of STA2 mRNA and, consequently, increased glucoamylase activity. Deletion of MSS11, located on chromosome XIII, results in media-de- pendent absence of glucoamylase synthesis. MSS11 has not been cloned previously and the encoded protein, Mss11p, is not homologous to any other known protein. An out- standing feature of Mss11p is that the protein contains re- gions of 33 asparagine residues interrupted by only three serine residues, and 35 glutamine residues interrupted by a single histidine residue. Epistasis studies showed that de- letion of MSS11 abolishes the activation of STA2 caused by the over-expression of MSS10, a previously identified gene. In turn, it was found that deletion of MSS10 still allows activation of STA2 by over-expression of MSS11. Mss11p therefore appears to be positioned below Mss10p in a signal transduction pathway. Key words Saccharomyces cerevisiae · Starch utilization · Glucoamylase · Transcriptional repression/activation Introduction The ability of S. cerevisiae to utilize starch is dependent on the expression of one of three homologous genes, STA1 (DEX2, MAL5), STA2 (DEX1) and STA3 (DEX3), situated on chromosomes IV, II and XIV, respectively (for recent reviews see Pretorius et al. 1991; Pretorius 1997). These three genes each encodes one of the glycosylated, extra- cellular glucoamylase isozymes, Sta1p, Sta2p or Sta3p, re- spectively (Erratt and Stewart 1978; Tamaki 1978). It was shown previously that glucoamylase levels produced in starch and glycerol/ethanol media are similar, and that dif- ferent components of the STA2 promoter are required for cells to grow on these media (Lambrechts et al. 1994). De- letion of nt-1026 to nt-1940 of STA2 caused a lack of glu- coamylase synthesis in starch medium, whereas in glyce- rol/ethanol medium this deletion mutant was still able to produce 50% of the glucoamylase synthesized by cells con- taining the wild-type STA2 promoter (Lambrechts et al. 1994). The STA1–3 genes are subject to complex regulation in- volving an interaction between positive and negative fac- tors. Transcription of STA1–3 depends on the presence of several positive regulatory genes, such as the SNF genes, which have been shown to be part of the RNA polymerase holoenzyme involved in the regulation of glucose-re- pressed genes. None of these individual genes specifically affect the expression of STA1–3 (Pretorius 1997). Tran- scription of the STA1–3 genes is regulated negatively by several mechanisms, namely, carbon-catabolite repres- sion, diploid-specific repression in most MATa/MAT α dip- loids, and, in most strains, repression through a genetically complex phenotype referred to as STA10 (Polaina and Wiggs 1983; Pretorius et al. 1986; Pretorius 1997). STA1–3 expression is also reduced in liquid synthetic media (pH related effect) and inhibited in respiratory deficient (rho o or petite) mutants (Pretorius et al. 1986; Dranginis 1989; Patel et al. 1990). The STA10 phenotype is characterized by a reduction of at least 20-fold in the expression of STA1–3 (Polaina and Wiggs 1983). It has been proposed Curr Genet (1997) 32: 260–266 © Springer-Verlag 1997 Received: 23 June 1997 Alice L. Webber · Marius G. Lambrechts Isak S. Pretorius MSS11, a novel yeast gene involved in the regulation of starch metabolism ORIGINAL PAPER A. L. Webber · M. G. Lambrechts · I. S. Pretorius () Institute for Wine Biotechnology and Department of Microbiology, University of Stellenbosch, Stellenbosch, ZA-7600, South Africa Communicated by F. K. Zimmermann