title: A PEST-like sequence in the N-terminal cytoplasmic domain of Saccharomyces maltose permease is required for glucose-induced proteolysis and rapid inactivation of transport activity.
abstract: Maltose permease is required for maltose transport into Saccharomyces cells. Glucose addition to maltose-fermenting cells causes selective delivery of this integral plasma membrane protein to the yeast vacuole via endocytosis for degradation by resident proteases. This glucose-induced degradation is independent of the proteasome but requires ubiquitin and certain ubiquitin conjugating enzymes. We used mutation analysis to identify target sequences in Mal61/HA maltose permease involved in its selective glucose-induced degradation. A nonsense mutation was introduced at codon 581, creating a truncated functional maltose permease. Additional missense mutations were introduced into the mal61/HA-581NS allele, altering potential phosphorylation and ubiquitination sites. No significant effect was seen on the rate of glucose-induced degradation of these mutant proteins. Deletion mutations were constructed, removing residues 2-30, 31-60, 61-90, and 49-78 of the N-terminal cytoplasmic domain, as well as a missense mutation of a dileucine motif. Results indicate that the proline-, glutamate-, aspartate-, serine-, and threonine-rich (PEST) sequence found in the N-terminal cytoplasmic domain, particularly residues 49-78, is required for glucose-induced degradation of Mal61/HAp and for the rapid glucose-induced inactivation of maltose transport activity. The decreased rate of glucose-induced degradation correlates with a decrease in the level of glucose-induced ubiquitination of the DeltaPEST mutant permease. In addition, newly synthesized mutant permease proteins lacking residues 49-78 or carrying an alteration in the dileucine motif, residues 69 and 70, are resistant to glucose-induced inactivation of maltose transport activity. This N-terminal PEST-like sequence is the target of both the Rgt2p-dependent and the Glc7p-Reg1p-dependent glucose signaling pathways.
authors.affiliation: Biology Department, Queens College and the Graduate School of the City University of New York, 65-30 Kissena Boulevard, Flushing, New York 11367, USA.
authors.forename: I
authors.initials: I
authors.lastname: Medintz
authors.forename: X
authors.initials: X
authors.lastname: Wang
authors.forename: T
authors.initials: T
authors.lastname: Hradek
authors.forename: C A
authors.initials: CA
authors.lastname: Michels
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D001692:Biological Transport / Q000187:drug effects; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D005285:Fermentation; D005656:Fungal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D005947:Glucose / Q000494:pharmacology*; D006207:Half-Life; D007930:Leucine / Q000235:genetics / Q000378:metabolism; D008320:Maltose / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000502:physiology; D026901:Membrane Transport Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009004:Monosaccharide Transport Proteins / Q000235:genetics / Q000502:physiology; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D017384:Sequence Deletion / Q000235:genetics; D015398:Signal Transduction / Q000187:drug effects; D027981:Symporters; D014452:Ubiquitins / Q000378:metabolism
chemical_list: D002352:Carrier Proteins; D005656:Fungal Proteins; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D009004:Monosaccharide Transport Proteins; C112512:RGT2 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D027981:Symporters; D014452:Ubiquitins; C067575:maltose transport system, S cerevisiae; D008320:Maltose; C021534:maltose permease; D007930:Leucine; D005947:Glucose