In response to changes in nutrient conditions, cells rearrange the composition of plasma membrane (PM) transporters to optimize their metabolic flux. Not only transcriptional gene regulation, but also inactivation of specific transporters is important for fast rearrangement of the PM. In eukaryotic cells, endocytosis plays a role in transporter inactivation, which is triggered by ubiquitination of these transporters. The Nedd4 family E3 ubiquitin ligase is responsible for ubiquitination of the PM transporters and requires that a series of α-arrestin proteins are targeted to these transporters. The mechanism by which an α-arrestin recognizes its cognate transporters in response to environmental signals is of intense scientific interest. Excess substrates or signal transduction pathways are known to initiate recognition of transporters by α-arrestins. Here, we identified an endocytic-sorting signal in the monocarboxylate transporter Jen1 from yeast (), whose endocytic degradation depends on the Snf1-glucose signaling pathway. We found that the C-terminal 20-amino acid-long region of Jen1 contains an amino acid sequence required for association of Jen1 to the α-arrestin Rod1, as well as lysine residues important for glucose-induced Jen1 ubiquitination. Notably, fusion of this region to the methionine permease, Mup1, whose endocytosis is normally induced by excess methionine, was sufficient for Mup1 to undergo glucose-induced, Rod1-mediated endocytosis. Taken together, our results demonstrate that the Jen1 C-terminal region acts as a glucose-responding degron for α-arrestin-mediated endocytic degradation of Jen1.
In the yeast Saccharomyces cerevisiae, the mitochondrial branched-chain amino acid (BCAA) aminotransferase Bat1 plays an important role in the synthesis of BCAAs (valine, leucine and isoleucine). Our upcoming study [Large et al. 2020
Awamori shochu is a traditional distilled alcoholic beverage made from steamed rice in Okinawa, Japan. Although it has a unique aroma that is distinguishable from that of other types of shochu, no studies have been reported on the breeding of awamori yeasts. In yeast, isoamyl alcohol (i-AmOH), known as the key flavor of bread, is mainly produced from α-ketoisocaproate in the pathway of L-leucine biosynthesis, which is regulated by end-product inhibition of α-isopropylmalate synthase (IPMS). Here, we isolated mutants resistant to the L-leucine analog 5,5,5-trifluoro-DL-leucine (TFL) derived from diploid awamori yeast of Saccharomyces cerevisiae. Some of the mutants accumulated a greater amount of intracellular L-leucine, and among them, one mutant overproduced i-AmOH in awamori brewing. This mutant carried an allele of the LEU4 gene encoding the Ser542Phe/Ala551Val variant IPMS, which is less sensitive to feedback inhibition by L-leucine. Interestingly, we found that either of the constituent mutations (LEU4(S542F) and LEU4(A551V)) resulted in the TFL tolerance of yeast cells and desensitization to L-leucine feedback inhibition of IPMS, leading to intracellular L-leucine accumulation. Homology modeling also suggested that L-leucine binding was drastically inhibited in the Ser542Phe, Ala551Val, and Ser542Phe/Ala551Val variants due to steric hindrance in the cavity of IPMS. As we expected, awamori yeast cells expressing LEU4(S542F), LEU4(A551V), and LEU4(S542F/A551V) showed a prominent increase in extracellular i-AmOH production, compared with that of cells carrying the vector only. The approach described here could be a practical method for the breeding of novel awamori yeasts to expand the diversity of awamori taste and flavor.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.