Excretion of putrescine and spermidine by the protein encoded by YKL174c (TPO5) in Saccharomyces cerevisiae.
The properties of the protein encoded by YKL174c (TPO5) were studied. It was found that TPO5 excretes putrescine effectively and spermidine less effectively. Gamma-aminobutyric acid slightly inhibited the excretion of putrescine, but basic amino acids did not affect excretion, suggesting that TPO5 preferentially recognizes polyamines. Accordingly, yeast cells transformed with the plasmid encoding YKL174c (TPO5) were resistant to toxicity caused by 120 mm putrescine or by 3 mm spermidine, and a mutant with a disrupted YKL174c (TPO5) gene was sensitive to toxicity by 90 mm putrescine. The growth of this mutant was faster than that of the wild-type strain. In parallel, there was an increase in putrescine and spermidine content of the YKL174c (TPO5) mutant compared with wild-type. It is noted that TPO5 functions as a suppressor of cell growth by excreting polyamines. The level of YKL174c (TPO5) mRNA was increased by the addition of polyamines to the medium. The degree of induction of the mRNA was spermine > spermidine > putrescine. The subcellular localization of TPO5 was determined by immunostaining of hemagglutinin-tagged TPO5, and it was found on Golgi or post-Golgi secretory vesicles. Excretion of putrescine and spermidine by TPO5 was reduced in cells that have mutations in the secretory or endocytic pathways, indicating that both processes are involved in the excretion of polyamines.
Tachihara
Ken
K
Graduate School of Pharmaceutical Sciences, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba 260-8675, Japan.
Uemura
Takeshi
T
Kashiwagi
Keiko
K
Igarashi
Kazuei
K
D015152:Blotting, Northern; D015153:Blotting, Western; D027682:Cation Transport Proteins / Q000737:chemistry* / Q000502:physiology*; D049109:Cell Proliferation; D004705:Endocytosis; D019084:Fluorescent Antibody Technique, Indirect; D006056:Golgi Apparatus / Q000378:metabolism; D009154:Mutation; D010802:Phylogeny; D010957:Plasmids / Q000378:metabolism; D011073:Polyamines / Q000737:chemistry; D011700:Putrescine / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000502:physiology*; D013095:Spermidine / Q000378:metabolism*; D013347:Subcellular Fractions; D013395:Sucrose / Q000494:pharmacology; D013696:Temperature; D013997:Time Factors; D005680:gamma-Aminobutyric Acid / Q000378:metabolism
D027682:Cation Transport Proteins; D011073:Polyamines; D012333:RNA, Messenger; D029701:Saccharomyces cerevisiae Proteins; C500357:TPO5 protein, S cerevisiae; D005680:gamma-Aminobutyric Acid; D013395:Sucrose; D013095:Spermidine; D011700:Putrescine