Sed1p interacts with Arn3p physically and mediates ferrioxamine B uptake in Saccharomyces cerevisiae.
Two-hybrid analysis can be used to study protein function and metabolic pathways. Using yeast two-hybrid analysis to identify a siderophore uptake pathway in the yeast Saccharomyces cerevisiae, we found that the C-terminal part of the cell-wall protein Sed1p interacts with the N-terminal region of Arn3p. To confirm the physical interaction between the Sed1p C-terminal fragment and the hydrophilic N-terminal fragment of Arn3p, we used an in vitro co-immunoprecipitation assay and a growth test of the strain with bait and SED1 plasmids in quadruple amino acid-depleted medium. The expression of SED1 was upregulated by overexpression of AFT1-1(up) under the control of the GAL promoter. This occurred despite the lack of an Aft1p-binding consensus region on the upstream region of SED1 or a high concentration of free iron. Although free-iron uptake activity in the Deltased1 strain did not differ from that in the parental strain, ferrioxamine bound-iron uptake activity was reduced in the Deltased1 strain. Moreover, the Deltased1 strain showed low viability at high iron concentrations. Taken together, these results suggest that Sed1p mediates siderophore transport and confers iron resistance in S. cerevisiae.
Park
Yong-Sung
YS
College of life science/Graduate School of Biotechnology, Korea University, Anam-dong, Sungbuk-gu, Seoul, Korea.
Jeong
Ho-Sang
HS
Sung
Ha-Chin
HC
Yun
Cheol-Won
CW
D001693:Biological Transport, Active / Q000235:genetics; D003676:Deferoxamine / Q000378:metabolism*; D005290:Ferric Compounds / Q000378:metabolism*; D007502:Iron Chelating Agents; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000502:physiology*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*
C410163:Arn1 protein, S cerevisiae; D005290:Ferric Compounds; D007502:Iron Chelating Agents; D008562:Membrane Glycoproteins; D026901:Membrane Transport Proteins; C077394:SED1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C002577:ferrioxamine B; D003676:Deferoxamine