Identification of the plasma membrane H+-biotin symporter of Saccharomyces cerevisiae by rescue of a fatty acid-auxotrophic mutant.
Bakers' yeast is auxotrophic for biotin (vitamin H) and depends on the efficient uptake of this compound from the environment. A mutant strain with strongly reduced biotin uptake and with reduced levels of protein biotinylation was identified. The strain was auxotrophic for long-chain fatty acids, and this auxotrophy could be suppressed with high levels of biotin in the medium. After transformation of this mutant with a yeast genomic library, the unassigned open reading frame YGR065C was identified to complement this mutation. This gene codes for a protein with 593 amino acids and 12 putative transmembrane helices. Northern blot analysis revealed that, in wild-type cells, the corresponding mRNA levels were increased at low biotin concentrations. Likewise, cellular biotin uptake was increased with decreasing biotin availability. Expression of YGR065C under the control of the constitutive ADH1 promoter resulted in very high biotin transport rates across the plasma membrane that were no longer regulated by the biotin concentration in the growth medium. We conclude that YGR065C encodes the first biotin transporter identified for a non-mammalian organism and designate this gene VHT1 for vitamin H transporter 1.
Stolz
J
J
Lehrstuhl Botanik II, Molekulare Pflanzenphysiologie, Universität Erlangen-Nürnberg, Staudtstrasse 5, D-91058 Erlangen, Germany.
Hoja
U
U
Meier
S
S
Sauer
N
N
Schweizer
E
E
D000595:Amino Acid Sequence; D001692:Biological Transport / Q000235:genetics; D001710:Biotin / Q000378:metabolism*; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism*; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D003001:Cloning, Molecular; D005230:Fatty Acids, Nonesterified / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics* / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation; D016366:Open Reading Frames; D012333:RNA, Messenger / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D027981:Symporters
D002352:Carrier Proteins; D005230:Fatty Acids, Nonesterified; D008565:Membrane Proteins; D012333:RNA, Messenger; D027981:Symporters; C119981:biotin transporter; D001710:Biotin