Identification of a novel selD homolog from eukaryotes, bacteria, and archaea: is there an autoregulatory mechanism in selenocysteine metabolism?
Escherichia coli selenophosphate synthetase (SPS, the selD gene product) catalyzes the production of monoselenophosphate, the selenium donor compound required for synthesis of selenocysteine (Sec) and seleno-tRNAs. We report the molecular cloning of human and mouse homologs of the selD gene, designated Sps2, which contains an in-frame TGA codon at a site corresponding to the enzyme's putative active site. These sequences allow the identification of selD gene homologs in the genomes of the bacterium Haemophilus influenzae and the archaeon Methanococcus jannaschii, which had been previously misinterpreted due to their in-frame TGA codon. Sps2 mRNA levels are elevated in organs previously implicated in the synthesis of selenoproteins and in active sites of blood cell development. In addition, we show that Sps2 mRNA is up-regulated upon activation of T lymphocytes and have mapped the Sps2 gene to mouse chromosome 7. Using the mouse gene isolated from the hematopoietic cell line FDCPmixA4, we devised a construct for protein expression that results in the insertion of a FLAG tag sequence at the N terminus of the SPS2 protein. This strategy allowed us to document the readthrough of the in-frame TGA codon and the incorporation of 75Se into SPS2. These results suggest the existence of an autoregulatory mechanism involving the incorporation of Sec into SPS2 that might be relevant to blood cell biology. This mechanism is likely to have been present in ancient life forms and conserved in a variety of living organisms from all domains of life.
Guimarães
M J
MJ
Department of Molecular Biology, DNAX Research Institute of Molecular and Cellular Biology, Palo Alto, CA 94304, USA.
Peterson
D
D
Vicari
A
A
Cocks
B G
BG
Copeland
N G
NG
Gilbert
D J
DJ
Jenkins
N A
NA
Ferrick
D A
DA
Kastelein
R A
RA
Bazan
J F
JF
Zlotnik
A
A
D000595:Amino Acid Sequence; D000818:Animals; D001105:Archaea / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D001483:Base Sequence; D019556:COS Cells; D002874:Chromosome Mapping; D003001:Cloning, Molecular; D029721:Drosophila Proteins; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D005260:Female; D005819:Genetic Markers; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D010770:Phosphotransferases; D010802:Phylogeny; D016133:Polymerase Chain Reaction; D012643:Selenium / Q000378:metabolism; D017279:Selenocysteine / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D014162:Transfection
D001426:Bacterial Proteins; D029721:Drosophila Proteins; D005819:Genetic Markers; D017279:Selenocysteine; D010770:Phosphotransferases; C062762:selenophosphate synthetase; D012643:Selenium