The canonical pathway for selenocysteine insertion is dispensable in Trypanosomes.
The micronutrient selenium is found in proteins as selenocysteine (Sec), the 21st amino acid cotranslationally inserted in response to a UGA codon. In vitro studies in archaea and mouse showed that Sec-tRNA(Sec) formation is a 3-step process starting with serylation of tRNA(Sec) by seryl-tRNA synthetase (SerRS), phosphorylation of serine to form phosphoserine (Sep)-tRNA(Sec) by phosphoseryl-tRNA(Sec) kinase (PSTK), and conversion to Sec-tRNA(Sec) by Sep-tRNA:Sec-tRNA synthase (SepSecS). However, a complete study of eukaryotic selenoprotein synthesis has been lacking. Here, we present an analysis of Sec-tRNA(Sec) formation in the parasitic protozoon Trypanosoma brucei in vivo. Null mutants of either PSTK or SepSecS abolished selenoprotein synthesis, demonstrating the essentiality of both enzymes for Sec-tRNA(Sec) formation. Growth of the 2 knockout strains was not impaired; thus, unlike mammals, trypanosomes do not require selenoproteins for viability. Analysis of conditional RNAi strains showed that SerRS, selenophosphate synthase, and the Sec-specific elongation factor, EFSec, are also essential for selenoprotein synthesis. These results with T. brucei imply that eukaryotes have a single pathway of Sec-tRNA(Sec) synthesis that requires Sep-tRNA(Sec) as an intermediate.
Aeby
Eric
E
Department of Chemistry and Biochemistry, University of Bern, Freiestrasse 3, CH-3012 Bern, Switzerland.
Palioura
Sotiria
S
Pusnik
Mascha
M
Marazzi
Janine
J
Lieberman
Allyson
A
Ullu
Elisabetta
E
Söll
Dieter
D
Schneider
André
A
D000818:Animals; D001105:Archaea; D053858:Metabolic Networks and Pathways; D051379:Mice; D010770:Phosphotransferases / Q000378:metabolism; D012345:RNA, Transfer, Amino Acid-Specific / Q000378:metabolism; D012346:RNA, Transfer, Amino Acyl / Q000378:metabolism; D017279:Selenocysteine / Q000378:metabolism*; D051140:Selenoproteins / Q000096:biosynthesis*; D014346:Trypanosoma brucei brucei / Q000201:enzymology / Q000378:metabolism*
D012345:RNA, Transfer, Amino Acid-Specific; D012346:RNA, Transfer, Amino Acyl; D051140:Selenoproteins; C112705:selenocysteinyl-tRNA; C036928:tRNA, selenocysteine-; D017279:Selenocysteine; D010770:Phosphotransferases