Initiator methionine tRNA is essential for Ty1 transposition in yeast.
The yeast retrotransposon Ty1 transposes through an RNA intermediate by a mechanism similar to that of retroviral reverse transcription and integration. Ty1 RNA contains a putative minus strand primer binding site (-PBS) that is complementary to the 3' acceptor stem of the initiator methionine tRNA (tRNA(iMet)). Here we demonstrate that the tRNA(iMet) is used as a primer for Ty1 reverse transcription. Mutations in the Ty1 element that alter 5 of 10 nucleotides that are complementary to the tRNA(iMet) abolish Ty1 transposition, even though they are silent with regard to Ty1 protein coding. We have constructed a yeast strain lacking wild-type tRNA(iMet) that is dependent on a mutant derivative of tRNA(iMet) that has an altered acceptor stem sequence, engineered to restore homology with the Ty1 -PBS mutant. The compensatory mutations made in the tRNA(iMet) alleviate the transposition defect of the Ty1 -PBS mutant. The mutant and wild-type tRNA(iMet) are enriched within Ty1 virus-like particles irrespective of complementarity to the Ty1 -PBS. Thus, complementarity between the Ty1 -PBS and tRNA(iMet) is essential for transposition but is not necessary for packaging of the tRNA inside virus-like particles.
Chapman
K B
KB
Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, MD 21205.
Byström
A S
AS
Boeke
J D
JD
D001483:Base Sequence; D004251:DNA Transposable Elements; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010957:Plasmids; D012331:RNA, Fungal / Q000235:genetics / Q000302:isolation & purification; D012358:RNA, Transfer, Met / Q000235:genetics*; D015183:Restriction Mapping; D012190:Retroviridae / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000502:physiology; D013172:Spores, Fungal / Q000502:physiology
D004251:DNA Transposable Elements; D012331:RNA, Fungal; D012358:RNA, Transfer, Met