Functional organization of the retrotransposon Ty from Saccharomyces cerevisiae: Ty protease is required for transposition.
We used several mutations generated in vitro to further characterize the functions of the products encoded by the TyB gene of the transpositionally active retrotransposon TyH3 from Saccharomyces cerevisiae. Mutations close to a core protein domain of TyB, which is homologous to retroviral proteases, have striking effects on Ty protein processing, the physiology of Ty viruslike particles, and transposition. The Ty protease is required for processing of both TyA and TyB proteins. Mutations in the protease resulted in the synthesis of morphologically and functionally aberrant Ty viruslike particles. The mutant particles displayed reverse transcriptase activity, but did not synthesize Ty DNA in vitro. Ty RNA was present in the mutant particles, but at very low levels. Transposition of a genetically tagged element ceased when the protease domain was mutated, demonstrating that Ty protease is essential for transposition. One of these mutations also defined a segment of TyB encoding an active reverse transcriptase. These results indicate that the Ty protease, like its retroviral counterpart, plays an important role in particle assembly, replication, and transposition of these elements.
Youngren
S D
SD
Bionetics Research, Inc., National Cancer Institute-Frederick Cancer Research Facility, Maryland 21701.
Boeke
J D
JD
Sanders
N J
NJ
Garfinkel
D J
DJ
D004251:DNA Transposable Elements; D010450:Endopeptidases / Q000235:genetics* / Q000378:metabolism; D005796:Genes; D005800:Genes, Fungal; D005838:Genotype; D008854:Microscopy, Electron; D009154:Mutation; D010957:Plasmids; D012194:RNA-Directed DNA Polymerase / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics* / Q000648:ultrastructure
D004251:DNA Transposable Elements; D012194:RNA-Directed DNA Polymerase; D010450:Endopeptidases