Biomedical subjects

F-X Wilhelm

Publications and source records attributed to F-X Wilhelm.

4 recordsLinked to original sources

PubMed2006

Cooperation between reverse transcriptase and integrase during reverse transcription and formation of the preintegrative complex of Ty1.

Reverse transcriptase (RT) and integrase (IN) play a central role in the replication and transposition of retroelements. Increasing evidence suggests that the interaction between these two enzymes is functional and plays an important role in replication. In the yeast Saccharomyces cerevisiae retrotransposon Ty1, the interaction of IN with RT is critical for the formation of an active conformation of RT. We show here that the RT associated with VLPs is active only if it is in close interaction with IN. To probe the IN-RT cis-trans relationship, we have used a complementation assay based on coexpressing two transposons. We show that IN acts in cis to activate RT and that a functional integrase provided in trans is not able to complement replication and transposition defects of IN deletion or IN active-site mutant elements. Our data support a model in which IN not only interacts closely with RT during reverse transcription but also remains associated with RT during the formation of the preintegrative complex.

Binding Sites↗

PubMed2005

Role of integrase in reverse transcription of the Saccharomyces cerevisiae retrotransposon Ty1.

Reverse transcriptase (RT) with its associated RNase H (RH) domain and integrase (IN) are key enzymes encoded by retroviruses and retrotransposons. Several studies have implied a functional role of the interaction between IN and RT during the replication of retroviral and retrotransposon genomes. In this study, IN deletion mutants were used to investigate the role of IN on the RT activity of the yeast Saccharomyces cerevisiae retrotransposon Ty1. We have identified two domains of Ty1 integrase which have effects on RT activity in vivo. The deletion of a domain spanning amino acid residues 233 to 520 of IN increases the exogenous specific activity of RT up to 20-fold, whereas the removal of a region rich in acidic amino acid residues between residues 521 and 607 decreases its activity. The last result complements our observation that an active recombinant RT protein can be obtained if a small acidic tail mimicking the acidic domain of IN is fused to the RT-RH domain. We suggest that interaction between these acidic amino acid residues of IN and a basic region of RT could be critical for the correct folding of RT and for the formation of an active conformation of the enzyme.

Amino Acid Sequence↗

PubMed2005

Reverse transcriptase and integrase of the Saccharomyces cerevisiae Ty1 element.

Integrase (IN) and reverse transcriptase (RT) play a central role in transposition of retroelements. The mechanism of integration by IN and the steps of the replication process mediated by RT are briefly described here. Recently, active recombinant forms of Ty1 IN and RT have been obtained. This has allowed a more detailed understanding of their biochemical and structural properties and has made possible combined in vitro and in vivo analyses of their functions. A focus of this review is to discuss some of the results obtained thus far with these two recombinant proteins and to propose future directions.

Amino Acid Sequence↗

PubMed2003

The central PPT of the yeast retrotransposon Ty1 is not essential for transposition.

The yeast retrotransposon Ty1 has structural and functional similarities to retroviruses. We report here that, as in retroviruses, the plus-strand DNA of Ty1 is synthesized as two segments. A central DNA flap is formed during reverse transcription consecutive to elongation (with strand displacement) of the upstream segment beyond the central polypurine tract (cPPT) until the replication machinery is stopped at the central termination sequence. Comparison of wild-type and cPPT-mutant Ty1 elements shows that the mutant element lacking the central DNA flap is only twofold defective in transposition.

Base Sequence↗