An RNA secondary structure bias for non-homologous reverse transcriptase-mediated deletions in vivo

TY - JOUR

T1 - An RNA secondary structure bias for non-homologous reverse transcriptase-mediated deletions in vivo

AU - Duch, Mogens R.

AU - Carrasco, Maria L

AU - Jespersen, Thomas

AU - Aagaard, Lars

AU - Pedersen, Finn Skou

AU - Jespersen, Thomas

PY - 2004/1/1

Y1 - 2004/1/1

N2 - Murine leukemia viruses harboring an internal ribosome entry site (IRES)-directed translational cassette are able to replicate, but undergo loss of heterologous sequences upon continued passage. While complete loss of heterologous sequences is favored when these are flanked by a direct repeat, deletion mutants with junction sites within the heterologous cassette may also be retrieved, in particular from vectors without flanking repeats. Such deletion mutants were here used to investigate determinants of reverse transcriptase-mediated non-homologous recombination. Based upon previous structural analysis the individual recombination sites within the IRES could be assigned to either base-paired or unpaired regions of RNA. This assignment showed a significant bias (P = 0.000082) towards recombination within unpaired regions of the IRES. We propose that the events observed in this in vivo system result from template switching during first-strand cDNA synthesis and that the choice of acceptor sites for non-homologous recombination are guided by non-paired regions. Our results may have implications for recombination events taking place within structured regions of retroviral RNA genomes, especially in the absence of longer stretches of sequence similarity.

AB - Murine leukemia viruses harboring an internal ribosome entry site (IRES)-directed translational cassette are able to replicate, but undergo loss of heterologous sequences upon continued passage. While complete loss of heterologous sequences is favored when these are flanked by a direct repeat, deletion mutants with junction sites within the heterologous cassette may also be retrieved, in particular from vectors without flanking repeats. Such deletion mutants were here used to investigate determinants of reverse transcriptase-mediated non-homologous recombination. Based upon previous structural analysis the individual recombination sites within the IRES could be assigned to either base-paired or unpaired regions of RNA. This assignment showed a significant bias (P = 0.000082) towards recombination within unpaired regions of the IRES. We propose that the events observed in this in vivo system result from template switching during first-strand cDNA synthesis and that the choice of acceptor sites for non-homologous recombination are guided by non-paired regions. Our results may have implications for recombination events taking place within structured regions of retroviral RNA genomes, especially in the absence of longer stretches of sequence similarity.

KW - Animals

KW - Base Sequence

KW - Cell Line

KW - Mice

KW - Molecular Sequence Data

KW - Nucleic Acid Conformation

KW - RNA, Viral

KW - RNA-Directed DNA Polymerase

KW - Recombination, Genetic

KW - Regulatory Sequences, Ribonucleic Acid

KW - Retroviridae

KW - Sequence Deletion

KW - Sequence Homology, Nucleic Acid

KW - Virus Replication

U2 - 10.1093/nar/gkh513

DO - 10.1093/nar/gkh513

M3 - Journal article

C2 - 15069126

SN - 0261-3166

VL - 32

SP - 2039

EP - 2048

JO - Nucleic acids symposium series

JF - Nucleic acids symposium series

IS - 6

ER -