Integration bias of gammaretrovirus vectors following transduction and growth of primary mouse hematopoietic progenitor cells with and without selection.
The recent recognition that recombinant retrovirus vectors can induce oncogenic transformation has stimulated much interest in the pattern of vector integration sites. We report here on the integration pattern of a gammaretrovirus reporter vector following transduction and ex vivo culture of primary mouse bone marrow progenitor cells in the absence and presence of drug selection. Using a novel method of cloning junction fragments, we observed no bias for integrations within genes, but did observe a bias for integrations within gene-dense regions and especially near transcriptional start sites of highly active genes, similar to previous reports in other cell types. We also document a novel bias for integrations within or near a class of genes that encode nuclear-localized proteins. We found that drug selection resulted in an increase in the frequency of recovered integration events that were located within the beginning of genes, integration events that were located in less gene-dense regions, and integration events that were oriented in an antisense direction relative to flanking gene transcription. Taken together, these studies provide new insights into the nature of retrovirus vector integration patterns in primary cells and demonstrate that selection based on vector expression can bias the integration site repertoire.
Aker
Mari
M
Department of Medicine, Division of Medical Genetics, University of Washington, Box 357720, Seattle, 98195, USA.
Tubb
Julie
J
Miller
Daniel G
DG
Stamatoyannopoulos
George
G
Emery
David W
DW
D000818:Animals; D003001:Cloning, Molecular / Q000379:methods; D005260:Female; D017988:Gammaretrovirus / Q000235:genetics*; D005822:Genetic Vectors; D006412:Hematopoietic Stem Cells; D051379:Mice; D009355:Neomycin / Q000494:pharmacology; D024363:Transcription Initiation Site; D014161:Transduction, Genetic; D016662:Virus Integration / Q000187:drug effects
D009355:Neomycin