Relationship between retroviral DNA-integration-site selection and host cell transcription

Abstract

Retroviral DNA integration occurs throughout the genome; however, local ''hot spots'' exist where a strong preference for certain sites over others are seen, and more global preferences associated with genes have been reported. Previous data from our laboratory suggested that there are fewer integration events into a DNA template when it is undergoing active transcription than when it is not. Because these data were generated by using a stably transfected foreign gene that was only weakly inducible, we have extended this observation by comparing integration events into a highly inducible endogenous gene under both induced and uninduced transcriptional states. To examine the influence of transcription on site selection directly, we analyzed the frequency and distribution of integration of avian retrovirus DNA into the metallothionein gene, before and after its induction to a highly sustained level of expression by addition of ZnSO 4. We found a 6-fold reduction in integration events after 100-fold induction of transcription. This result implies that, despite an apparent preference for integration of retroviral DNA into transcribed regions of host DNA, high-level transcription can be inhibitory to the integration process. Several possible models for our observation are as follows. First, when a DNA template is undergoing active transcription, integration might be blocked by the RNA polymerase II complex because of steric hindrance. Alternatively, the integrase complex may require DNA to be in a double-stranded conformation, which would not be the case during active transcription. Last, transcription might lead to remodeling of chromatin into a structure that is less favorable for integration.retrovirus ͉ alpharetrovirus ͉ chromatin ͉ RAV-1 ͉ metallothionein

Integrase, LEDGF/p75 and HIV replication

“…The numerous proteins that have been suspected to influence integration can be grouped into three main categories: those that (i) interact with IN directly; (ii) bind to the viral DNA to influence the process; or (iii) participate in the final gap repair step. Of equivalent interest is deciphering the roles cellular proteins are likely to play in genome-wide patterns of retroviral integration [34][35][36][37][38][39][40][41][42][43][44][45][46]. Experimental capabilities in this area changed with the availability of the human genome sequence, commensurately advancing bioinformatics, and more recently, a second wave of high-throughput sequencing methodologies [47,48]).…”

The ups and downs of gene expression and retroviral DNA integration

“…The experiment was repeated several times, yielding a total of 47 events, 5 of which were duplicate hotspots. Yet during side-by-side comparison, the same number of repetitions yielded only eight integrations in the presence of Zn 2ϩ , leading to an overall 6-fold reduction in integration (2). Because Coffin and coworkers (3) established that an Ϸ5-to 7-fold increase in transcription decreased integration into coding-region DNA by Ϸ25-40%, active transcription has suppressed gene-specific integration over a range of expression levels.…”

“…Although different cloning strategies have indicated an overall preference for transcription units, the mechanism of retroviral integration site selection is for the most part unknown. In this issue of PNAS, Maxfield et al (2) assayed RAV-1 (a subgroup A Alpharetrovirus) integration into the quail metallothionein (MT) gene with and without transcriptional activation. A 100-fold increase in gene expression reduced integration 6-fold.…”

“…Maxfield et al (2) examined the frequency and distribution of RAV-1 integration along the quail MT gene plus͞ minus transcriptional activation. The avian gene was chosen for three main reasons.…”

Crystal structures of catalytic core domain of BIV integrase: implications for the interaction between integrase and target DNA

“…Integration is an obligate step in productive retrovirus infection (Weidhaas et al, 2000;Maxfield et al, 2005). Integration of the retroviral DNA into the host cell genome requires the interaction of the retroviral integrase protein with the outer ends of both viral long-terminal repeats (LTRs), thereby removing two nucleotides from the 3¢ ends (3¢ processing), and the newly created 3¢OH attacking a phosphate of the target DNA (strand transfer) (Engelman et al, 1991;Craigie, 2001;Vandegraaff and Engelman, 2007;Delelis et al, 2008).…”