Nucleotide Sequence of the Long Terminal Repeat and Flanking Cellular Sequences of Avian Endogenous Retrovirus
            <i>ev</i>
            -2: Variation in Rous-Associated Virus-0 Expression Cannot Be Explained by Differences in Primary Sequence

A strong enhancer element is located within the long terminal repeats (LTRs) of exogenous, oncogenic avian retroviruses, such as Rous sarcoma virus (RSV) and the avian leukosis viruses. The LTRs of a second class of avian retroviruses, the endogenous viruses (evs), lack detectable enhancer function, a property that correlates with major sequence differences between the LTRs of these two virus groups. Despite this lack of independent enhancer activity, we previously identified sequences in ev LTRs that were able to functionally replace essential enhancer domains from the RSV enhancer with which they share limited sequence similarity. To identify candidate enhancer domains in ev LTRs that are functionally equivalent to those in RSV LTRs, we analyzed and compared ev and RSV LTR-specific DNA-protein interactions. Using this approach, we identified two candidate enhancer domains and one deficiency in ev LTRs. One of the proposed ev enhancer domains was identified as a CArG box, a motif also found upstream of several muscle-specific genes, and as the core sequence of the c-fos serum response element. The RSV LTR contains two CArG motifs, one at a previously identified site and one identified in this report at the same relative location as the ev CArG motif. A second factor binding site that interacts with a heat-stable protein was also identified in ev LTRs and, contrary to previous suggestions, appears to be different from previously described exogenous virus enhancer binding proteins. Finally, a deficiency in factor binding was found within the one inverted CCAAT box in ev LTRs, affirming the importance of sequences that flank CCAAT motifs in factor binding and providing a candidate defect in the ev enhancer.

Section: Resultsmentioning

confidence: 74%

mentioning

confidence: 99%

CArG, CCAAT, and CCAAT-like protein binding sites in avian retrovirus long terminal repeat enhancers

Zachow

Conklin

1992

“…A comparison of the nucleotide sequences of avian retroviral LTRs shows that major differences exist between LTRs of exogenous and endogenous viral origin. Endogenous retroviral LTRs are relatively short (-277 base pairs [bp]) and are highly interrelated in sequence (20). The LTRs of exogenous retroviruses such as Rous sarcoma virus (RSV) and avian leukosis virus (ALV) differ significantly from the endogenous viral LTRs but show extensive sequence homology to one another (4).…”

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confidence: 99%

Transcriptional activity of avian retroviral long terminal repeats directly correlates with enhancer activity

Cullen¹,

Raymond²,

Ju³

1985

Retroviral long terminal repeats (LTRs) contain elements responsible for the control of proviral transcription and gene expression. Molecular clones of the LTR region of a number of avian retroviruses have been isolated, and DNA sequence analysis of these clones reveals the existence of a related, but heterogenous, family of LTRs. To examine the functional significance of the observed sequence differences, we have directly tested the abilities of several different avian retrovirus LTRs to act as promoters and enhancers of mRNA transcription. Our results indicate that large differences in LTR transcriptional activity exist and that these differences in gene expression directly correlate with LTR enhancer activity. In particular, we show that the LTR of Fujinami sarcoma virus is intermediate in both transcriptional and enhancer activity when compared with the very active LTRs of the exogenous viruses RAV-2 and Schmidt-Ruppin B and the much less active LTRs of the endogenous virus RAV-0 and its provirus ev-2. These results suggest that LTR enhancer activity may be the primary determinant of avian retroviral LTR transcriptional activity and, hence, oncogenic potential.

“…In addition, these proviruses can be activated in vivo by treating cells which contain them with nucleotide analogs such as bromodeoxyuridine (37) or 5-azacytidine (7,20). Under these conditions, ev-2 gives rise to an infectious virus called Rousassociated virus-0 (RAV-0); the LTRs of ev-2 and RAV-0 are identical (23,41). Previous studies have indicated that the expression of individual evs correlates with the site of provirus integration (7,8,21,25,26,41) and with the pattern of proviral DNA methylation acquired during gametogenesis (19).…”

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confidence: 99%

“…Under these conditions, ev-2 gives rise to an infectious virus called Rousassociated virus-0 (RAV-0); the LTRs of ev-2 and RAV-0 are identical (23,41). Previous studies have indicated that the expression of individual evs correlates with the site of provirus integration (7,8,21,25,26,41) and with the pattern of proviral DNA methylation acquired during gametogenesis (19). However, sequence analysis of the LTR of an ev that normally is expressed in vivo (ev-3) showed minor nucleotide variations relative to the LTRs of ev-1 and ev-2.…”

mentioning

confidence: 99%

Activation of an endogenous retrovirus enhancer by insertion into a heterologous context

Conklin

1991

An enhancer element is located in the U3 portion of exogenous avian retrovirus long terminal repeats (LTRs). A similar element has not been detected in the LTRs of ev-l and ev-2, two avian endogenous viruses (evs) that normally are not expressed in vivo. Experiments were initiated to determine whether minor nucleotide differences in the U3 region of a previously untested ev that is ubiquitously expressed in vivo (ev-3) might confer enhancer function on the LTR of this provirus. This question was addressed by inserting U3 regions from ev-3 and from ev-1 and/or ev-2 both upstream of the herpesvirus thymidine kinase gene promoter and in place of the major enhancer domains of the Rous sarcoma virus LTR and determining their relative effects on transcription. U3 regions from all evs tested were unable to enhance transcription from the thymidine kinase gene promoter, indicating that nucleotide differences in the ev U3 regions do not affect their relative enhancer function and therefore are unlikely to play a role in their differential expression in vivo. Unexpectedly, however, all ev U3 regions were able to augment transcription in an orientation-independent manner in the ev-Rous sarcoma virus hybrid LTRs. Further experiments conducted to determine why this enhancer activity is not detectable in intact ev LTRs demonstrated that it was not due to removal of repressor sequences in the ev fragments used that might normally be present in intact ev LTRs. The lack of detectable enhancer activity in intact ev LTRs also was not explained by a defect in ev promoters that makes them unresponsive to enhancers in cis. These experiments therefore identify sequences that, although unable to function detectably as enhancers in their natural context, can function efficiently in a heterologous context. Data are discussed in terms of the modularity of enhancer elements and possible interactions between enhancers and promoter-specific sequences.