Gain of Sp1 Sites and Loss of Repressor Sequences Associated with a Young, Transcriptionally Active Subset of HERV-H Endogenous Long Terminal Repeats

Abstract

HERV-H sequences comprise a large family of human endogenous retrovirus-like elements. Previous DNA sequence comparisons of HERV-H long terminal repeats (LTRs) have led to their classification into three subtypes, Types I, Ia, and II. Type Ia appears to have been generated by recombination between Type I and Type II LTRs. These subtypes differ in evolutionary age and transcriptional activity with Type Ia LTRs being younger in evolutionary terms and possessing stronger promoter function than the other two subtypes. In this study, possible mechanisms responsible for the functional difference between LTRs have been explored. Types I and II LTRs each contain different sets of repeated segments in their U3 regions which are disrupted in Type Ia LTRs. Using reporter gene assays, we have shown that both types of repeated segments can suppress activity of the human β-globin gene promoter when cloned at a distant site. Both sets of repeats also repress promoter activity of a Type Ia LTR when directly inserted within its U3 region. In addition, using deletion constructs, we have localized two positive regulatory segments within the Type Ia LTR, both of which contain a potential binding site for the transcription factor Sp1. Gel mobility shift assays demonstrated that fragments containing these sites do bind Sp1. Although Type I LTRs are generally similar to Type Ia LTRs in the regions surrounding the Sp1 sites, there are sequence differences within the sites. Gel-shift analysis revealed no or much reduced Sp1 binding of Type I LTR fragments containing these sites. Thus, it appears that the loss of repeated suppresser elements and the acquisition of Sp1-binding sites have both contributed to the relatively strong transcriptional activity of the Type Ia LTRs.