A Trinucleotide Repeat-Associated Increase in the Level of Alu RNA-Binding Protein Occurred during the Same Period as the Major Alu Amplification That Accompanied Anthropoid Evolution

Abstract

Nearly 1 million Alu elements in human DNA were inserted by an RNA-mediated retroposition-amplification process that clearly decelerated about 30 million years ago. Since then, Alu sequences have proliferated at a lower rate, including within the human genome, in which Alu mobility continues to generate genetic variability. Initially derived from 7SL RNA of the signal recognition particle (SRP), Alu became a dominant retroposon while retaining secondary structures found in 7SL RNA. We previously identified a human Alu RNA-binding protein as a homolog of the 14-kDa Alu-specific protein of SRP and have shown that its expression is associated with accumulation of 3-processed Alu RNA. Here, we show that in early anthropoids, the gene encoding SRP14 Alu RNA-binding protein was duplicated and that SRP14-homologous sequences currently reside on different human chromosomes. In anthropoids, the active SRP14 gene acquired a GCA trinucleotide repeat in its 3-coding region that produces SRP14 polypeptides with extended C-terminal tails. A C3G substitution in this region converted the mouse sequence CCA GCA to GCA GCA in prosimians, which presumably predisposed this locus to GCA expansion in anthropoids and provides a model for other triplet expansions. Moreover, the presence of the trinucleotide repeat in SRP14 DNA and the corresponding C-terminal tail in SRP14 are associated with a significant increase in SRP14 polypeptide and Alu RNA-binding activity. These genetic events occurred during the period in which an acceleration in Alu retroposition was followed by a sharp deceleration, suggesting that Alu repeats coevolved with C-terminal variants of SRP14 in higher primates.Ample evidence indicates that amplification of short interspersed elements (SINEs) occurred via reverse transcription of small RNA intermediaries in a process termed retroposition (52). The SINEs of many organisms are related to one or another tRNA, and these have been found widely distributed in nature, including in plants, fish, and mammals (see references 13, 43, and 44 and references therein). The Alu family of SINEs was derived from the terminal portions of the 7SL RNA component of the signal recognition particle (SRP), a small ribonucleoprotein involved in intracellular protein trafficking (6,24,27,70,71,74,77). In contrast to tRNA-like SINEs, and despite the ubiquity of 7SL SRP RNAs, Alu-related SINEs are for unknown reasons mostly limited to rodents and primates (for a recent review, see reference 13).Evolution of Alu-related SINEs apparently occurred in two major phases: an ancient period during which Alu sequences emerged as monomeric elements followed by a period of remodelling and proliferation (see reference 50 and references therein). The discovery of fossil Alu sequences provided evidence that an Alu monomer originated in an ancestor common to rodents and primates (50). The monomeric Alu-equivalent SINE referred to as B1 has since undergone substantial divergence from 7SL during its amplification to ϳ50,000 to 80,000 copies per haploid genom...

The Decline in Human Alu Retroposition Was Accompanied by an Asymmetric Decrease in SRP9/14 Binding to Dimeric Alu RNA and Increased Expression of Small Cytoplasmic Alu RNA

“…This deregulation is associated with genetic polymorphisms in SRP14 and occurred during the primate evolutionary period that witnessed the greatest burst of Alu amplification known (11). Moreover, coincidences between structural changes in SRP9/14 and calculated rates of Alu amplification strengthen the contention that Alu amplification was modulated by adaptations in SRP9/14 (11,32). As discussed above, the Alu sequence itself changed during primate evolution, and although this might have affected its interaction with SRP9/14, a way to test this had been lacking.…”

“…Alu monomer electrophoretic mobility shift assays (EMSAs) were performed with SRP9/14 that was purified from HeLa cells (heparin agarose fraction) (10). Alu dimer EMSAs were performed with cytoplasmic extract of owl monkey cells as described previously (11,19). RNAs were synthesized in parallel reactions using a common pool of nucleoside triphosphates and [␣- 32 P]GTP to ensure [ 32 P]RNA products of the same specific activity.…”

“…Conservation of the Alu cruciform structure by flAlu consensuses suggests that interaction with SRP9/14 has had a positive effect on Alu retroposition, presumably at the level of RNA stability. A role for SRP9/14 in Alu mobility is also suggested by the fact that in humans and modern primates SRP9/14 levels are 10-to 20-fold higher than 7SL RNA levels (5,11). This deregulation is associated with genetic polymorphisms in SRP14 and occurred during the primate evolutionary period that witnessed the greatest burst of Alu amplification known (11).…”

“…A role for SRP9/14 in Alu mobility is also suggested by the fact that in humans and modern primates SRP9/14 levels are 10-to 20-fold higher than 7SL RNA levels (5,11). This deregulation is associated with genetic polymorphisms in SRP14 and occurred during the primate evolutionary period that witnessed the greatest burst of Alu amplification known (11). Moreover, coincidences between structural changes in SRP9/14 and calculated rates of Alu amplification strengthen the contention that Alu amplification was modulated by adaptations in SRP9/14 (11,32).…”

“…Association between a relatively large flAlu [ . To more convincingly demonstrate direct binding we used owl monkey cell extract as a source of SRP9/14, which, being larger, produces a more retarded RNP complex (11 (10). Reconstitution products were electrophoresed on 6% native polyacrylamide gels, dried, and exposed to X-ray film.…”