LINEs, SINEs and other retroelements: do birds of a feather flock together?

Abstract

Mobile elements account for almost half of the mass of the human genome. Only the retroelements from the non-LTR (long terminal repeat) retrotransposon family, which include the LINE-1 (L1) and its non-autonomous partners, are currently active and contributing to new insertions. Although these elements seem to share the same basic amplification mechanism, the activity and success of the different types of retroelements varies. For example, Alu-induced mutagenesis is responsible for the majority of the documented instances of human disease induced by insertion of retroelements. Using copy number in mammals as an indicator, some SINEs have been vastly more successful than other retroelements, such as the retropseudogenes and even L1, likely due to differences in post-insertion selection and ability to overcome cellular controls. SINE and LINE integration can be differentially influenced by cellular factors, indicating some differences between in their amplification mechanisms. We focus on the known aspects of this group of retroelements and highlight their similarities and differences that may significantly influence their biological impact.

Figure 1

Schematic representation of the structural organization of active human retroelements and their transcripts. Alu, SVA and LINE-1 belong to the non-LTR group of currently active human retroelements. Each panel shows the schematic of the element with the expected transcript (represented as a line and labeled RNA) shown below it. The boundaries of a retroelement are defined by the presence of tandem site duplications (TSDs) (shown as black arrowheads) generated during the retrotransposition process and which are a hallmark of retrotransposed sequences. Ns represent the flanking genomic sequence. The transcription start sites are represented by a green arrow. LINE-1 (L1) elements are composed of a 5’UTR that contains an internal RNA polymerase II promoter, followed by two open reading frames (ORF1 and ORF2) and a 3’UTR containing a polyadenylation signal (pA) and an A-tail. Full-length L1 transcripts are polyadenylated and capped (represented as a brown hexagon). Alu elements are about 300 bp in length (shown at the top of the panel, with a larger version that includes structural details shown below). Alu elements are composed of two non-identical sequences (monomers shown in yellow) separated by an adenine-rich region and flanked at the 3’ end by a longer A-rich region commonly referred to as an “A-tail”. The Alu left monomer contains the internal bipartite RNA pol III promoter composed of an A and B box (shown in blue). Alu transcripts will usually contain the 3’ flanking unique genomic sequence (shown in gray) past the A-tail due to the lack of a RNA pol III terminator sequence (TTTT) within the Alu sequence. Alu transcripts are thought to have a cap-like structure composed of a gamma-monomethyl phosphate (represented as P-P-P) that has been shown to be present in other pol IIII transcripts (135, 136). SVA elements are composite elements that contain a 5’ region with a variable number of CCCTCT repeats, an antisense Alu-like sequence, a variable number tandem repeat (VNTR) region, and a HERVK-like region previously referred to as SINE-R. A promoter has yet to be identified (represented as “?”), but it is presumed that RNA polymerase II likely drives transcription to generate a capped (brown hexagon) and polyadenylated transcript. Retropseudogenes or processed pseudogenes can originate from the retrotransposition of either RNA pol II or pol III transcripts. Retropseudogenes also are flanked by the hallmark TSD. Although the retrotransposed copies are not expected to contain a promoter sequence with them when they mobilize, sometimes the new flanking genomic sequence into which they insert provides promoter function, and thus some retropseudogenes generate transcripts.

Figure 2

RNA profiles of L1 and Alu elements. Schematic representations of northern blot analysis of L1 (A) and Alu (B) are shown. L1 transcripts are extensively processed both by premature polyadenylation and splicing (42, 44). An L1 RNA profile may show full-length (FL) transcripts (indicated by a black arrow), in addition to a variety of processed products depending on the human tissue analyzed (45). Alu RNA profiles will vary depending on the probe utilized. The structure of a typical Alu transcript is shown at the bottom. Probes (shown as blue boxes marked with asterisks *) that hybridize with the Alu body sequence will detect both large (over 1 kb) and small (100–600 bp) RNA products (schematic shown on the left). The large products observed include RNA pol II transcripts that contain Alu sequences, while the smaller products observed include the RNA pol III transcribed Alu RNAs of various lengths, the left monomer, a degradation intermediate known as small cytoplasmic Alu (scAlu (54), indicated by an open arrowhead) and the 7SL transcript (56), indicated by a dotted arrow) due to shared sequence homology. The use of a probe that hybridizes to the unique sequence (derived from the genomic flank) will only detect those RNA pol III Alu transcripts generated from that specific locus (schematic shown on the right).

Figure 3

Proximity as a requirement for efficient retrotransposition. A. Proposed model explaining the reduced efficiency of retrotransposition of non-L1 mRNAs (retropseudogenes generation). Schematic representation of a full-length capped L1 transcript in a polyribosomal complex undergoing translation. As the L1 ORF1 protein is translated, the ORF1 monomers interact forming a trimer. Due to the cis-preference shown by L1 it is expected that the ORF1 trimer will bind the L1 RNA that generated it, likely driven by proximity. Although ORF2p is made in lower amounts, it also is thought to preferentially interact with the same L1 transcript. The increased number of ORF1p likely coats the L1 RNA, possibly preventing the reassembly of the ribosome and contributing to the “escape” from the translational machinery, allowing for the newly formed L1 RNP to progress through the retrotransposition cycle. In contrast, because mRNA from cellular genes will be undergoing translation in their own polyribosomes, these transcripts will be physically restricted from coming in contact with the L1 proteins. The lack of the ORF1p interaction with the mRNA will enable ribosomes to continue to assemble and continue translation, until the mRNA is targeted to the degradation pathway. [The CAP complex is represented as a brown circle at the 5’ end of the transcript. Ribosomes are shown in green, and the L1 ORF1 and ORF2 proteins are shown in purple and orange respectively.] B. Proposed model for Alu sequestration of ORF2 protein. Due to the lack of coding sequences, Alu transcripts are not engaged by the translation machinery. Thus the Alu transcript can exist as a “free” ribonucleoprotein (RNP) complex (see inset). The inset shows a liberal representation of the potential folding of the two 7SL derived monomers to form the propeller structure and the interaction with the SRP9 and SRP14 proteins (depicted in blue): only SRP9p is shown. In addition to SRP9/14p, the Poly-A binding protein (PABP, depicted in purple) is thought to interact with the A-tail of the Alu RNA. PABP is shown interacting with the 3’ A-tail of the Alu RNA. The structure is likely to fold to form a compact RNP. The bound SRP14p and PABP may play a role in targeting Alu RNPs to the ribosomes. SRP14p interacts with ribosome through direct contacts with amino acids located in its carboxy-terminus. The PABP bound to the Alu A-tail may also help direct the Alu RNP to the translating L1 transcript by interacting with the cap complex (brown circle) of the L1 RNA. These interactions (indicated by short black arrows) may favor the localization of Alu RNPs to translating ribosomes to increase their probability of coming in close proximity to newly synthesized ORF2p (in orange). For illustrative purposes, the L1 transcript bound to ribosome is shown as a linear molecule. However, the cap complex of the L1 RNA likely interacts with the PABP present in its own A-tail circularizing the RNA.

Figure 4

Target primed reverse transcription (TPRT). A schematic representation of the TPRT step of the retrotransposition process is shown for L1, Alu, and tailless tRNA-derived transcripts. The genomic DNA is depicted as dark lines. The consensus 5’-TTAAAA-3’ is shown on the top strand of the insertion sites for the L1 and Alu, while a representative sequence of the 5’-TTWX1-17-3’ consensus is shown for the tailless retropseudogene. The bottom strand shows the nicked DNA with the exposed nucleotides (e.g. TTTT) thought to anneal with the RNA and provide the priming site for reverse transcription by ORF2p (depicted in orange). The Alu and tailless retropseudogene transcripts are shown with extended 3’ sequences (shown as Ns) which represent the unique region of Alu or part of the tRNA-related sequences followed by the RNA pol III terminator (shown as Us).