User:R2element

R2 elements are ancient molecular parasites found throughout arthropods. The R2 element is a type of retrotransposon: a transposon whose reproduction proceeds through an RNA intermediate (similar to retroviruses, e.g. HIV). Three regions of conserved secondary structure appear in the R2 RNA; secondary structure is known or suspected to be important in several aspect of R2 biology: RNA processing, genome incorporation (retrotransposition), and translation.

RNA Processing: The R2 Ribozyme


The R2 element is co-transcribed with host organism 28S ribosomal RNA (rRNA). To become a fully mature R2 messenger RNA (mRNA), requires that the initial R2 transcript be processed to remove the 28S rRNA. This occurs by a self cleaving ribozyme (RNA enzyme) that occurs close to the R2/rRNA junction site. The R2 ribozyme has marked structural and sequence correspondence to the Hepatitis Delta Virus (HDV) ribozyme. (REF) The mature R2 mRNA contains a single open reading frame (ORF) for the multi-functional R2 protein, and two untranslated regions(UTRs) at the 5' and 3' ends.

Genome Integration: The 5' and 3' R2 RNA structured regions
R2 elements reproduce by site specific integration into host genome 28S rRNA genes. Integration proceeds via sequence specidfic DNA strand cleavage followed by DNA synthesis mediated by a RNA-protein complex. The complex is made up of two R2 proteins, bound to an R2 mRNA molecule at the structured 5' and 3' regions,. The mRNA-protein complex recognizes the insertion site in the 28S rRNA gene. The 3′ bound R2 protein nicks one DNA strand providing a 3′-OH group to prime reverse-transcription of the R2 complementary DNA(cDNA). The 5' bound R2 protein then cleaves the other DNA strand and acts as a DNA templated DNA polymerase, using the R2 cDNA as template. The 3′ R2 protein binding site occurs within the mRNA UTR and has a conserved secondary structure determined from thermodynmamic energy minimization, sequence comparison and structure probing with chemical reagents. Secondary structure conservation occurs within silk moths, drosophila and between the two groups.

The 5′ R2 protein binding site (in Bombyx mori) occurs in a region that spans part of the 5' UTR and the start of the R2 ORF. These region also has a conserved secondary structure, which has been deduced from binding to oligonucleotide microarrays, structure probing, and free energy minimization. To date, conservation of structure has only been described between silk moth species.

5' R2 Pseudoknot is Conserved Across 5 Silk Moth Species
Secondary structure is important for biological function of some non-coding RNAs (ncRNA). A 5' psuedoknot structure was found in 5 different moth species; Bombyx mori (R2Bm), Samia cynthia (R2Sc), Coscinocera hercules (R2Ch), Callosamia promethea (R2Cpr), Saturnia pyri (R2Spy). The psuedoknot is highly conserved between the 5 silk moth species. Sequence comparisons show evidence for compensatory mutations within the helical regions indicating the secondary structure of the RNA is of biological importance. The single stranded regions, on the other hand, possess less evolutionary constraints shown by the high frequency of base substitutions, thereby hinting how uninportant the sequence is of the single strand regions to the biological function of the psuedoknot.

R2 Translation (putative): The R2 Pseudoknot
Within the 5' structure of B. mori, 74 nucleotides fold into a complex RNA pseudoknot, which is supported by NMR spectra and sequence comparison. Pseudoknots are unusual structural elements and they often play important roles in biological processes. Alignments of R2 RNA and predicted R2 ORFs suggests a transition from functions relating solely to RNA secondary structure to protein coding potential only. Alignments also suggest the possibility of an unusual mode of translation initiation: as the R2 transcript has no 5' cap structure, has multiple in-frame stop codons in its 5' region, and from the observation that conservation of the protein coding sequence only occurs after the conserved pseudoknot.

It is possible that this structure, alone or with other parts of the 5' conserved region, may be able to recruit the ribosome in a process similar to internal ribosomal entry sites (IRES). Pseudoknots have been previously reported to be involved viral IRES domains.