Investigation of transposon-like elements in the genome of Physarum polycephalum
Tp1 is a long highly-abundant repetitive element found in the genome of Physarum polycephalum. It predominates in a hypermethylated, high molecular weight HpaII-resistant domain. The structural properties of this element resemble those of eukaryotic retrotransposons such as copia in Drosophila melanogaster. Preliminary evidence has indicated that segments of Tp1 are transcribed in both plasmodia and amoebae. Hence, from the structural analogy with retrotransposons it was supposed that the long terminal repeats (LTRs) of Tp1 may serve as transcriptional promoters. A plasmid (pE1oriCAT) was constructed which contains a CAT reporter gene located 3' to the Tp1 LTR and a Physarum origin of replication. CAT activity observed in cell lysates prepared from pE1oriCAT-transfected amoebae suggested that the Tp1 LTR can function as a transcriptional promoter. Nucleotide sequence information was obtained for the right-hand end of a copy of Tp1. From its predicted amino acid sequence a high degree of homology was found with the reverse transcriptase domain in the C-terminal half of the pol polypeptide in copia from Drosophila melanogaster, Ty912 from Saccharomyces cerevisiae and Ta1 from Arabidopsis thaliana. The high degree of identity between the reverse transcriptase domains in all of the above elements suggests that they may share a common ancestor. A preliminary search for Tp1 virus-like particles (VLPs) was carried out. Fractions collected from sucrose density gradients, prepared from amoebal cell lysates, were examined for VLP structures by transmission electron microscopy. Although putative VLPs were observed, which were morphologically similar to VLPs associated with the yeast retrotransposon Ty, they did not react immunologically with an antibody prepared against Ty VLPs. A member of another transposon-like family, designated Tp2, has also been identified in the genome of Physarum. Tp2 is 1.68kb in length, has 180bp LTR sequences at its termini and is blanked by 5bp direct repeats. Analysis of the predicted amino acid sequence of Tp2 identified an amino acid motif that is homologous to a highly conserved RNA binding domain in the gag polypeptide of retroviruses and retrotransposons.