Bioinformatic analysis of Entamoeba histolytica SINE1 elements

Abstract

BackgroundInvasive amoebiasis, caused by infection with the human parasite Entamoeba histolytica remains a major cause of morbidity and mortality in some less-developed countries. Genetically E. histolytica exhibits a number of unusual features including having approximately 20% of its genome comprised of repetitive elements. These include a number of families of SINEs - non-autonomous elements which can, however, move with the help of partner LINEs. In many eukaryotes SINE mobility has had a profound effect on gene expression; in this study we concentrated on one such element - EhSINE1, looking in particular for evidence of recent transposition.ResultsEhSINE1s were detected in the newly reassembled E. histolytica genome by searching with a Hidden Markov Model developed to encapsulate the key features of this element; 393 were detected. Examination of their sequences revealed that some had an internal structure showing one to four 26-27 nt repeats. Members of the different classes differ in a number of ways and in particular those with two internal repeats show the properties expected of fairly recently transposed SINEs - they are the most homogeneous in length and sequence, they have the longest (i.e. the least decayed) target site duplications and are the most likely to show evidence (in a cDNA library) of active transcription. Furthermore we were able to identify 15 EhSINE1s (6 pairs and one triplet) which appeared to be identical or very nearly so but inserted into different sites in the genome; these provide good evidence that if mobility has now ceased it has only done so very recently.ConclusionsOf the many families of repetitive elements present in the genome of E. histolytica we have examined in detail just one - EhSINE1. We have shown that there is evidence for waves of transposition at different points in the past and no evidence that mobility has entirely ceased. There are many aspects of the biology of this parasite which are not understood, in particular why it is pathogenic while the closely related species E. dispar is not, the great genetic diversity found amongst patient isolates and the fact, which may be related, that only a small proportion of those infected develop clinical invasive amoebiasis. Mobile genetic elements, with their ability to alter gene expression may well be important in unravelling these puzzles.

Recombinant SINEs are formed at high frequency during induced retrotransposition in vivo

“…To check this, we did multiple alignment of 63 full-length EhSINE1 copies thought to have retrotransposed most recently 12 , and looked at all positions that were polymorphic in at least 20% of the SINEs. We found 16 such positions.…”

“…Hits showing 100% coverage and maximum similarity were recorded. The coordinates of EhSINEs in various scaffolds are as per previous report 12 . …”

“…Rather they may correspond to genomic SINEs. It is estimated that 142 SINE copies are transcribed in E. histolytica 12 , some of which may be mobilized in our cell line upon tet induction.Sequence comparison showed that seven of the eight sequences in set II indeed showed 98-99% identity with genomic EhSINE1 sequences found in the E. histolytica EST database (3-11 mismatches with best data base hits when compared with the full-length SINE sequence; Table 1, set II). The number of mismatches seen in separate comparisons of 5′-half and 3′-half with the EhSINE1 sequences in the data base is explained later.…”

“…‡ 25 nucleotides were removed from both ends for BLAsT analysis. § Length variation is due to different numbers of internal 26-mer repeats in the EhsInE copies 12 .…”

“…The sequence similarity of 78 bp between EhLInE1 and EhsInE1 at their 3′ ends is shown by a solid black bar. The region marked 'C' (100 bp) in EhsInE1 is conserved in all EhsInEs and may contain the sInE promoter 12 . (b) northern hybridization was performed with total cellular RnA isolated from normally proliferating trophozoites with the hybridization probes as indicated in (a).…”