Analysis of cDNA expressed sequence tags from Entamoeba histolytica: Identification of two highly abundant polyadenylated transcripts with no overt open reading frames

Upon analysis of 304 expressed sequence tags derived from the protozoan par asite Entamoeba histolytica, a number of novel protein encoding amoeba sequ ences were isolated. In addition, two unrelated, abundantly expressed trans cripts were identified, and designated, ehapt1 and ehapt2. Although these t ranscripts do not contain any overt open reading frame, both are polyadenyl ated and together represent about 19% of total polyA(+)-RNA(11.6% for ehapt 1 and 7.5% for ehapt2), thus being the most highly expressed polya-containi ng transcripts so far identified in E. histolytica trophozoites. Northern b lot and primer extension analyses revealed single-sized transcripts of 0.5 and 0.6 kb for ehapt1 and ehapt2, respectively, and Southern blot analysis suggests that both are encoded by multiple genes, which are distributed thr oughout the amoeba genome. Comparison between various ehapt1- and ehapt2-de rived cDNAs indicated that both transcripts are highly polymorphic. Whereas nucleotide substitutions in ehapt2 are distributed throughout the sequence , variations in ehapt1 are mainly restricted to two regions, one of which c omprises a deletion of variable length within an 8 nt tandem repeat unit. A t present there is no convincing explanation for the possible role of ehapt 1 and ehapt2 in E. histolytica, and analogous sequences have not been descr ibed so far for any other organism. Most likely they might represent regula tory RNAs or transcribed transposable elements.