Unusual genome organisation in Entamoeba histolytica leads to two overlapping transcripts

Abstract

We have isolated homologs of the mini chromosome maintenance (MCM) gene family from the parasitic protozoan Entamoeba histolytica. The full length genomic and cDNA clones for the Eh MCM3 gene have been characterised. The Eh MCM3 gene is much smaller than the Saccharomyces cerevisiae MCM3 gene and other eukaryotic homologs of the MCM3/P1 family. The predicted Eh Mcm3 protein was 597 amino acids long and showed 37 and 46% positional identity with the Sc Mcm3 and the mouse P1 homologs respectively. While proceeding along the chromosome from the Eh MCM3 gene, we have identified a homolog (Eh PAK) of the murine p²¹ activated kinase (Rn KPAK), or S. cerevisiae STE20. Eh PAK lies 126 bp upstream of the Eh MCM3 gene. The predicted Eh p²¹ activated kinase protein was 459 amino acids long and showed 33% positional identity with the murine p²¹ activated kinase and its yeast homolog Ste20. Analysis of cDNA and genomic sequences shows that the 3' untranslated region (UTR) of the Eh PAK mRNA and the 5' UTR of the Eh MCM3 mRNA are transcribed from a common 40 bp genomic segment. This is the first report of an amoeba gene being physically linked to a second gene such that their transcripts are overlapping and there is no non-transcribed intergenic region between the two genes. Primer extension studies have confirmed that unlike most E. histolytica genes, which have short 5' UTRs, the Eh MCM3 mRNA has a 126 bp long 5' UTR and the Eh PAK mRNA has a 265 bp long 5' UTR.