Transcriptional analysis of three major putative phosphatidylinositol kinase genes in a parasitic protozoan, Giardia lamblia.

The current investigation evaluates the expression of phosphatidylinositol kinase (PIK) genes in the parasitic protozoan, Giardia lamblia. The G. lamblia Genome Database revealed the presence of two putative phosphatidylinositol-3-kinase (gPI3K) and one phosphatidylinositol-4-kinase (gPI4K) genes resembling the catalytic subunit of eukaryotic PIKs. Primers, designed to amplify mRNA of these three genes, were used to measure transcription by quantitative reverse-transcriptase polymerase chain reactions. Results suggest that all three PIK genes are expressed in non-encysting and encysting trophozoites. The relative levels of the mRNA were highest in parasites cultured in pre-encysting medium that contained no bile. Two inhibitors of PI3K, LY 294002 and wortmannin were found to inhibit the growth of the trophozoite in culture. However, wortmannin was more effective than LY294002. Altogether, the present study indicates that Giardia is capable of expressing PIKs that are necessary for the growth and differentiation of this pathogen.

Inferring protein function from genomic sequence: Giardia lamblia expresses a phosphatidylinositol kinase-related kinase similar to yeast and mammalian TOR.

Functional assays of genes have historically led to insights about the activities of a protein or protein cascade. However, the rapid expansion of genomic and proteomic information for a variety of diverse taxa is an alternative and powerful means of predicting function by comparing the enzymes and metabolic pathways used by different organisms. As part of the Giardia lamblia genome sequencing project, we routinely survey the complement of predicted proteins and compare those found in this putatively early diverging eukaryote with those of prokaryotes and more recently evolved eukaryotic lineages. Such comparisons reveal the minimal composition of conserved metabolic pathways, suggest which proteins may have been acquired by lateral transfer, and, by their absence, hint at functions lost in the transition from a free-living to a parasitic lifestyle. Here, we describe the use of bioinformatic approaches to investigate the complement and conservation of proteins in Giardia involved in the regulation of translation. We compare an FK506 binding protein homologue and phosphatidylinositol kinase-related kinase present in Giardia to those found in other eukaryotes for which complete genomic sequence data are available. Our investigation of the Giardia genome suggests that PIK-related kinases are of ancient origin and are highly conserved.