MicroRNAs (miRNAs) are a family of ~22 nucleotide small, non-coding RNAs that negatively regulate gene expression at the transcriptional and post-transcriptional level. miRNAs have been identified in many metazoan including humans, flies, nematodes and plants. Recent researches also reveal that Chlamydomonas reinhardtii, a unicellular green alga, encodes many miRNAs. Thus far, however, no miRNAs have been identified in Giardia lamblia which is considered as one of the most primitive extant eukaryotes. Does any miRNA exist in Giardia, which takes a paramount position in evolution? If yes, what are the characteristics of the miRNAs? What are the differences between the miRNA in Giardia and those in higher eukaryote and Chlamydomonas？Are the miRNA in Giardia related to the inducing of diarrhea? Several recent studies indicate that in Giardia genome, there exist RNAi-related Argonaute (AGO) protein family and Dicer. Interestingly, those key proteins which induce RNAi-AGO and Dicer-are also paramount components in miRNA system, so it implies that miRNA may also exist in Giardia and have functions. Some other researches reveal the existance of huge amount of non-coding transcripts, which are thought to be produced by bi-directional promoters ubiquitously located in Giardia genome. Whether some of the non-coding transcripts are regulational RNA molecules (such as miRNAs) requests further study. In this work, several approaches were combined to make computational prediction of potential miRNAs and their targets in Giardia lamblia. A total of 50 potential miRNAs have been identified following a range of strict filtering criteria. Using these potential miRNA sequences, we have searched their corresponding targets using BLASTN Giardia lamblia coding sequences with their 5’ and 3’ flanking 200bp sequences. Although most of the potential target genes of Giardia miRNAs encode functionally unknown proteins, other annotated target proteins are involved in various physiological functions. A case in point is VSP proteins (various surface proteins), which are a sort of surface antigen proteins. This result implies that Giardia miRNA may be pathogenicity-related. After testing the expression of 12 predicted Giardia miRNAs by RT-PCR and sequencing, we find that gla-mir-6 and gla-mir-35 are steadily expressed in Giardia trophozoite cells. Our study predicts the existance of microRNAs in Giardia by bioinformatics and also confirms the prediction by wet-lab experiments for the first time, which paves the way for studying the functions of those microRNAs in Giardia.
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