Both endo-siRNAs and tRNA-derived small RNAs are involved in the differentiation of primitive eukaryote Giardia lamblia

Significance Small RNAs (sRNAs) are most important regulators in eukaryotes. Although different kinds of sRNAs have been extensively studied in higher eukaryotes, their role remains largely unknown in protozoa. We have systematically investigated in the full genome the sRNAs of Giardia lamblia , the most primitive eukaryote known. Surprisingly, we have found that two major types of sRNAs (i.e., endogenous siRNAs and tRNA-derived sRNAs) are largely encoded in the genome of G. lamblia , whereas canonical microRNAs could not be identified in this parasite. Additional studies showed that both sRNAs might be involved in the differentiation regulation of G. lamblia . This study indicates that these two kinds of eukaryotic sRNAs emerged in the early evolution of eukaryotes. Small RNAs (sRNAs), including microRNAs and endogenous siRNAs (endo-siRNAs), regulate most important biologic processes in eukaryotes, such as cell division and differentiation. Although sRNAs have been extensively studied in various eukaryotes, the role of sRNAs in the early emergence of eukaryotes is unclear. To address these questions, we deep sequenced the sRNA transcriptome of four different stages in the differentiation of Giardia lamblia , one of the most primitive eukaryotes. We identified a large number of endo-siRNAs in this fascinating parasitic protozoan and found that they were produced from live telomeric retrotransposons and three genomic regions (i.e., endo-siRNA generating regions [eSGRs]). eSGR-derived endo-siRNAs were proven to target mRNAs in trans . Gradual up-regulation of endo-siRNAs in the differentiation of Giardia suggested that they might be involved in the regulation of this process. This hypothesis was supported by the impairment of the differentiation ability of Giardia when GLDICER, essential for the biogenesis of endo-siRNAs, was knocked down. Endo-siRNAs are not the only sRNA regulators in Giardia differentiation, because a great number of tRNAs-derived sRNAs showed more dramatic expression changes than endo-siRNAs in this process. We totally identified five novel kinds of tRNAs-derived sRNAs and found that the biogenesis in four of them might be correlated with that of stress-induced tRNA-derived RNA (sitRNA), which was discovered in our previous studies. Our studies reveal an unexpected complex panorama of sRNA in G. lamblia and shed light on the origin and functional evolution of eukaryotic sRNAs.