Natural Variation of the Amino-Terminal Glutamine-Rich Domain in Drosophila Argonaute2 Is Not Associated with Developmental Defects

Daniel Hain,H.-Arno J Müller,Haruhiko Siomi,Gyorgy Hutvagner,Eric C Lai,Brian R Bettencourt,Michael Welte,Tibor Csorba,Wibke Meyer,Jason Biggerstaff,Katsutomo Okamura,Zhigang Jin

doi:10.1371/journal.pone.0015264

Abstract

The Drosophila argonaute2 (ago2) gene plays a major role in siRNA mediated RNA silencing pathways. Unlike mammalian Argonaute proteins, the Drosophila protein has an unusual amino-terminal domain made up largely of multiple copies of glutamine-rich repeats (GRRs). We report here that the ago2 locus produces an alternative transcript that encodes a putative short isoform without this amino-terminal domain. Several ago2 mutations previously reported to be null alleles only abolish expression of the long, GRR-containing isoform. Analysis of drop out (dop) mutations had previously suggested that variations in GRR copy number result in defects in RNAi and embryonic development. However, we find that dop mutations genetically complement transcript-null alleles of ago2 and that ago2 alleles with variant GRR copy numbers support normal development. In addition, we show that the assembly of the central RNAi machinery, the RISC (RNA induced silencing complex), is unimpaired in embryos when GRR copy number is altered. In fact, we find that GRR copy number is highly variable in natural D. melanogaster populations as well as in laboratory strains. Finally, while many other insects share an extensive, glutamine-rich Ago2 amino-terminal domain, its primary sequence varies drastically between species. Our data indicate that GRR variation does not modulate an essential function of Ago2 and that the amino-terminal domain of Ago2 is subject to rapid evolution.

Highlights

Argonaute proteins play key roles in diverse mechanisms of gene regulation from plants to fungi and animals, including humans
We show that the dop mutation is not allelic to ago2 and that glutamine-rich repeats (GRRs) variation does not cause striking defects in embryonic development
We discovered variants in the GRR pattern and found that they were associated with severe disruption of embryonic development [16]

Summary

Introduction

Argonaute proteins play key roles in diverse mechanisms of gene regulation from plants to fungi and animals, including humans. The presence of multiple, differentially expressed family members in a single species indicates diverse biological functions of Argonaute proteins. Argonaute subfamily proteins interact with siRNAs and/or miRNAs and in animals exert diverse functions during development of various somatic lineages. These functions range from selective degradation of unwanted maternal mRNAs in early development to the regulation of transcripts important for cell fate decisions during neuronal and muscle development as well as degradation of viral RNA in adult organisms [5]. An important function for the RNAi pathway was demonstrated in innate immune response against viruses in animals and plants [6,7,8,9]

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