PgmNr W454: The Argonaute VSRA-1 Regulates Gene Expression through Multiple Small RNA Pathways.

Authors:
Julie M. Claycomb 1 ; Monica Z. Wu 1 ; Shikui Tu 2 ; Zhiping Weng 2


Institutes
1) Univ. of Toronto, Toronto, ON, Canada; 2) Univ. of Massachusetts Medical School, Worcester, MA, USA.


Keyword: RNAi and gene silencing

Abstract:

At the core of small RNA pathways are Argonaute (AGO) proteins, which interact with small RNAs to regulate gene expression in a sequence-specific manner. The characterization of AGOs across species, including a subset of the 26 AGOs in C. elegans, has demonstrated that AGOs generally interact with non-overlapping complements of small RNAs to perform distinct regulatory functions. Our lab’s previous studies on the CSR-1 22G-small RNA pathway in C. elegans provided mechanistic insights into a paradigm-shifting role for nuclear small RNA pathways in activating germline gene expression, instead of silencing as occurs in the vast majority of small RNA pathways, like RNAi.

We set out to characterize a well-conserved nematode AGO that was most closely related to CSR-1. Remarkably, we found that this AGO interacts with multiple classes of small RNAs, including the essential mir-51 family of microRNAs (Hs-mir-100), as well as specific subsets of piRNAs and CSR-1 class 22G-RNAs. Owing to this versatile repertoire of small RNA partners, we named this AGO VSRA-1 (Versatile Small RNAs Argonaute).

Loss of vsra-1 leads to slower larval development. Consistent with functions related to multiple small RNA classes, vsra-1 displays synthetic genetic interactions with the AGOs that have previously been shown to act with each type of small RNA (alg-1/miRNAs, prg-1/piRNAs, csr-1/22G-RNAs). Using a combination of CRISPR-Cas9 GFP knock-in strains and antibody staining against endogenous VSRA-1, we determined that VSRA-1 is broadly expressed across most developmental stages and tissues including the germline. In vivo reporter assays and mRNA-seq data implicate VSRA-1 in silencing gene expression, including a large portion of germline-expressed genes. Moreover, sequencing of VSRA-1-small RNA complexes indicates that VSRA-1 associated small RNAs all appear to target CSR-1 pathway genes. Thus, we propose a model in which VSRA-1, through a complex network of small RNAs, functions antagonistically to the CSR-1 pathway to maintain germline homeostasis.

We are currently pursuing structure-function analyses, using CRISPR-Cas9 and biochemical experiments, to dissect the loading and specification of VSRA-1 small RNAs. Due to its unique property of association with multiple classes of small RNAs, characterization of this fascinating AGO will reveal novel mechanisms regulating germline gene expression.



Wormbase Genetic Index
1. C04F12.1
2. VSRA-1
3. CSR-1
4. ALG-1
5. PRG-1