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Enhanced neuronal RNAi in C. elegans using SID-1

Nature Methods volume 7, pages 554–559 (2010)Cite this article

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Abstract

We expressed SID-1, a transmembrane protein from Caenorhabditis elegans that is required for systemic RNA interference (RNAi), in C. elegans neurons. This expression increased the response of neurons to double-stranded (ds)RNA delivered by feeding. Mutations in the lin-15b and lin-35 genes enhanced this effect. Worms expressing neuronal SID-1 showed RNAi phenotypes when fed with bacteria expressing dsRNA for known neuronal genes and for uncharacterized genes with no previously known neuronal phenotypes. Neuronal expression of sid-1 decreased nonneuronal RNAi, suggesting that neurons expressing transgenic sid-1(+) served as a sink for dsRNA. This effect, or a sid-1(–) background, can be used to uncover neuronal defects for lethal genes. Expression of sid-1(+) from cell-specific promoters in sid-1 mutants results in cell-specific feeding RNAi. We used these strains to identify a role for integrin signaling genes in mechanosensation.

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Figure 1: Expression of sid-1 in neurons enhances neuronal RNAi.
Figure 2: Mutations in lin-35 and lin-15b enhance RNAi in neurons expressing sid-1.
Figure 3: Enhanced RNAi for genes needed for touch sensitivity.
Figure 4: Expression of sid-1 in neurons decreases RNAi responses in nonneuronal tissues.
Figure 5: Eliminating integrin signaling proteins by RNAi in neurons.

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Acknowledgements

We thank A. Grishok for helpful discussions, J. Kratz for generating the Psng-1yfp plasmid, S. Karimzadegan for generating the Punc-4mdm2∷gfp; Punc-119sid-1 strain. Some C. elegans strains used in this work were provided by the Caenorhabditis Genetics Center, which is funded by the US National Institutes of Health National Center for Research Resources, the C. elegans Gene Knockout Consortium and the National Bioresource Project of Japan. I.T. was supported by an EMBO Long Term fellowship (ALTF 298-2004) and a Human Frontier Science Program Long Term fellowship (LT00776/2005-l/1). This work was supported by US National Institutes of Health grant GM30997 to M.C.

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Author notes

  1. Andrea Calixto & Dattananda Chelur

    Present address: Present addresses: Centro de Envejecimiento y Regeneración, Centro de Regulación Celular y Patología Joaquin V. Luco, Instituto Milenio de Biología Fundamentaly Aplicada, Facultad de Ciencias Biológicas, P. Universidad Católica de Chile, Santiago, Chile (A.C.) and LifeSensors, Inc., Malvern, Pennsylvania, USA (D.C.).,

  2. Andrea Calixto, Dattananda Chelur and Irini Topalidou: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Biological Sciences, Columbia University, New York, New York, USA

    Andrea Calixto, Dattananda Chelur, Irini Topalidou, Xiaoyin Chen & Martin Chalfie

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  1. Andrea Calixto
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Contributions

All authors designed experiments; D.C. generated the initial constructs and constructed the multiply mutant lines; X.C. generated the cell-specific constructs and constructed strains with the sid-1 mutation; A.C., D.C., I.T. and X.C. conducted the RNAi tests; and A.C., I.T. and M.C. wrote the paper.

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Correspondence to Martin Chalfie.

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Calixto, A., Chelur, D., Topalidou, I. et al. Enhanced neuronal RNAi in C. elegans using SID-1. Nat Methods 7, 554–559 (2010). https://doi.org/10.1038/nmeth.1463

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