Efficacy of auxin-inducible protein degradation inC. eleganstissues using different auxins and TIR1-expressing strains

Author:

Sharma Nidhi,Marques Filipe,Kratsios PaschalisORCID

Abstract

SummaryThe auxin-inducible degradation system has emerged as a powerful tool to deplete proteins of interest in cells and tissues of various model organisms, includingC. elegans2–5. Here, we present a detailed protocol to perform AID-driven spatiotemporal depletion of specific proteins inC. eleganstissues. First, we introduced the AID degron and a fluorescent reporter at two conserved proteins: (a) the transcription factor CFI-1 (human ARID3), which is expressed in the nucleus of multipleC. elegansneurons and head muscle cells6,7, and (b) the broadly expressed translation initiation factor Y47D3A.21 (human DENR) that localizes in the cytoplasm. Second, we provide a step-by-step guide on how to generateC. elegansstrains suitable for AID-mediated protein (CFI-1 and DENR) depletion. Third, we find that the degree of CFI-1 and DENR depletion inC. eleganstissues is comparable upon treatment with either natural auxin (indole-3-acetic acid (IAA) or a water-soluble synthetic auxin analog (K-NAA). Last, we compare the degree of AID-mediated CFI-1 depletion inC. elegansneurons when the transport inhibitor response 1 (TIR1), component of the SCF ubiquitin ligase complex, is provided in neurons or all somatic cells. Altogether, this protocol provides side-by-side comparisons of different auxins and TIR1-expressing lines. Such comparisons may benefit future studies of AID-mediated protein depletion inC. elegans.Graphical abstractImage provided as pdf (together with Figures).HighlightsEfficient protein depletion inC. eleganstissues upon treatment with either natural or synthetic auxins.Pansomatic TIR1 expression leads to efficient depletion of CFI-1 and DENR.Panneuronal TIR1 expression leads to neuron-specific, yet variable CFI-1 depletion.The AID system is compatible with fluorescence microscopy, Western blotting and behavioral assays.

Publisher

Cold Spring Harbor Laboratory

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