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A small molecule with differential effects on the PTS1 and PTS2 peroxisome matrix import pathways
Author(s) -
Brown LauraAnne,
O’LearySteele Catherine,
Brookes Paul,
Armitage Lynne,
Kepinski Stefan,
Warriner Stuart L.,
Baker Alison
Publication year - 2011
Publication title -
the plant journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.058
H-Index - 269
eISSN - 1365-313X
pISSN - 0960-7412
DOI - 10.1111/j.1365-313x.2010.04473.x
Subject(s) - peroxisome , endoplasmic reticulum , microbiology and biotechnology , auxin , biochemistry , cytosol , peroxisomal targeting signal , organelle , chemistry , green fluorescent protein , brefeldin a , golgi apparatus , biology , receptor , enzyme , gene
Summary The use of small molecules has great power to dissect biological processes. This study presents the identification and characterisation of an inhibitor of peroxisome matrix protein import. A mini‐screen was carried out to identify molecules that cause alteration in peroxisome morphology, or mislocalization of a peroxisome targeted fluorescent reporter protein. A benzimidazole lead compound (LDS‐003655) was identified that resulted in reduced GFP fluorescence in peroxisomes and cytosolic GFP accumulation. The effect of the compound was specific to peroxisomes as Golgi bodies, endoplasmic reticulum and the actin cytoskeleton were unaffected even at 25 μ m , whereas peroxisome import via the PTS1 pathway was compromised at 100 n m . When seedlings were grown on 25 μ m LDS‐003655 they displayed morphology typical of seedlings grown in the presence of auxin, and expression of the auxin reporter DR5::GFP was induced. Analysis of a focussed library of LDS‐003655 derivatives in comparison with known auxins led to the conclusion that the auxin‐like activity of LDS‐003655 is attributable to its in situ hydrolysis giving rise to 2,5‐dichlorobenzoic acid, whereas the import inhibiting activity of LDS‐003655 requires the whole molecule. None of the auxins tested had any effect on peroxisome protein import. Matrix import by the PTS2 import pathway was relatively insensitive to LDS‐003655 and its active analogues, with effects only seen after prolonged incubation on high concentrations. Steady‐state protein levels of PEX5, the PTS1 import pathway receptor, were reduced in the presence of 100 n m LDS‐003655, suggesting a possible mechanism for the import inhibition.