Fig. 3: Overexpression of PEX39 impairs the import of PTS2-containing proteins.
From: PEX39 facilitates the peroxisomal import of PTS2-containing proteins
a, Immunoblot analysis of HEK293T cells overexpressing GAPDH (negative control) or HsPEX39. The solid and open red arrowheads indicate mature and precursor forms, respectively. b, Quantification of changes in mature and precursor forms using the band intensities of immunoblots prepared per a. The precursor and mature form intensities were normalized by the intensities of the corresponding loading controls. The data represent means ± s.e.m. (n = 3). Statistical significance was determined by unpaired, two-tailed t-test. c,d, [35S]ACAA1 in vitro import assays in the presence of increasing concentrations of H6HsPEX39 (c) or in the presence or absence of H6HsPEX39 and FLAG-PEX7 (d). After incubation, the reactions were treated with trypsin and organelles were isolated by centrifugation and analysed by SDS-PAGE and autoradiography. Protection from trypsin and maturation of ACAA1 reflect import into peroxisomes. Precursor and mature ACAA1 are indicated by open and solid red arrowheads, respectively. I represents input (5% of the reticulocyte lysate containing the [35S]ACAA1 used in each reaction). e, Growth of the indicated yeast strains in oleic acid medium. pPEX39_OE represents a plasmid containing Scpex39 under the control of a TEF2 promoter for overexpression. The data for the wild type, Scpex39Δ and Scpex39Δ + pPEX39 are the same as those shown in Fig. 2a. The data represent means ± s.d. (n = 4). f, Immunoblots of cellular fractions from yeast overexpressing ScPex39. The experiment was performed as described for Fig. 2b using Scpex39Δ cells transformed with a plasmid containing Scpex39 under the control of the endogenous promoter (pPEX39) or an overexpressing TEF2 promoter (pPEX39_OE).
