skip to main content

Explore Research Products in the NSF-PAR

  • NSF Public Access
  • Search Results
  • Disparate peroxisome‐related defects in Arabidopsis pex6 and pex26 mutants link peroxisomal retrotranslocation and oil body utilization
Title: Disparate peroxisome‐related defects in Arabidopsis pex6 and pex26 mutants link peroxisomal retrotranslocation and oil body utilization
Summary

Catabolism of fatty acids stored in oil bodies is essential for seed germination and seedling development in Arabidopsis. This fatty acid breakdown occurs in peroxisomes, organelles that sequester oxidative reactions. Import of peroxisomal enzymes is facilitated by peroxins includingPEX5, a receptor that delivers cargo proteins from the cytosol to the peroxisomal matrix. After cargo delivery, a complex of thePEX1 andPEX6ATPases and thePEX26 tail‐anchored membrane protein removes ubiquitinatedPEX5 from the peroxisomal membrane. We identified Arabidopsispex6andpex26mutants by screening for inefficient seedling β‐oxidation phenotypes. The mutants displayed distinct defects in growth, response to a peroxisomally metabolized auxin precursor, and peroxisomal protein import. The lowPEX5 levels in these mutants were increased by treatment with a proteasome inhibitor or by combiningpex26with peroxisome‐associated ubiquitination machinery mutants, suggesting that ubiquitinatedPEX5 is degraded by the proteasome when the function ofPEX6 orPEX26 is reduced. Combiningpex26with mutations that increasePEX5 levels either worsened or improvedpex26physiological and molecular defects, depending on the introduced lesion. Moreover, elevatingPEX5 levels via a35S:PEX5transgene exacerbatedpex26defects and ameliorated the defects of only a subset ofpex6alleles, implying that decreasedPEX5 is not the sole molecular deficiency in these mutants. We found peroxisomes clustered around persisting oil bodies inpex6andpex26seedlings, suggesting a role for peroxisomal retrotranslocation machinery in oil body utilization. The disparate phenotypes of thesepexalleles may reflect unanticipated functions of the peroxisomalATPase complex.

  more » « less
NSF-PAR ID:
10042013
Author(s) / Creator(s):
 ;  ;  ;  ;  ;  ;  
Publisher / Repository:
Wiley-Blackwell
Date Published:
Journal Name:
The Plant Journal
Volume:
92
Issue:
1
ISSN:
0960-7412
Page Range / eLocation ID:
p. 110-128
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
More Like this
  1. ; ; ; ( , Plant Direct)
    Abstract

    The sorting of eukaryotic proteins to various organellar destinations requires receptors that recognize cargo protein targeting signals and facilitate transport into the organelle. One such receptor is the peroxinPEX5, which recruits cytosolic cargo carrying a peroxisome‐targeting signal (PTS) type 1 (PTS1) for delivery into the peroxisomal lumen (matrix). In plants and mammals,PEX5 is also indirectly required for peroxisomal import of proteins carrying aPTS2 signal becausePEX5 binds thePTS2 receptor, bringing the associatedPTS2 cargo to the peroxisome along withPTS1 cargo. DespitePEX5 being thePTS1 cargo receptor, previously identified Arabidopsispex5mutants display either impairment of bothPTS1 andPTS2 import or defects only inPTS2 import. Here, we report the first Arabidopsispex5mutant with an exclusivePTS1 import defect. In addition to markedly diminishedGFP‐PTS1 import and decreased pex5‐2 protein accumulation, thispex5‐2mutant shows typical peroxisome‐related defects, including inefficient β‐oxidation and reduced growth. Growth at reduced or elevated temperatures ameliorated or exacerbatedpex5‐2peroxisome‐related defects, respectively, without markedly changing pex5‐2 protein levels. In contrast to the diminishedPTS1 import,PTS2 processing was only slightly impaired andPTS2‐GFPimport appeared normal inpex5‐2. This finding suggests that even minor peroxisomal localization of thePTS1 proteinDEG15, thePTS2‐processing protease, is sufficient to maintain robustPTS2 processing.

     
    more » « less
  2. ; ; ; ( , The Plant Journal)
    Summary

    Peroxisomes are dynamic organelles crucial for a variety of metabolic processes during the development of eukaryotic organisms, and are functionally linked to other subcellular organelles, such as mitochondria and chloroplasts. Peroxisomal matrix proteins are imported by peroxins (PEX proteins), yet the modulation of peroxin functions is poorly understood. We previously reported that, besides its known function in chloroplast protein import, the Arabidopsis E3 ubiquitin ligase SP1 (suppressor of ppi1 locus1) also targets to peroxisomes and mitochondria, and promotes the destabilization of the peroxisomal receptor–cargo docking complex components PEX13 and PEX14. Here we present evidence that in Arabidopsis, SP1's closest homolog SP1‐like 1 (SPL1) plays an opposite role to SP1 in peroxisomes. In contrast tosp1, loss‐of‐function ofSPL1led to reduced peroxisomal β‐oxidation activity, and enhanced the physiological and growth defects ofpex14andpex13mutants. Transient co‐expression of SPL1 and SP1 promoted each other's destabilization. SPL1 reduced the ability of SP1 to induce PEX13 turnover, and it is the N‐terminus of SP1 and SPL1 that determines whether the protein is able to promote PEX13 turnover. Finally, SPL1 showed prevalent targeting to mitochondria, but rather weak and partial localization to peroxisomes. Our data suggest that these two members of the same E3 protein family utilize distinct mechanisms to modulate peroxisome biogenesis, where SPL1 reduces the function of SP1. Plants and possibly other higher eukaryotes may employ this small family of E3 enzymes to differentially modulate the dynamics of several organelles essential to energy metabolism via the ubiquitin‐proteasome system.

     
    more » « less
  3. ; ; ( , The Plant Journal)
    Summary

    Light signal provides the spatial and temporal information for plants to adapt to the prevailing environmental conditions. Alterations in light quality and quantity can trigger robust changes in global gene expression. InArabidopsis thaliana, two groups of key factors regulating those changes in gene expression areCONSTITUTIVE PHOTOMORPHOGENESIS/DEETIOLATED/FUSCA(COP/DET/FUS) and a subset of basic helix‐loop‐helix transcription factors calledPHYTOCHROME‐INTERACTING FACTORS(PIFs). Recently, rapid progress has been made in characterizing the E3 ubiquitin ligases for the light‐induced degradation ofPIF1,PIF3 andPIF4; however, the E3 ligase(s) forPIF5 remains unknown. Here, we show that theCUL4COP1–SPAcomplex is necessary for the red light‐induced degradation ofPIF5. Furthermore,COP1 andSPAproteins stabilizePIF5 in the dark, but promote the ubiquitination and degradation ofPIF5 in response to red light through the 26S proteasome pathway. Genetic analysis illustrates that overexpression ofPIF5can partially suppress bothcop1‐4andspaQseedling de‐etiolation phenotypes under dark and red‐light conditions. In addition, thePIF5 protein level cycles under both diurnal and constant light conditions, which is also defective in thecop1‐4andspaQbackgrounds. Bothcop1‐4andspaQshow defects in diurnal growth pattern. Overexpression ofPIF5partially restores growth defects incop1‐4andspaQunder diurnal conditions, suggesting that theCOP1–SPAcomplex plays an essential role in photoperiodic hypocotyl growth, partly through regulating thePIF5 level. Taken together, our data illustrate how theCUL4COP1–SPAE3 ligase dynamically controls thePIF5 level to regulate plant development.

     
    more » « less
  4. ; ; ; ; ; ; ( , Plant Direct)
    Abstract

    Photorespiration recovers carbon that would be otherwise lost following the oxygenation reaction of rubisco and production of glycolate. Photorespiration is essential in plants and recycles glycolate into usable metabolic products through reactions spanning the chloroplast, mitochondrion, and peroxisome. Catalase in peroxisomes plays an important role in this process by disproportionating H2O2resulting from glycolate oxidation into O2and water. We hypothesize that catalase in the peroxisome also protects against nonenzymatic decarboxylations between hydrogen peroxide and photorespiratory intermediates (glyoxylate and/or hydroxypyruvate). We test this hypothesis by detailed gas exchange and biochemical analysis ofArabidopsis thalianamutants lacking peroxisomal catalase. Our results strongly support this hypothesis, with catalase mutants showing gas exchange evidence for an increased stoichiometry of CO2release from photorespiration, specifically an increase in the CO2compensation point, a photorespiratory‐dependent decrease in the quantum efficiency of CO2assimilation, increase in the12CO2released in a13CO2background, and an increase in the postillumination CO2burst. Further metabolic evidence suggests this excess CO2release occurred via the nonenzymatic decarboxylation of hydroxypyruvate. Specifically, the catalase mutant showed an accumulation of photorespiratory intermediates during a transient increase in rubisco oxygenation consistent with this hypothesis. Additionally, end products of alternative hypotheses explaining this excess release were similar between wild type and catalase mutants. Furthermore, the calculated rate of hydroxypyruvate decarboxylation in catalase mutant is much higher than that of glyoxylate decarboxylation. This work provides evidence that these nonenzymatic decarboxylation reactions, predominately hydroxypyruvate decarboxylation, can occur in vivo when photorespiratory metabolism is genetically disrupted.

     
    more » « less
  5. ; ; ; ; ( , Physiologia Plantarum)
    Abstract

    Brassinosteroids (BRs) are essential plant growth‐promoting hormones involved in many processes throughout plant development, from seed germination to flowering time. SinceBRsdo not undergo long‐distance transport, cell‐ and tissue‐specific regulation of hormone levels involves both biosynthesis and inactivation. To date, tenBR‐inactivating enzymes, with at least five distinct biochemical activities, have been experimentally identified in the model plantArabidopsis thaliana. Epigenetic interactions betweenT‐DNAinsertion alleles and genetic linkage have hindered analysis of higher‐order null mutants in these genes. A previous study demonstrated that thebas1‐2 sob7‐1 ben1‐1triple‐null mutant could not be characterized due to epigenetic interactions between the exonicT‐DNAinsertions inbas1‐2andsob7‐1,causing the intronicT‐DNAinsertion ofben1‐1to revert to a partial loss‐of‐function allele. We usedCRISPR‐Cas9genome editing to avoid this problem and generated thebas1‐2 sob7‐1 ben1‐3triple‐null mutant. This triple‐null mutant resulted in an additive seedling long‐hypocotyl phenotype. We also uncovered a role forBEN1‐mediatedBR‐inactivation in seedling cotyledon petiole elongation that was not observed in the singleben1‐2null mutant but only in the absence of bothBAS1andSOB7. In addition, genetic analysis demonstrated thatBEN1does not contribute to the early‐flowering phenotype, whichBAS1andSOB7redundantly regulate. Our results show thatBAS1,BEN1,andSOB7have overlapping and independent roles based on their differential spatiotemporal tissue expression patterns

     
    more » « less
  • Citation Formats
  • MLA
  • APA
  • Chicago
  • BibTeX
  • Save / Share this Record
  • Send to Email