E3 ubiquitin ligase SP1 regulates peroxisome biogenesis in Arabidopsis
Friday, 2016/11/18 | 07:40:00
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Ronghui Pan, John Satkovich, and Jianping Hu SignificancePeroxisomes are eukaryotic organelles crucial for development. Peroxisomal matrix proteins are imported by the peroxisome import machinery composed of peroxins (PEX proteins), but how the function of these PEX proteins is regulated is largely unknown. We discovered in Arabidopsis that the ubiquitin–proteasome system regulates peroxisome protein import via an E3 ubiquitin ligase, SP1 (suppressor of ppi1 locus1), which targets PEX13 and possibly several other components of the peroxisome matrix protein import machinery for degradation. Our data demonstrate that the same E3 ubiquitin ligase can be shared by metabolically linked peroxisomes and chloroplasts to promote the destabilization of distinct components of the two import machineries, suggesting that the ubiquitin–proteasome system may represent an important regulatory mechanism coordinating the biogenesis of functionally associated organelles. AbstractPeroxisomes are ubiquitous eukaryotic organelles that play pivotal roles in a suite of metabolic processes and often act coordinately with other organelles, such as chloroplasts and mitochondria. Peroxisomes import proteins to the peroxisome matrix by peroxins (PEX proteins), but how the function of the PEX proteins is regulated is poorly understood. In this study, we identified the Arabidopsis RING (really interesting new gene) type E3 ubiquitin ligase SP1 [suppressor of plastid protein import locus 1 (ppi1) 1] as a peroxisome membrane protein with a regulatory role in peroxisome protein import. SP1 interacts physically with the two components of the peroxisome protein docking complex PEX13–PEX14 and the (RING)-finger peroxin PEX2. Loss of SP1 function suppresses defects of the pex14-2 and pex13-1 mutants, and SP1 is involved in the degradation of PEX13 and possibly PEX14 and all three RING peroxins. An in vivo ubiquitination assay showed that SP1 has the ability to promote PEX13 ubiquitination. Our study has revealed that, in addition to its previously reported function in chloroplast biogenesis, SP1 plays a role in peroxisome biogenesis. The same E3 ubiquitin ligase promotes the destabilization of components of two distinct protein-import machineries, indicating that degradation of organelle biogenesis factors by the ubiquitin–proteasome system may constitute an important regulatory mechanism in coordinating the biogenesis of metabolically linked organelles in eukaryotes.
See http://www.pnas.org/content/113/46/E7307.abstract.html?etoc PNAS 15 November 2016; vol.113; no.46: E7307–E7316
Fig. 1. Localization of the SP1 protein. (A) Domain structure of the SP1 protein. TM, transmembrane domain. RING, RING domain. C-term, C terminus. Amino acid numbers are indicated. T-DNA insertion sites of in the two sp1 mutants are also indicated. (B) Peroxisome localization of SP1-YFP. Confocal images were taken in leaf epidermal cells of 2-wk-old Arabidopsis T2 plants coexpressing SP1-YFP and CFP-PTS1. (Scale bar, 10 μm.) (C) Assessment of the purity of peroxisomes isolated from the leaf tissue of transgenic Arabidopsis plants coexpressing SP1-YFP and CFP-PTS1. Organelle-specific antibodies used were against Arabidopsis VDAC (Voltage-dependent anion channel) (mitochondrial), FtsZ1 (Filamenting temperature-sensitive mutant Z) (chloroplast), and PEX11d (peroxisomal) proteins. CE, crude extract. (D) Peroxisomal membrane association of SP1-YFP. Purified peroxisomes were treated with TE, NaCl, or Na2CO3 (pH 11.0) and were separated into soluble (S) and pellet (P) fractions by centrifugation. SP1-YFP and CFP-PTS1 were detected by α-GFP. CFP-PTS1 and PEX11d are controls for matrix and integral membrane proteins, respectively. The matrix protein CFP-PTS1 is often detected in the pellet as well as in the soluble fraction, as has been observed in previous studies. |
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