Shot-Gun Proteomic Analysis on Roots of Arabidopsis pldα1 Mutants Suggesting New Roles of PLDα1 in Mitochondrial Protein Import, Vesicular Trafficking and Glucosinolate Biosynthesis

Phospholipase Dα1 (PLDα1) belongs to phospholipases, a large phospholipid hydrolyzing protein family. PLDα1 has a substrate preference for phosphatidylcholine leading to enzymatic production of phosphatidic acid, a lipid second messenger with multiple cellular functions. PLDα1 itself is implicated in biotic and abiotic stress responses. We present here a shot-gun differential proteomic analysis on roots of two pldα1 mutants compared to the Col-0 wild type. Our data suggest new roles of PLDα1 in endomembrane transport, mitochondrial protein import and protein quality control and glucosinolate biosynthesis. Thus, we identified proteins involved in endocytosis, endoplasmic reticulum-Golgi transport and attachment sites of endoplasmic reticulum and plasma membrane (V-type proton ATPases, protein transport protein SEC13 homolog A, vesicle-associated protein 1-2, vacuolar protein sorting-associated protein 29, syntaxin-32, all upregulated in the mutants), mitochondrial import and electron transport chain (mitochondrial import inner membrane translocase subunits TIM23-2 and TIM13, mitochondrial NADH dehydrogenases, ATP synthases, cytochrome c oxidase subunit 6b-1, ADP,ATP carrier protein 2, downregulated in the mutants) and glucosinolate biosynthesis (3-isopropylmalate dehydrogenases 1, 2 and 3, methylthioalkylmalate synthase 1, cytochrome P450 83B1, Glutathione S-transferase F9, indole glucosinolate O-methyltransferase 1, adenylyl-sulfate kinase 1, all upregulated in mutants). Our results suggest broader biological roles of PLDα1 as anticipated so far.

Download Full-text

Shot-Gun Proteomic Analysis on Roots of Arabidopsis pldα1 Mutants Suggesting the Involvement of PLDα1 in Mitochondrial Protein Import, Vesicular Trafficking and Glucosinolate Biosynthesis

International Journal of Molecular Sciences ◽

10.3390/ijms20010082 ◽

2018 ◽

Vol 20 (1) ◽

pp. 82 ◽

Cited By ~ 1

Author(s):

Tomáš Takáč ◽

Olga Šamajová ◽

Pavol Vadovič ◽

Tibor Pechan ◽

Jozef Šamaj

Keyword(s):

Endoplasmic Reticulum ◽

Stress Responses ◽

Proteomic Analysis ◽

Protein Import ◽

Protein Quality ◽

Mitochondrial Protein ◽

Biotic And Abiotic Stress ◽

Mitochondrial Protein Import ◽

Enzymatic Production ◽

Glucosinolate Biosynthesis

Phospholipase Dα1 (PLDα1) belongs to phospholipases, a large phospholipid hydrolyzing protein family. PLDα1 has a substrate preference for phosphatidylcholine leading to enzymatic production of phosphatidic acid, a lipid second messenger with multiple cellular functions. PLDα1 itself is implicated in biotic and abiotic stress responses. Here, we present a shot-gun differential proteomic analysis on roots of two Arabidopsis pldα1 mutants compared to the wild type. Interestingly, PLDα1 deficiency leads to altered abundances of proteins involved in diverse processes related to membrane transport including endocytosis and endoplasmic reticulum-Golgi transport. PLDα1 may be involved in the stability of attachment sites of endoplasmic reticulum to the plasma membrane as suggested by increased abundance of synaptotagmin 1, which was validated by immunoblotting and whole-mount immunolabelling analyses. Moreover, we noticed a robust abundance alterations of proteins involved in mitochondrial import and electron transport chain. Notably, the abundances of numerous proteins implicated in glucosinolate biosynthesis were also affected in pldα1 mutants. Our results suggest a broader biological involvement of PLDα1 than anticipated thus far, especially in the processes such as endomembrane transport, mitochondrial protein import and protein quality control, as well as glucosinolate biosynthesis.

Download Full-text

From cytosol to mitochondria: the beginning of a protein journey

Biological Chemistry ◽

10.1515/hsz-2020-0110 ◽

2020 ◽

Vol 401 (6-7) ◽

pp. 645-661 ◽

Cited By ~ 5

Author(s):

Maria Clara Avendaño-Monsalve ◽

José Carlos Ponce-Rojas ◽

Soledad Funes

Keyword(s):

Mitochondrial Biogenesis ◽

Stress Responses ◽

Protein Import ◽

Protein Targeting ◽

Mitochondrial Protein ◽

Past Research ◽

Membrane Receptors ◽

Mitochondrial Proteins ◽

Mitochondrial Protein Import ◽

The Past

AbstractMitochondrial protein import is one of the key processes during mitochondrial biogenesis that involves a series of events necessary for recognition and delivery of nucleus-encoded/cytosol-synthesized mitochondrial proteins into the organelle. The past research efforts have mainly unraveled how membrane translocases ensure the correct protein sorting within the different mitochondrial subcompartments. However, early steps of recognition and delivery remain relatively uncharacterized. In this review, we discuss our current understanding about the signals on mitochondrial proteins, as well as in the mRNAs encoding them, which with the help of cytosolic chaperones and membrane receptors support protein targeting to the organelle in order to avoid improper localization. In addition, we discuss recent findings that illustrate how mistargeting of mitochondrial proteins triggers stress responses, aiming to restore cellular homeostasis.

Download Full-text

The amino terminus of the F1-ATPase beta-subunit precursor functions as an intramolecular chaperone to facilitate mitochondrial protein import.

Molecular and Cellular Biology ◽

10.1128/mcb.17.12.7169 ◽

1997 ◽

Vol 17 (12) ◽

pp. 7169-7177 ◽

Cited By ~ 14

Author(s):

P Hájek ◽

J Y Koh ◽

L Jones ◽

D M Bedwell

Keyword(s):

Protein Import ◽

Mitochondrial Protein ◽

Beta Subunit ◽

Amino Terminus ◽

Missense Mutations ◽

Primary Defect ◽

Mitochondrial Protein Import ◽

Mitochondrial Import ◽

Yeast Saccharomyces Cerevisiae ◽

F1 Atpase

Mitochondrial import signals have been shown to function in many steps of mitochondrial protein import. Previous studies have shown that the F1-ATPase beta-subunit precursor (pre-F1beta) of the yeast Saccharomyces cerevisiae contains an extended, functionally redundant mitochondrial import signal at its amino terminus. However, the full significance of this functionally redundant targeting sequence has not been determined. We now report that the extended pre-F1beta signal acts to maintain the precursor in an import-competent conformation prior to import, in addition to its previously characterized roles in mitochondrial targeting and translocation. We found that this extended signal is required for the efficient posttranslational mitochondrial import of pre-F1beta both in vivo and in vitro. To determine whether the pre-F1beta signal directly influences precursor conformation, fusion proteins that contain wild-type and mutant forms of the pre-F1beta import signal attached to the model passenger protein dihydrofolate reductase (DHFR) were constructed. Deletions that reduced the import signal to a minimal functional unit decreased both the half-time of precursor folding and the efficiency of mitochondrial import. To confirm that the reduced mitochondrial import associated with this truncated signal was due to a defect in its ability to maintain DHFR in a loosely folded conformation, we introduced structurally destabilizing missense mutations into the DHFR passenger to block precursor folding independently of the import signal. We found that the truncated signal imported this destabilized form of DHFR as efficiently as the intact targeting signal, indicating that the primary defect associated with the minimal signal is an inability to maintain the precursor in a loosely folded conformation. Our results suggest that the loss of this intramolecular chaperone function leads to defects in the early stages of the import process.

Download Full-text

Role of the Mitochondrial Protein Import Machinery and Protein Processing in Heart Disease

Frontiers in Cardiovascular Medicine ◽

10.3389/fcvm.2021.749756 ◽

2021 ◽

Vol 8 ◽

Author(s):

Fujie Zhao ◽

Ming-Hui Zou

Keyword(s):

Heart Disease ◽

Protein Import ◽

Protein Quality ◽

Mitochondrial Dynamics ◽

Mitochondrial Protein ◽

Nuclear Genome ◽

Major Protein ◽

Intermembrane Space ◽

Mitochondrial Protein Import ◽

Mitochondrial Import

Mitochondria are essential organelles for cellular energy production, metabolic homeostasis, calcium homeostasis, cell proliferation, and apoptosis. About 99% of mammalian mitochondrial proteins are encoded by the nuclear genome, synthesized as precursors in the cytosol, and imported into mitochondria by mitochondrial protein import machinery. Mitochondrial protein import systems function not only as independent units for protein translocation, but also are deeply integrated into a functional network of mitochondrial bioenergetics, protein quality control, mitochondrial dynamics and morphology, and interaction with other organelles. Mitochondrial protein import deficiency is linked to various diseases, including cardiovascular disease. In this review, we describe an emerging class of protein or genetic variations of components of the mitochondrial import machinery involved in heart disease. The major protein import pathways, including the presequence pathway (TIM23 pathway), the carrier pathway (TIM22 pathway), and the mitochondrial intermembrane space import and assembly machinery, related translocases, proteinases, and chaperones, are discussed here. This review highlights the importance of mitochondrial import machinery in heart disease, which deserves considerable attention, and further studies are urgently needed. Ultimately, this knowledge may be critical for the development of therapeutic strategies in heart disease.

Download Full-text