Plant UBX Domain-containing Protein 1, PUX1, Regulates the Oligomeric Structure and Activity of Arabidopsis CDC48

Rancour, David M.; Park, Sookhee; Knight, Seth D.; Bednarek, Sebastian Y.

doi:10.1074/jbc.m405498200

Cited by 99 publications

(125 citation statements)

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“…Samples were incubated for 30 min on ice followed by centrifugation (Beckman TLA100.1, 90,000 rpm, 30 min, 4°C). Pellet fractions were washed with MIB DTT and 20 g of protein equivalents of supernatant and pellet fractions were analyzed by immunoblotting using antisera specific for FAC1 (see supplementary Materials and Methods), cytosolic (PUX1) (21), integral membrane (AtSEC12) (22), and peripheral membrane (AtCDC48) (20) proteins. Prior to immunoblotting, the nitrocellulose membrane was analyzed by Ponceau S staining to confirm protein recovery and equal loading.…”

Section: Methodsmentioning

confidence: 99%

Membrane Association, Mechanism of Action, and Structure of Arabidopsis Embryonic Factor 1 (FAC1)

Han

Bingman

Mahnke

et al. 2006

Journal of Biological Chemistry

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Embryonic factor 1 (FAC1) is one of the earliest expressed plant genes and encodes an AMP deaminase (AMPD), which is also an identified herbicide target. This report identifies an N-terminal transmembrane domain in Arabidopsis FAC1, explores subcellular fractionation, and presents a 3.3-Å globular catalytic domain x-ray crystal structure with a bound herbicide-based transition state inhibitor that provides the first glimpse of a complete AMPD active site. FAC1 contains an (␣/␤) 8 -barrel characterized by loops in place of strands 5 and 6 that places it in a small subset of the amidohydrolase superfamily with imperfect folds. Unlike tetrameric animal orthologs, FAC1 is a dimer and each subunit contains an exposed Walker A motif that may be involved in the dramatic combined K m (25-80-fold lower) and V max (5-6-fold higher) activation by ATP. Normal mode analysis predicts a hinge motion that flattens basic surfaces on each monomer that flank the dimer interface, which suggests a reversible association between the FAC1 globular catalytic domain and intracellular membranes, with N-terminal transmembrane and disordered linker regions serving as the anchor and attachment to the globular catalytic domain, respectively.Embryonic factor 1 (FAC1) 5 was recently identified as one of the earliest expressed plant genes and is essential for the zygote to embryo transition in Arabidopsis thaliana (1). The zygote-lethal phenotype is characterized by developmental arrest at the 8 -16-cell stage and mutant embryo shriveling 2-3 days after fertilization. The Arabidopsis FAC1 locus encodes an AMP deaminase (AMPD; EC 3.5.4.6), which is a eukaryotic enzyme that catalyzes the hydrolytic deamination of AMP to IMP. AMPD has also been identified as the intracellular target for a class of herbicides that are produced by fungal pathogens. Carbocyclic coformycin was initially discovered in Saccharothrix (2), and plant cells can take up this diffusible nucleoside and 5Ј-phosphorylate it to produce a potent transition state inhibitor of AMPD (3). Exposure to carbocyclic coformycin results in cessation of seedling growth, followed by paling and necrosis at the apical meristem (3). Coformycin, a structurally related compound produced by a number of microbes (4, 5), also has herbicidal properties (5). Although the intracellular metabolism of this compound in plants has not been examined, its mode of action is presumably similar because coformycin 5Ј-phosphate is a transition state inhibitor of rabbit muscle AMPD (6). Both herbicides are inhibitors of mammalian adenosine deaminase (ADA) (3, 6), but the lack of this enzyme in plants (3, 7-9) supports the argument that AMPD is the intracellular target for their nucleotide derivatives. Taken together, these observations suggest that AMPD is essential throughout the plant life cycle. However, the underlying basis for lethality of a FAC1 null phenotype and for herbicidal toxicity related to the catalytic inhibition of this enzyme is not known.AMPD catalyzes the initial step in adenine to guanine ribon...

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Section: Methodsmentioning

confidence: 99%

Membrane Association, Mechanism of Action, and Structure of Arabidopsis Embryonic Factor 1 (FAC1)

Han

Bingman

Mahnke

et al. 2006

Journal of Biological Chemistry

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“…S4) is encoded by a gene that is part of at least a 15-member gene family (PUX) in Arabidopsis (77). Notably, this family also includes PUX1, which encodes a protein that, similar to the yeast and mammalian Shp1 proteins, binds to and modulates the activity of Cdc48 (52, 75,76). Studies aimed at determining whether Arabidopsis PUX4 and PUX1 and/or other members of the PUX gene family, as well as putative Arabidopsis DOA1 and UFD2 genes, are involved in Fad3 regulation are currently being pursued in our laboratories through a combination of reverse genetic studies and expression of epitope-tagged Fad3 in mutant transgenic plants.…”

Section: Identification Of Specific Components Of the Erad Pathway Inmentioning

confidence: 99%

Temperature-sensitive Post-translational Regulation of Plant Omega-3 Fatty-acid Desaturases Is Mediated by the Endoplasmic Reticulum-associated Degradation Pathway

O'Quin

Bourassa

Zhang

et al. 2010

Journal of Biological Chemistry

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Changes in ambient temperature represent a major physiological challenge to membranes of poikilothermic organisms. In plants, the endoplasmic reticulum (ER)-localized omega-3 fatty-acid desaturases (Fad3) increase the production of polyunsaturated fatty acids at cooler temperatures, but the FAD3 genes themselves are typically not up-regulated during this adaptive response. Here, we expressed two closely related plant FAD3 genes in yeast cells and found that their enzymes produced significantly different amounts of omega-3 fatty acids and that these differences correlated to differences in rates of protein turnover. Domain-swapping and mutagenesis experiments revealed that each protein contained a degradation signal in its N terminus and that the charge density of a PEST-like sequence within this region was largely responsible for the differences in rates of protein turnover. The half-life of each Fad3 protein was increased at cooler temperatures, and protein degradation required specific components of the ER-associated degradation pathway including the Cdc48 adaptor proteins Doa1, Shp1, and Ufd2. Expression of the Fad3 proteins in tobacco cells incubated with the proteasomal inhibitor MG132 further confirmed that they were degraded via the proteasomal pathway in plants. Collectively, these findings indicate that Fad3 protein abundance is regulated by a combination of cis-acting degradation signals and the ubiquitin-proteasome pathway and that modulation of Fad3 protein amounts in response to temperature may represent one mechanism of homeoviscous adaptation in plants.

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“…Rhodamine-conjugated anti-rabbit (Sigma; http://www.sigmaaldrich.com) and Cy3 TM -conjugated anti-mouse (Jackson Labs) were used as secondary antibodies at 1:300. For Western blots, anti-GFP (Davis and Vierstra, 1998) was used at 1:8000 and anti-PUX1 (Rancour et al, 2004) and anti-PEP12 (da Silva Conceiçã o et al, 1997) were used at 1:5000. Detection of positive signals was performed using HP-conjugated anti-rabbit at 1:20 000 (Sigma).…”

Section: Antibodies and In Situ Immunolocalizationmentioning

confidence: 99%

ARL2, ARG1 and PIN3 define a gravity signal transduction pathway in root statocytes

Harrison

Masson²

2007

The Plant Journal

171

170

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SummaryALTERED RESPONSE TO GRAVITY1 (ARG1) and its paralog ARG1-LIKE2 (ARL2) are J-domain proteins that are required for normal root and hypocotyl gravitropism. In this paper, we show that both ARL2 and ARG1 function in a gravity signal transduction pathway with PIN3, an auxin efflux facilitator that is expressed in the statocytes. In gravi-stimulated roots, PIN3 relocalizes to the lower side of statocytes, a process that is thought to, in part, drive the asymmetrical redistribution of auxin toward the lower flank of the root. We show that ARL2 and ARG1 are required for PIN3 relocalization and asymmetrical distribution of auxin upon gravistimulation. ARL2 is expressed specifically in the root statocytes, where it localizes to the plasma membrane. Upon ectopic expression, ARL2 is also found at the cell plate of dividing cells during cytokinesis, an area of intense membrane dynamics. Mutations in ARL2 and ARG1 also result in auxin-related expansion of the root cap columella, consistent with a role for ARL2 and ARG1 in regulating auxin flux through the root tip. Together these data suggest that ARL2 and ARG1 functionally link gravity sensation in the statocytes to auxin redistribution through the root cap.

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Plant UBX Domain-containing Protein 1, PUX1, Regulates the Oligomeric Structure and Activity of Arabidopsis CDC48

Cited by 99 publications

References 75 publications

Membrane Association, Mechanism of Action, and Structure of Arabidopsis Embryonic Factor 1 (FAC1)

Membrane Association, Mechanism of Action, and Structure of Arabidopsis Embryonic Factor 1 (FAC1)

Temperature-sensitive Post-translational Regulation of Plant Omega-3 Fatty-acid Desaturases Is Mediated by the Endoplasmic Reticulum-associated Degradation Pathway

ARL2, ARG1 and PIN3 define a gravity signal transduction pathway in root statocytes

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