Arabidopsis thaliana G3BP Ortholog Rescues Mammalian Stress Granule Phenotype across Kingdoms

Stress granules (SGs) are dynamic RNA–protein complexes localized in the cytoplasm that rapidly form under stress conditions and disperse when normal conditions are restored. The formation of SGs depends on the Ras-GAP SH3 domain-binding protein (G3BP). Formations, interactions and functions of plant and human SGs are strikingly similar, suggesting a conserved mechanism. However, functional analyses of plant G3BPs are missing. Thus, members of the Arabidopsis thaliana G3BP (AtG3BP) protein family were investigated in a complementation assay in a human G3BP knock-out cell line. It was shown that two out of seven AtG3BPs were able to complement the function of their human homolog. GFP-AtG3BP fusion proteins co-localized with human SG marker proteins Caprin-1 and eIF4G1 and restored SG formation in G3BP double KO cells. Interaction between AtG3BP-1 and -7 and known human G3BP interaction partners such as Caprin-1 and USP10 was also demonstrated by co-immunoprecipitation. In addition, an RG/RGG domain exchange from Arabidopsis G3BP into the human G3BP background showed the ability for complementation. In summary, our results support a conserved mechanism of SG function over the kingdoms, which will help to further elucidate the biological function of the Arabidopsis G3BP protein family.

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Cep192, a Novel Missing Link between the Centrosomal Core and Corona in Dictyostelium Amoebae

Cells ◽

10.3390/cells10092384 ◽

2021 ◽

Vol 10 (9) ◽

pp. 2384

Author(s):

Valentin Pitzen ◽

Sophia Sander ◽

Otto Baumann ◽

Ralph Gräf ◽

Irene Meyer

Keyword(s):

Research Group ◽

Fusion Proteins ◽

Outer Core ◽

Core Structure ◽

The Core ◽

Marker Proteins ◽

Interaction Partners ◽

Core Components ◽

Corona Structure ◽

Centrosomal Proteins

The Dictyostelium centrosome is a nucleus-associated body with a diameter of approx. 500 nm. It contains no centrioles but consists of a cylindrical layered core structure surrounded by a microtubule-nucleating corona. At the onset of mitosis, the corona disassembles and the core structure duplicates through growth, splitting, and reorganization of the outer core layers. During the last decades our research group has characterized the majority of the 42 known centrosomal proteins. In this work we focus on the conserved, previously uncharacterized Cep192 protein. We use superresolution expansion microscopy (ExM) to show that Cep192 is a component of the outer core layers. Furthermore, ExM with centrosomal marker proteins nicely mirrored all ultrastructurally known centrosomal substructures. Furthermore, we improved the proximity-dependent biotin identification assay (BioID) by adapting the biotinylase BioID2 for expression in Dictyostelium and applying a knock-in strategy for the expression of BioID2-tagged centrosomal fusion proteins. Thus, we were able to identify various centrosomal Cep192 interaction partners, including CDK5RAP2, which was previously allocated to the inner corona structure, and several core components. Studies employing overexpression of GFP-Cep192 as well as depletion of endogenous Cep192 revealed that Cep192 is a key protein for the recruitment of corona components during centrosome biogenesis and is required to maintain a stable corona structure.

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Coordination and assembly of protein complexes encoded across mitochondrial and nuclear genomes is assisted by CLPP2 in Arabidopsis thaliana

10.1101/2020.01.22.907055 ◽

2020 ◽

Author(s):

Jakob Petereit ◽

Owen Duncan ◽

Monika W Murcha ◽

Ricarda Fenske ◽

Emilia Cincu ◽

...

Keyword(s):

Arabidopsis Thaliana ◽

Protein Degradation ◽

Protein Import ◽

Protein Complexes ◽

Single Gene ◽

Mitochondrial Protein ◽

Protein Homeostasis ◽

Knock Out ◽

Genes Encoding ◽

Nuclear Genomes

AbstractProtein homeostasis in eukaryotic organelles and their progenitor prokaryotes is regulated by a series of proteases including the caseinolytic protease (CLPP). CLPP has essential roles in chloroplast biogenesis and maintenance, but the significance of the plant mitochondrial CLPP remains unknown and factors that aid coordination of nuclear and mitochondrial encoded subunits for complex assembly in mitochondria await discovery. We generated knock-out lines of the single gene for the mitochondrial CLP protease subunit, CLPP2, in Arabidopsis thaliana. Mutants had higher abundance of transcripts from mitochondrial genes encoding OXPHOS protein complexes, while transcripts for nuclear genes encoding other subunits of the same complexes showed no change in abundance. In contrast, the protein abundance of specific nuclear-encoded subunits in OXPHOS complexes I and V increased in CLPP2 knockouts, without accumulation of mitochondrial-encoded counterparts in the same complex. Protein complexes mainly or entirely encoded in the nucleus were unaffected. Analysis of protein import, assembly and function of Complex I revealed that while function was retained, protein homeostasis was disrupted through decreased assembly, leading to accumulation of soluble subcomplexes of nuclear-encoded subunits. Therefore, CLPP2 contributes to the mitochondrial protein degradation network through supporting coordination and assembly of protein complexes encoded across mitochondrial and nuclear genomes.One sentence summaryCLPP contributes to the mitochondrial protein degradation network through supporting coordination and assembly of protein complexes encoded across mitochondrial and nuclear genomes.

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Faculty Opinions recommendation of An Arabidopsis thaliana knock-out mutant of the chloroplast triose phosphate/phosphate translocator is severely compromised only when starch synthesis, but not starch mobilisation is abolished.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1010844.178261 ◽

2003 ◽

Author(s):

Sjef Smeekens

Keyword(s):

Arabidopsis Thaliana ◽

Starch Synthesis ◽

Knock Out ◽

Triose Phosphate ◽

Phosphate Translocator

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Alu RNA-protein complexes formed in vitro react with a novel lupus autoantibody.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(17)39098-1 ◽

1985 ◽

Vol 260 (21) ◽

pp. 11781-11786

Author(s):

R Kole ◽

L D Fresco ◽

J D Keene ◽

P L Cohen ◽

R A Eisenberg ◽

...

Keyword(s):

Protein Complexes ◽

Alu Rna ◽

Rna Protein Complexes

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Ca2+-Dependent Protein Kinase 6 Enhances KAT2 Shaker Channel Activity in Arabidopsis thaliana

International Journal of Molecular Sciences ◽

10.3390/ijms22041596 ◽

2021 ◽

Vol 22 (4) ◽

pp. 1596

Author(s):

Elsa Ronzier ◽

Claire Corratgé-Faillie ◽

Frédéric Sanchez ◽

Christian Brière ◽

Tou Cheu Xiong

Keyword(s):

Arabidopsis Thaliana ◽

Resonance Energy Transfer ◽

Resonance Energy ◽

Physical Interaction ◽

Fluorescence Lifetime Imaging ◽

Dependent Manner ◽

In Planta ◽

Knock Out ◽

Calcium Dependent ◽

Dependent Protein

Post-translational regulations of Shaker-like voltage-gated K+ channels were reported to be essential for rapid responses to environmental stresses in plants. In particular, it has been shown that calcium-dependent protein kinases (CPKs) regulate Shaker channels in plants. Here, the focus was on KAT2, a Shaker channel cloned in the model plant Arabidopsis thaliana, where is it expressed namely in the vascular tissues of leaves. After co-expression of KAT2 with AtCPK6 in Xenopuslaevis oocytes, voltage-clamp recordings demonstrated that AtCPK6 stimulates the activity of KAT2 in a calcium-dependent manner. A physical interaction between these two proteins has also been shown by Förster resonance energy transfer by fluorescence lifetime imaging (FRET-FLIM). Peptide array assays support that AtCPK6 phosphorylates KAT2 at several positions, also in a calcium-dependent manner. Finally, K+ fluorescence imaging in planta suggests that K+ distribution is impaired in kat2 knock-out mutant leaves. We propose that the AtCPK6/KAT2 couple plays a role in the homeostasis of K+ distribution in leaves.

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Analysis of an Arabidopsis thaliana protein family, structurally related to cytochromes b 561 and potentially involved in catecholamine biochemistry in plants

Journal of Plant Physiology ◽

10.1078/0176-1617-01064 ◽

2004 ◽

Vol 161 (2) ◽

pp. 175-181 ◽

Cited By ~ 14

Author(s):

W.i.m. Verelst ◽

H.a.n. Asard

Keyword(s):

Arabidopsis Thaliana ◽

Protein Family ◽

Catecholamine Biochemistry

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Molecular biology approaches in bioadhesion research

Beilstein Journal of Nanotechnology ◽

10.3762/bjnano.5.112 ◽

2014 ◽

Vol 5 ◽

pp. 983-993 ◽

Cited By ~ 12

Author(s):

Marcelo Rodrigues ◽

Birgit Lengerer ◽

Thomas Ostermann ◽

Peter Ladurner

Keyword(s):

Molecular Biology ◽

Biological Function ◽

Molecular Approach ◽

Research Groups ◽

Blast Search ◽

Search Facility ◽

New Research ◽

And Function ◽

Functional Analyses

The use of molecular biology tools in the field of bioadhesion is still in its infancy. For new research groups who are considering taking a molecular approach, the techniques presented here are essential to unravelling the sequence of a gene, its expression and its biological function. Here we provide an outline for addressing adhesion-related genes in diverse organisms. We show how to gradually narrow down the number of candidate transcripts that are involved in adhesion by (1) generating a transcriptome and a differentially expressed cDNA list enriched for adhesion-related transcripts, (2) setting up a BLAST search facility, (3) perform an in situ hybridization screen, and (4) functional analyses of selected genes by using RNA interference knock-down. Furthermore, latest developments in genome-editing are presented as new tools to study gene function. By using this iterative multi-technologies approach, the identification, isolation, expression and function of adhesion-related genes can be studied in most organisms. These tools will improve our understanding of the diversity of molecules used for adhesion in different organisms and these findings will help to develop innovative bio-inspired adhesives.

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