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Overlapping functions and protein-protein interactions of LRR-extensins in Arabidopsis.

ABSTRACT: Plant cell growth requires the coordinated expansion of the protoplast and the cell wall, which is controlled by an elaborate system of cell wall integrity (CWI) sensors linking the different cellular compartments. LRR-eXtensins (LRXs) are cell wall-attached extracellular regulators of cell wall formation and high-affinity binding sites for RALF (Rapid ALkalinization Factor) peptide hormones that trigger diverse physiological processes related to cell growth. LRXs function in CWI sensing and in the case of LRX4 of Arabidopsis thaliana, this activity was shown to involve interaction with the transmembrane Catharanthus roseus Receptor-Like Kinase1-Like (CrRLK1L) protein FERONIA (FER). Here, we demonstrate that binding of RALF1 and FER is common to most tested LRXs of vegetative tissue, including LRX1, the main LRX protein of root hairs. Consequently, an lrx1-lrx5 quintuple mutant line develops shoot and root phenotypes reminiscent of the fer-4 knock-out mutant. The previously observed membrane-association of LRXs, however, is FER-independent, suggesting that LRXs bind not only FER but also other membrane-localized proteins to establish a physical link between intra- and extracellular compartments. Despite evolutionary diversification of various LRX proteins, overexpression of several chimeric LRX constructs causes cross-complementation of lrx mutants, indicative of comparable functions among members of this protein family. Suppressors of the pollen-growth defects induced by mutations in the CrRLK1Ls ANXUR1/2 also alleviate lrx1 lrx2-induced mutant root hair phenotypes. This suggests functional similarity of LRX-CrRLK1L signaling processes in very different cell types and indicates that LRX proteins are components of conserved processes regulating cell growth.

SUBMITTER: Herger A

PROVIDER: S-EPMC7357788 | biostudies-literature | 2020 Jun

REPOSITORIES: biostudies-literature

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Overlapping functions and protein-protein interactions of LRR-extensins in Arabidopsis.

Herger Aline A Gupta Shibu S Kadler Gabor G Franck Christina Maria CM Boisson-Dernier Aurélien A Ringli Christoph C

PLoS genetics 20200619 6

Plant cell growth requires the coordinated expansion of the protoplast and the cell wall, which is controlled by an elaborate system of cell wall integrity (CWI) sensors linking the different cellular compartments. LRR-eXtensins (LRXs) are cell wall-attached extracellular regulators of cell wall formation and high-affinity binding sites for RALF (Rapid ALkalinization Factor) peptide hormones that trigger diverse physiological processes related to cell growth. LRXs function in CWI sensing and in ...[more]

PMID: 32559234

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Project description:BACKGROUND:Grafting is a technique widely used in horticulture. The processes involved in grafting are diverse, and the technique is commonly employed in studies focusing on the mechanisms that regulate cell differentiation or response of plants to abiotic stress. Information on the changes in the composition of the cell wall that occur during the grafting process is scarce. Therefore, this study was carried out for analyzing the composition of the cell wall using Arabidopsis hypocotyls as an example. During the study, the formation of a layer that covers the surface of the graft union was observed. So, this study also aimed to describe the histological and cellular changes that accompany autografting of Arabidopsis hypocotyls and to perform preliminary chemical and structural analyses of extracellular material that seals the graft union. RESULTS:During grafting, polyphenolic and lipid compounds were detected, along with extracellular deposition of carbohydrate/protein material. The spatiotemporal changes observed in the structure of the extracellular material included the formation of a fibrillar network, polymerization of the fibrillar network into a membranous layer, and the presence of bead-like structures on the surface of cells in established graft union. These bead-like structures appeared either "closed" or "open". Only three cell wall epitopes, namely: LM19 (un/low-methyl-esterified homogalacturonan), JIM11, and JIM20 (extensins), were detected abundantly on the cut surfaces that made the adhesion plane, as well as in the structure that covered the graft union and in the bead-like structures, during the subsequent stages of regeneration. CONCLUSIONS:To the best of our knowledge, this is the first report on the composition and structure of the extracellular material that gets deposited on the surface of graft union during Arabidopsis grafting. The results showed that unmethyl-esterified homogalacturonan and extensins are together involved in the adhesion of scion and stock, as well as taking part in sealing the graft union. The extracellular material is of importance not only due to the potential pectin-extensin interaction but also due to its origin. The findings presented here implicate a need for studies with biochemical approach for a detailed analysis of the composition and structure of the extracellular material.

Project description:Purpose: Bacillus anthracis produces three regulators, AtxA, AcpA and AcpB, which control virulence gene transcription and belong to an emerging class of regulators termed ‘PCVRs’ (Phosphoenolpyruvate-dependent phosphotransferase regulation Domain-Containing Virulence Regulators). AtxA, named for its control of toxin gene expression, is the master virulence regulator and archetype PCVR. AcpA and AcpB are less well studied. Reports of PCVR activity suggest overlapping function. We used RNA-Seq to assess the regulons of the paralogous regulators in strains constructed to express individual PCVRs at native levels. Methods: Cultures were incubated in toxin-inducing conditions, and expression of each PCVR in the atxAacpAacpB-null background strain was induced with an IPTG concentration that yielded native protein levels for each PCVR. RNA was extracted when cultures reached late-exponential phase using saturated acid phenol extraction, followed by RNA precipitation from the aqueous phase. Creation of libraries for NGS analysis used total RNA (1.0 μg). Samples were treated with Ribo-Zero (Epicentre) to remove ribosomal RNA prior to fragmentation using divalent cations and heat. Libraries were created using an Illumina TruSeq sample preparation kit following the protocol as recommended by the manufacturer. NGS sequencing was performed as a paired-end 50 base sequence using an Illumina HiSeq 1500 following the protocol recommended by the manufacturer. Results: Plasmid and chromosome-borne genes were PCVR controlled, with AtxA, AcpA and AcpB having a ≥4-fold effect on transcript levels of 145, 130 and 49 genes respectively. Several genes were coregulated by two or three PCVRs. Of the 203 transcripts associated with pXO1, 15 were altered at least fourfold: 13 by AtxA and 2 by AcpA. No pXO1-derived transcripts were affected fourfold or greater by AcpB. Read maps revealed that many of the regulated transcripts from pXO1 were associated with the 35 kb region that lies within a larger 44.8 kb pathogenicity island (PAI) (Thorne, 1993). We detected transcripts for 110 genes on pXO2, of which 21 were regulated fourfold or greater by at least one of the three PCVRs. The highly regulated genes clustered within a 35.5 kb region of the plasmid, which includes the capsule biosynthetic operon capBCADE followed by the weakly co-transcribed acpB gene. A significantly smaller proportion of PCVR-regulated genes mapped to the chromosome. Of the 5593 transcripts, 198 were altered fourfold or greater by one or more PCVRs. Clustering of PCVR-regulated chromosome genes was not apparent. Among the most highly regulated chromosomal genes were many genes associated with branched chain amino acid (BCAA) synthesis and uptake. Of the 17 genes implicated in BCAA biosynthesis, expression of 13 of these genes is repressed by AtxA and AcpB. In this work, we have shown that the three PRD-containing virulence regulators of B. anthracis exhibit overlapping and divergent target gene specificities. Conclusions: In this work, we show the relative activities of the PCVRs for specific and co-regulated genes. The results reveal the vast effects of the PCVRs on B. anthracis gene expression and indicate a high degree of functional similarity among the regulators. Our investigations provide insight into control of B. anthracis gene expression and expand our knowledge of structure and function of this emerging class of virulence gene regulators.

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Dataset Information

Overlapping functions and protein-protein interactions of LRR-extensins in Arabidopsis.

Publications

Overlapping functions and protein-protein interactions of LRR-extensins in Arabidopsis.

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