A contribution of DNA methylation to defense against invading nucleic acids and maintenance of genome integrity is uncontested; however, our understanding of the extent of involvement of this ...epigenetic mark in genome-wide gene regulation and plant developmental control is incomplete. Here, we knock out all five known DNA methyltransferases in Arabidopsis, generating DNA methylation-free plants. This quintuple mutant exhibits a suite of developmental defects, unequivocally demonstrating that DNA methylation is essential for multiple aspects of plant development. We show that CG methylation and non-CG methylation are required for a plethora of biological processes, including pavement cell shape, endoreduplication, cell death, flowering, trichome morphology, vasculature and meristem development, and root cell fate determination. Moreover, we find that DNA methylation has a strong dose-dependent effect on gene expression and repression of transposable elements. Taken together, our results demonstrate that DNA methylation is dispensable for Arabidopsis survival but essential for the proper regulation of multiple biological processes.
Brassinosteroids (BRs) play key roles in diverse plant growth processes through a complex signaling pathway. Components orchestrating the BR signaling pathway include receptors such as kinases, ...transcription factors, protein kinases and phosphatases. The proper functioning of the receptor kinase BRI1 and the transcription factors BES1/BZR1 depends on their dephosphorylation by type 2A protein phosphatases (PP2A). In this work, we report that an additional phosphatase family, type one protein phosphatases (PP1), contributes to the regulation of the BR signaling pathway. Co-immunoprecipitation and BiFC experiments performed in Arabidopsis plants overexpressing durum wheat TdPP1 showed that TdPP1 interacts with dephosphorylated BES1, but not with the BRI1 receptor. Higher levels of dephosphorylated, active BES1 were observed in these transgenic lines upon BR treatment, indicating that TdPP1 modifies the BR signaling pathway by activating BES1. Moreover, ectopic expression of durum wheat TdPP1 lead to an enhanced growth of primary roots in comparison to wild-type plants in presence of BR. This phenotype corroborates with a down-regulation of the BR-regulated genes CPD and DWF4. These data suggest a role of PP1 in fine-tuning BR-driven responses, most likely via the control of the phosphorylation status of BES1.
Reversible phosphorylation is an essential mechanism regulating signal transduction during development and environmental stress responses. An important number of dephosphorylation events in the cell ...are catalyzed by type one protein phosphatases (PP1), which catalytic activity is driven by the binding of regulatory proteins that control their substrate specificity or subcellular localization. Plants harbor several PP1 isoforms accounting for large functional redundancies. While animal PP1s were reported to play relevant roles in controlling multiple cellular processes, plant orthologs remain poorly studied. To decipher the role of plant PP1s, we compared PP1 genes from three monocot species, Brachypodium, common wheat and rice at the genomic and transcriptomic levels. To gain more insight into the wheat PP1 proteins, we identified and characterized TdPP1a, the first wheat type one protein phosphatase from a Tunisian durum wheat variety Oum Rabiaa3. TdPP1a is highly conserved in sequence and structure when compared to mammalian, yeast and other plant PP1s. We demonstrate that TdPP1a is an active, metallo-dependent phosphatase in vitro and is able to interact with AtI2, a typical regulator of PP1 functions. Also, TdPP1a is capable to complement the heat stress sensitivity of the yeast mutant indicating that TdPP1a is functional also in vivo. Moreover, transient expression of TdPP1a::GFP in tobacco leaves revealed that it is ubiquitously distributed within the cell, with a strong accumulation in the nucleus. Finally, transcriptional analyses showed similar expression levels in roots and leaves of durum wheat seedlings. Interestingly, the expression in leaves is significantly induced following salinity stress, suggesting a potential role of TdPP1a in wheat salt stress response.
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Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The control of optimal root growth and plant stress responses depends largely on a variety of phytohormones among which auxin and brassinosteroids (BRs) are the most influential. We have previously ...reported that the durum wheat type 1 protein phosphatase TdPP1 participates in the control of root growth by modulating BR signaling. In this study, we pursue our understanding of how TdPP1 fulfills this regulatory function on root growth by evaluating the physiological and molecular responses of Arabidopsis TdPP1 over-expressing lines to abiotic stresses. Our results showed that when exposed to 300 mM Mannitol or 100 mM NaCl, the seedlings of TdPP1 over-expressors exhibit modified root architecture with higher lateral root density, and longer root hairs concomitant with a lower inhibition of the primary root growth. These lines also exhibit faster gravitropic response and a decrease in primary root growth inhibition when exposed to high concentrations of exogenous IAA. On another hand, a cross between TdPP1 overexpressors and DR5:GUS marker line was performed to monitor auxin accumulation in roots. Remarkably, the TdPP1 overexpression resulted in an enhanced auxin gradient under salt stress with a higher accumulation in primary and lateral root tips. Moreover, TdPP1 transgenics exhibit a significant induction of a subset of auxin-responsive genes under salt stress conditions. Therefore, our results reveal a role of PP1 in enhancing auxin signaling to help shape greater root plasticity thus improving plant stress resilience.
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•The root system is expanded under abiotic stress upon PP1 over-expression.•PP1 contributes to inducing auxin transport and signaling.•PP1 promotes abiotic stress resilience by increasing the plasticity of the root system.
Xylem patterning in the root is established through the creation of opposing gradients of miRNAs and their targets, transcripts of the HD-ZIP III family of transcriptions factors, enabled by the ...cell-to-cell spread of the former. The miRNAs regulating xylem patterning, miR165/6, move through plasmodesmata, but how their trafficking is regulated remains elusive. Here, we describe that simultaneous mutation of the plasma membrane- and plasmodesmata-localized receptor-like kinases (RLKs) BARELY ANY MERISTEM (BAM) 1 and 2 or expression of the geminivirus-encoded BAM1/2-interactor C4 results in higher accumulation and broader distribution of the HD-ZIP III transcripts despite normal total accumulation of miR165/6, and ultimately causes defects in xylem patterning, which depend on the function of the aforementioned miRNA targets. Taken together, our results show that BAM1 and BAM2 are redundantly required for proper xylem patterning in the
root, by ensuring the proper distribution and accumulation of miR165/6-targeted transcripts.
Blockchain-based applications are basically built on smart contracts, which are widely different in regards of the encoded logic and the used standards. When talking about Ethereum standards, ERC-721 ...is a well-known standard interface developed for Non-Fungible Tokens. Even though it is standard-based contracts that are more and more exploited, prior work on smart contracts verification mostly investigates efforts in regards of specific vulnerabilities. To address this gap, this paper introduces a formal modeling and verification approach for Ethereum smart contracts including the standard-based ones. We propose a model checking framework that, according to a Solidity smart contract provided as an input, uses ERC guidelines as a standard template to extract the related security properties. Another added benefit of our proposal consists on modeling ERC contracts using the extended finite state machine formalism. As a proof of concept, we illustrate our model checking approach through an NFT contract.
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