Cell wall integrity modulates HOOKLESS1 and PHYTOCHROME INTERACTING FACTOR4 expression controlling apical hook formation.

Formation of the apical hook in etiolated dicot seedlings results from differential growth in the hypocotyl apex and is tightly controlled by environmental cues and hormones, among which auxin and gibberellins (GAs) play an important role. Cell expansion is tightly regulated by the cell wall, but whether and how feedback from this structure contributes to hook development is still unclear. Here, we show that etiolated seedlings of the Arabidopsis (Arabidopsis thaliana) quasimodo2-1 (qua2) mutant, defective in pectin biosynthesis, display severe defects in apical hook formation and maintenance, accompanied by loss of asymmetric auxin maxima and of differential cell expansion. Moreover, qua2 seedlings show reduced expression of HOOKLESS 1 (HLS1) and PHYTOCHROME INTERACTING FACTOR 4 (PIF4), which are positive regulators of hook formation. Treatment of wild-type seedlings with the cellulose inhibitor isoxaben (isx) also prevents hook development and represses HLS1 and PIF4 expression. Exogenous GAs, loss of DELLA proteins or HLS1 overexpression partially restore hook development in qua2 and isx-treated seedlings. Interestingly, increased agar concentration in the medium restores, both in qua2 and isx-treated seedlings, hook formation, asymmetric auxin maxima and PIF4 and HLS1 expression. Analysis of plants expressing a FRET-based GA sensor indicate that isx reduces accumulation of GAs in the apical hook region in a turgor-dependent manner. Lack of the cell wall integrity sensor THESEUS 1, which modulates turgor loss point, restores hook formation in qua2 and isx-treated seedlings. We propose that turgor-dependent signals link changes in cell wall integrity to the PIF4-HLS1 signalling module to control differential cell elongation during hook formation.

Mode Conversion Trimming in Asymmetric Directional Couplers Enabled by Silicon Ion Implantation.

An on-chip asymmetric directional coupler (DC) can convert fundamental modes to higher-order modes and is one of the core components of mode-division multiplexing (MDM) technology. In this study, we propose that waveguides of the asymmetric DC can be trimmed by silicon ion implantation to tune the effective refractive index and facilitate mode conversion into higher-order modes. Through this method of tuning, transmission changes of up to 18 dB have been realized with one ion implantation step. In addition, adjusting the position of the ion implantation on the waveguide can provide a further degree of control over the transmission into the resulting mode. The results of this work present a promising new route for the development of high-efficiency, low-loss mode converters for integrated photonic platforms, and aim to facilitate the application of MDM technology in emerging photonic neuromorphic computing.

The Cotton Wall-Associated Kinase GhWAK7A Mediates Responses to Fungal Wilt Pathogens by Complexing with the Chitin Sensory Receptors.

Plant receptor-like kinases (RLKs) are important players in response to pathogen infections. Verticillium and Fusarium wilts, caused by Verticillium dahliae (Vd) and Fusarium oxysporum f. sp vasinfectum (Fov), respectively, are among the most devastating diseases in cotton (Gossypium spp). To understand the cotton response to these soil-borne fungal pathogens, we performed a genome-wide in silico characterization and functional screen of diverse RLKs for their involvement in cotton wilt diseases. We identified Gossypium hirsutum GhWAK7A, a wall-associated kinase, that positively regulates cotton response to both Vd and Fov infections. Chitin, the major constituent of the fungal cell wall, is perceived by lysin-motif-containing RLKs (LYKs/CERK1), leading to the activation of plant defense against fungal pathogens. A conserved chitin sensing and signaling system is present in cotton, including chitin-induced GhLYK5-GhCERK1 dimerization and phosphorylation, and contributes to cotton defense against Vd and Fov Importantly, GhWAK7A directly interacts with both GhLYK5 and GhCERK1 and promotes chitin-induced GhLYK5-GhCERK1 dimerization. GhWAK7A phosphorylates GhLYK5, which itself does not have kinase activity, but requires phosphorylation for its function. Consequently, GhWAK7A plays a crucial role in chitin-induced responses. Thus, our data reveal GhWAK7A as an important component in cotton response to fungal wilt pathogens by complexing with the chitin receptors.

The plasma membrane-associated Ca2+ -binding protein, PCaP1, is required for oligogalacturonide and flagellin-induced priming and immunity.

Early signalling events in response to elicitation include reversible protein phosphorylation and re-localization of plasma membrane (PM) proteins. Oligogalacturonides (OGs) are a class of damage-associated molecular patterns (DAMPs) that act as endogenous signals to activate the plant immune response. Previous data on early phosphoproteome changes in Arabidopsis thaliana upon OG perception uncovered the immune-related phospho-regulation of several membrane proteins, among which PCaP1, a PM-anchored protein with actin filament-severing activity, was chosen for its potential involvement in OG- and flagellin-triggered responses. Here, we demonstrate that PCaP1 is required for late, but not early, responses induced by OGs and flagellin. Moreover, pcap1 mutants, unlike the wild type, are impaired in the recovery of full responsiveness to a second treatment with OGs performed 24 h after the first one. Localization studies on PCaP1 upon OG treatment in plants expressing a functional PCaP1-GFP fusion under the control of PCaP1 promoter revealed fluorescence on the PM, organized in densely packed punctate structures, previously reported as microdomains. Fluorescence was found to be associated also with endocytic vesicles, the number of which rapidly increased after OG treatment, suggesting both an endocytic turnover of PCaP1 for maintaining its homeostasis at the PM and an OG-induced endocytosis.

The Arabidopsis thaliana LysM-containing Receptor-Like Kinase 2 is required for elicitor-induced resistance to pathogens.

In Arabidopsis thaliana, perception of chitin from fungal cell walls is mediated by three LysM-containing Receptor-Like Kinases (LYKs): CERK1, which is absolutely required for chitin perception, and LYK4 and LYK5, which act redundantly. The role in plant innate immunity of a fourth LYK protein, LYK2, is currently not known. Here we show that CERK1, LYK2 and LYK5 are dispensable for basal susceptibility to B. cinerea but are necessary for chitin-induced resistance to this pathogen. LYK2 is dispensable for chitin perception and early signalling events, though it contributes to callose deposition induced by this elicitor. Notably, LYK2 is also necessary for enhanced resistance to B. cinerea and Pseudomonas syringae induced by flagellin and for elicitor-induced priming of defence gene expression during fungal infection. Consistently, overexpression of LYK2 enhances resistance to B. cinerea and P. syringae and results in increased expression of defence-related genes during fungal infection. LYK2 appears to be required to establish a primed state in plants exposed to biotic elicitors, ensuring a robust resistance to subsequent pathogen infections.

Extracellular DAMPs in Plants and Mammals: Immunity, Tissue Damage and Repair.

Innate immune receptors, well known mediators of response to non-self-molecules and inflammation, also act as mediators of immunity triggered by 'damage-associated molecular patterns' (DAMPs). Pathogen-associated molecular patterns (PAMPs) cause inflammation in mammals and a rapid immune response in plants, while DAMPs trigger more complex responses, including immunity, tissue maintenance and repair. DAMPs, their receptors and downstream transduction mechanisms are often conserved within a kingdom or, due to convergent evolution, are similar across the kingdoms of life. Herein, we describe the dynamics and functionality of specific extracellular DAMP classes and their receptors in immunity, inflammation and repair of tissue damage in plants and mammals.

Cell wall traits that influence plant development, immunity, and bioconversion.

The architecture of the plant cell wall is highly dynamic, being substantially re-modeled during growth and development. Cell walls determine the size and shape of cells and contribute to the functional specialization of tissues and organs. Beyond the physiological dynamics, the wall structure undergoes changes upon biotic or abiotic stresses. In this review several cell wall traits, mainly related to pectin, one of the major matrix components, will be discussed in relation to plant development, immunity and industrial bioconversion of biomass, especially for energy production. Plant cell walls are a source of oligosaccharide fragments with a signaling function for both development and immunity. Sensing cell wall damage, sometimes through the perception of released damage-associated molecular patterns (DAMPs), is crucial for some developmental and immunity responses. Methodological advances that are expected to deepen our knowledge of cell wall (CW) biology will also be presented.

Plant immunity triggered by engineered in vivo release of oligogalacturonides, damage-associated molecular patterns.

Oligogalacturonides (OGs) are fragments of pectin that activate plant innate immunity by functioning as damage-associated molecular patterns (DAMPs). We set out to test the hypothesis that OGs are generated in planta by partial inhibition of pathogen-encoded polygalacturonases (PGs). A gene encoding a fungal PG was fused with a gene encoding a plant polygalacturonase-inhibiting protein (PGIP) and expressed in transgenic Arabidopsis plants. We show that expression of the PGIP-PG chimera results in the in vivo production of OGs that can be detected by mass spectrometric analysis. Transgenic plants expressing the chimera under control of a pathogen-inducible promoter are more resistant to the phytopathogens Botrytis cinerea, Pectobacterium carotovorum, and Pseudomonas syringae. These data provide strong evidence for the hypothesis that OGs released in vivo act as a DAMP signal to trigger plant immunity and suggest that controlled release of these molecules upon infection may be a valuable tool to protect plants against infectious diseases. On the other hand, elevated levels of expression of the chimera cause the accumulation of salicylic acid, reduced growth, and eventually lead to plant death, consistent with the current notion that trade-off occurs between growth and defense.

Mixed-Mode Operation of Hybrid Phase-Change Nanophotonic Circuits.

Phase change materials (PCMs) are highly attractive for nonvolatile electrical and all-optical memory applications because of unique features such as ultrafast and reversible phase transitions, long-term endurance, and high scalability to nanoscale dimensions. Understanding their transient characteristics upon phase transition in both the electrical and the optical domains is essential for using PCMs in future multifunctional optoelectronic circuits. Here, we use a PCM nanowire embedded into a nanophotonic circuit to study switching dynamics in mixed-mode operation. Evanescent coupling between light traveling along waveguides and a phase-change nanowire enables reversible phase transition between amorphous and crystalline states. We perform time-resolved measurements of the transient change in both the optical transmission and resistance of the nanowire and show reversible switching operations in both the optical and the electrical domains. Our results pave the way toward on-chip multifunctional optoelectronic integrated devices, waveguide integrated memories, and hybrid processing applications.

Hot-spot relaxation time current dependence in niobium nitride waveguide-integrated superconducting nanowire single-photon detectors.

We investigate how the bias current affects the hot-spot relaxation dynamics in niobium nitride. We use for this purpose a near-infrared pump-probe technique on a waveguide-integrated superconducting nanowire single-photon detector driven in the two-photon regime. We observe a strong increase in the picosecond relaxation time for higher bias currents. A minimum relaxation time of (22 ± 1) ps is obtained when applying a bias current of 50% of the switching current at 1.7 K bath temperature. We also propose a practical approach to accurately estimate the photon detection regimes based on the reconstruction of the measured detector tomography at different bias currents and for different illumination conditions.

Cavity-Enhanced and Ultrafast Superconducting Single-Photon Detectors.

Ultrafast single-photon detectors with high efficiency are of utmost importance for many applications in the context of integrated quantum photonic circuits. Detectors based on superconductor nanowires attached to optical waveguides are particularly appealing for this purpose. However, their speed is limited because the required high absorption efficiency necessitates long nanowires deposited on top of the waveguide. This enhances the kinetic inductance and makes the detectors slow. Here, we solve this problem by aligning the nanowire, contrary to usual choice, perpendicular to the waveguide to realize devices with a length below 1 µm. By integrating the nanowire into a photonic crystal cavity, we recover high absorption efficiency, thus enhancing the detection efficiency by more than an order of magnitude. Our cavity enhanced superconducting nanowire detectors are fully embedded in silicon nanophotonic circuits and efficiently detect single photons at telecom wavelengths. The detectors possess subnanosecond decay (∼120 ps) and recovery times (∼510 ps) and thus show potential for GHz count rates at low timing jitter (∼32 ps). The small absorption volume allows efficient threshold multiphoton detection.

The Arabidopsis Class III Peroxidase AtPRX71 Negatively Regulates Growth under Physiological Conditions and in Response to Cell Wall Damage.

The structure of the cell wall has a major impact on plant growth and development, and alteration of cell wall structural components is often detrimental to biomass production. However, the molecular mechanisms responsible for these negative effects are largely unknown. Arabidopsis (Arabidopsis thaliana) plants with altered pectin composition because of either the expression of the Aspergillus niger polygalacturonase II (AnPGII; 35S:AnPGII plants) or a mutation in the QUASIMODO2 (QUA2) gene that encodes a putative pectin methyltransferase (qua2-1 plants), display severe growth defects. Here, we show that expression of Arabidopsis PEROXIDASE71 (AtPRX71), encoding a class III peroxidase, strongly increases in 35S:AnPGII and qua2-1 plants as well as in response to treatments with the cellulose synthase inhibitor isoxaben, which also impairs cell wall integrity. Analysis of atprx71 loss-of-function mutants and plants overexpressing AtPRX71 indicates that this gene negatively influences Arabidopsis growth at different stages of development, likely limiting cell expansion. The atprx71-1 mutation partially suppresses the dwarf phenotype of qua2-1, suggesting that AtPRX71 contributes to the growth defects observed in plants undergoing cell wall damage. Furthermore, AtPRX71 seems to promote the production of reactive oxygen species in qua2-1 plants as well as plants treated with isoxaben. We propose that AtPRX71 contributes to strengthen cell walls, therefore restricting cell expansion, during normal growth and in response to cell wall damage.

The Arabidopsis LYSIN MOTIF-CONTAINING RECEPTOR-LIKE KINASE3 regulates the cross talk between immunity and abscisic acid responses.

Transmembrane receptor-like kinases characterized by the presence of one or more lysin motif (LysM) domains in the extracytoplasmic portion (LysM-containing receptor-like kinases [LYKs]) mediate recognition of symbiotic and pathogenic microorganisms in plants. The Arabidopsis (Arabidopsis thaliana) genome encodes five putative LYKs; among them, AtLYK1/CHITIN ELICITOR RECEPTOR KINASE1 is required for response to chitin and peptidoglycan, and AtLYK4 contributes to chitin perception. More recently, AtLYK3 has been shown to be required for full repression, mediated by Nod factors, of Arabidopsis innate immune responses. In this work, we show that AtLYK3 also negatively regulates basal expression of defense genes and resistance to Botrytis cinerea and Pectobacterium carotovorum infection. Enhanced resistance of atlyk3 mutants requires PHYTOALEXIN-DEFICIENT3, which is crucial for camalexin biosynthesis. The expression of AtLYK3 is strongly repressed by elicitors and fungal infection and is induced by the hormone abscisic acid (ABA), which has a negative impact on resistance against B. cinerea and P. carotovorum. Plants lacking a functional AtLYK3 also show reduced physiological responses to ABA and are partially resistant to ABA-induced inhibition of PHYTOALEXIN-DEFICIENT3 expression. These results indicate that AtLYK3 is important for the cross talk between signaling pathways activated by ABA and pathogens.

Engineering the cell wall by reducing de-methyl-esterified homogalacturonan improves saccharification of plant tissues for bioconversion.

Plant cell walls represent an abundant, renewable source of biofuel and other useful products. The major bottleneck for the industrial scale-up of their conversion to simple sugars (saccharification), to be subsequently converted by microorganisms into ethanol or other products, is their recalcitrance to enzymatic saccharification. We investigated whether the structure of pectin that embeds the cellulose-hemicellulose network affects the exposure of cellulose to enzymes and consequently the process of saccharification. Reduction of de-methyl-esterified homogalacturonan (HGA) in Arabidopsis plants through the expression of a fungal polygalacturonase (PG) or an inhibitor of pectin methylesterase (PMEI) increased the efficiency of enzymatic saccharification. The improved enzymatic saccharification efficiency observed in transformed plants could also reduce the need for acid pretreatment. Similar results were obtained in PG-expressing tobacco plants and in PMEI-expressing wheat plants, indicating that reduction of de-methyl-esterified HGA may be used in crop species to facilitate the process of biomass saccharification.

Arabidopsis MPK3 and MPK6 play different roles in basal and oligogalacturonide- or flagellin-induced resistance against Botrytis cinerea.

Mitogen-activated protein kinases (MAPKs) are fundamental components of the plant innate immune system. MPK3 and MPK6 are Arabidopsis (Arabidopsis thaliana) MAPKs activated by pathogens and elicitors such as oligogalacturonides (OGs), which function as damage-associated molecular patterns, and flg22, a well-known microbe-associated molecular pattern. However, the specific contribution of MPK3 and MPK6 to the regulation of elicitor-induced defense responses is not completely defined. In this work we have investigated the roles played by these MAPKs in elicitor-induced resistance against the fungal pathogen Botrytis cinerea. Analysis of single mapk mutants revealed that lack of MPK3 increases basal susceptibility to the fungus, as previously reported, but does not significantly affect elicitor-induced resistance. Instead, lack of MPK6 has no effect on basal resistance but suppresses OG- and flg22-induced resistance to B. cinerea. Overexpression of the AP2C1 phosphatase leads to impaired OG- and flg22-induced phosphorylation of both MPK3 and MPK6, and to phenotypes that recapitulate those of the single mapk mutants. These data indicate that OG- and flg22-induced defense responses effective against B. cinerea are mainly dependent on MAPKs, with a greater contribution of MPK6.

Oligogalacturonide-auxin antagonism does not require posttranscriptional gene silencing or stabilization of auxin response repressors in Arabidopsis.

α-1-4-Linked oligogalacturonides (OGs) derived from plant cell walls are a class of damage-associated molecular patterns and well-known elicitors of the plant immune response. Early transcript changes induced by OGs largely overlap those induced by flg22, a peptide derived from bacterial flagellin, a well-characterized microbe-associated molecular pattern, although responses diverge over time. OGs also regulate growth and development of plant cells and organs, due to an auxin-antagonistic activity. The molecular basis of this antagonism is still unknown. Here we show that, in Arabidopsis (Arabidopsis thaliana), OGs inhibit adventitious root formation induced by auxin in leaf explants as well as the expression of several auxin-responsive genes. Genetic, biochemical, and pharmacological experiments indicate that inhibition of auxin responses by OGs does not require ethylene, jasmonic acid, and salicylic acid signaling and is independent of RESPIRATORY BURST OXIDASE HOMOLOGUE D-mediated reactive oxygen species production. Free indole-3-acetic acid levels are not noticeably altered by OGs. Notably, OG- as well as flg22-auxin antagonism does not involve any of the following mechanisms: (1) stabilization of auxin-response repressors; (2) decreased levels of auxin receptor transcripts through the action of microRNAs. Our results suggest that OGs and flg22 antagonize auxin responses independently of Aux/Indole-3-Acetic Acid repressor stabilization and of posttranscriptional gene silencing.

Spectroscopic measurement of the refractive index of ion-implanted diamond.

We present the results of variable-angle spectroscopic ellipsometry and transmittance measurements to determine the variation of the complex refractive index of ion-implanted single-crystal diamond. An increase is found in both real and imaginary parts at increasing damage densities. The index depth variation is determined in the whole wavelength range between 250 and 1690 nm. The dependence from the vacancy density is evaluated, highlighting a deviation from linearity in the high-damage-density regime. A considerable increase (up to 5%) in the real part of the index is observed, attributed to an increase in polarizability, thus offering new microfabrication possibilities for waveguides and other photonic structures in diamond.

Transient silencing of the grapevine gene VvPGIP1 by agroinfiltration with a construct for RNA interference.

Grapevine is an economically important crop, and the recent completion of its genome makes it possible to study the function of specific genes through reverse genetics. However, the analysis of gene function by RNA interference (RNAi) in grapevine is difficult, because the generation of stable transgenic plants has low efficiency and is time consuming. Recently, transient expression of genes in grapevine leaves has been obtained by Agrobacterium tumefaciens infiltration (agroinfiltration). We therefore tested the possibility to silence grapevine genes by agroinfiltration of RNAi constructs. A construct to express a double strand RNA (dsRNA) corresponding to the defense-related gene VvPGIP1, encoding a polygalacturonase-inhibiting protein (PGIP), was obtained and transiently expressed by agroinfiltration in leaves of grapevine plants grown in vitro. Expression of VvPGIP1 and accumulation of PGIP activity were strongly induced by infiltration with control bacteria, but not with bacteria carrying the dsRNA construct, indicating that the gene was efficiently silenced. In contrast, expression of another defense-related gene, VST1, encoding a stilbene synthase, was unaffected by the dsRNA construct. We have therefore demonstrated the possibility of transient down-regulation of grapevine genes by agroinfiltration of constructs for the expression of dsRNA. This system can be employed to evaluate the effectiveness of constructs that can be subsequently used to generate stable RNAi transgenic plants.

Tryptophan-derived metabolites are required for antifungal defense in the Arabidopsis mlo2 mutant.

Arabidopsis (Arabidopsis thaliana) genes MILDEW RESISTANCE LOCUS O2 (MLO2), MLO6, and MLO12 exhibit unequal genetic redundancy with respect to the modulation of defense responses against powdery mildew fungi and the control of developmental phenotypes such as premature leaf decay. We show that early chlorosis and necrosis of rosette leaves in mlo2 mlo6 mlo12 mutants reflects an authentic but untimely leaf senescence program. Comparative transcriptional profiling revealed that transcripts of several genes encoding tryptophan biosynthetic and metabolic enzymes hyperaccumulate during vegetative development in the mlo2 mlo6 mlo12 mutant. Elevated expression levels of these genes correlate with altered steady-state levels of several indolic metabolites, including the phytoalexin camalexin and indolic glucosinolates, during development in the mlo2 single mutant and the mlo2 mlo6 mlo12 triple mutant. Results of genetic epistasis analysis suggest a decisive role for indolic metabolites in mlo2-conditioned antifungal defense against both biotrophic powdery mildews and a camalexin-sensitive strain of the necrotrophic fungus Botrytis cinerea. The wound- and pathogen-responsive callose synthase POWDERY MILDEW RESISTANCE4/GLUCAN SYNTHASE-LIKE5 was found to be responsible for the spontaneous callose deposits in mlo2 mutant plants but dispensable for mlo2-conditioned penetration resistance. Our data strengthen the notion that powdery mildew resistance of mlo2 genotypes is based on the same defense execution machinery as innate antifungal immune responses that restrict the invasion of nonadapted fungal pathogens.

Host Cell Wall Damage during Pathogen Infection: Mechanisms of Perception and Role in Plant-Pathogen Interactions.

The plant cell wall (CW) is a complex structure that acts as a mechanical barrier, restricting the access to most microbes. Phytopathogenic microorganisms can deploy an arsenal of CW-degrading enzymes (CWDEs) that are required for virulence. In turn, plants have evolved proteins able to inhibit the activity of specific microbial CWDEs, reducing CW damage and favoring the accumulation of CW-derived fragments that act as damage-associated molecular patterns (DAMPs) and trigger an immune response in the host. CW-derived DAMPs might be a component of the complex system of surveillance of CW integrity (CWI), that plants have evolved to detect changes in CW properties. Microbial CWDEs can activate the plant CWI maintenance system and induce compensatory responses to reinforce CWs during infection. Recent evidence indicates that the CWI surveillance system interacts in a complex way with the innate immune system to fine-tune downstream responses and strike a balance between defense and growth.