Integration of phot1, phot2, and PhyB signalling in light-induced chloroplast movements

Abstract In Arabidopsis thaliana, chloroplasts move towards the periclinal cell walls upon exposure to low blue light intensities and to anticlinal walls under high light. The regulation of these chloroplast movements involves members of both the phototropin and phytochrome families of photoreceptors. Examination of fluence-rate response dependencies in phot1 and phot2 mutants revealed that although both photoreceptors are capable of inducing chloroplast accumulation under low-light conditions, the signals from these photoreceptors appear to be antagonistic. Chloroplast movements in wild-type plants were intermediate between those of the single phot mutants, consistent with each operating through separate signalling cascades. Mutants in phot2 showed transient chloroplast avoidance responses upon exposure to intense blue light, and slow but sustained chloroplast avoidance under intense white light, indicating that in the absence of phot2, phot1 is capable of generating both a low and a high-light response signal. Mutations in phytochrome B (phyB) caused an enhanced avoidance response at intermediate and high light intensities. Examination of phyB, phot1phyB, and phot2phyB mutants indicated that this enhancement is caused by PhyB inhibition of the high-light avoidance response in wild-type plants. In addition, our results suggest that the inhibition by PhyB is not exclusive to either of the phot1 or phot2 signalling pathways

Evolution and Expression of Tandem Duplicated Maize Flavonol Synthase Genes

Flavonoids are specialized compounds widely distributed and with diverse functions throughout the plant kingdom and with several benefits for human health. In particular, flavonols, synthesized by flavonol synthase (FLS), protect plants against UV-B radiation and are essential for male fertility in maize and other plants. We have recently characterized a UV-B inducible ZmFLS1, corresponding to the first to be described in monocot plants. Interestingly, the new assembly of the B73 maize genome revealed the presence of a second putative FLS gene (ZmFLS2), with very high identity with ZmFLS1. ZmFLSs expression was analyzed in different maize tissues, and by combining electrophoretic mobility shift assays and transient expression experiments, we show that both genes are direct targets of anthocyanin (C1/PL1 + R/B) and 3-deoxy flavonoid (P1) transcriptional regulators. ZmFLS expression analyses show higher levels of both transcripts in high altitude landraces than inbred lines, and both genes are regulated by UV-B radiation in all lines analyzed. Moreover, the high sequence conservation of the ZmFLS promoters between maize lines suggests that the differences observed in ZmFLS expression are due to allelic variations in the transcription factors that regulate their activities. Finally, we generated pFLS1::FLS1-RFP transgenic plants and analyzed ZmFLS1 expression in different maize tissues; we found that this enzyme is localized in the ER and the perinuclear region

Identification of protein interactions of grapevine fanleaf virus RNA-dependent RNA polymerase during infection of by affinity purification and tandem mass spectrometry.

The RNA-dependent RNA polymerase (1E) is involved in replication of grapevine fanleaf virus (GFLV, , ) and causes vein clearing symptoms in . Information on protein 1E interaction with other viral and host proteins is scarce. To study protein 1E biology, three GFLV infectious clones, i.e. GHu (a symptomatic wild-type strain), GHu-1E (an asymptomatic GHu mutant) and F13 (an asymptomatic wild-type strain), were engineered with protein 1E fused to a V5 epitope tag at the C-terminus. Following -mediated delivery of GFLV clones in and protein extraction at seven dpi, when optimal 1E:V5 accumulation was detected, two viral and six plant putative interaction partners of V5-tagged protein 1E were identified for the three GFLV clones by affinity purification and tandem mass spectrometry. This study provides insights into the protein interactome of 1E during GFLV systemic infection in and lays the foundation for validation work

Plastid Movement Impaired 2, a New Gene Involved in Normal Blue-Light-Induced Chloroplast Movements in Arabidopsis

Chloroplasts move in a light-dependent manner that can modulate the photosynthetic potential of plant cells. Identification of genes required for light-induced chloroplast movement is beginning to define the molecular machinery that controls these movements. In this work, we describe plastid movement impaired 2 (pmi2), a mutant in Arabidopsis (Arabidopsis thaliana) that displays attenuated chloroplast movements under intermediate and high light intensities while maintaining a normal movement response under low light intensities. In wild-type plants, fluence rates below 20 μmol m(−2) s(−1) of blue light lead to chloroplast accumulation on the periclinal cell walls, whereas light intensities over 20 μmol m(−2) s(−1) caused chloroplasts to move toward the anticlinal cell walls (avoidance response). However, at light intensities below 75 μmol m(−2) s(−1), chloroplasts in pmi2 leaves move to the periclinal walls; 100 μmol m(−2) s(−1) of blue light is required for chloroplasts in pmi2 to move to the anticlinal cell walls, indicating a shift in the light threshold for the avoidance response in the mutant. The pmi2 mutation has been mapped to a gene that encodes a protein of unknown function with a large coiled-coil domain in the N terminus and a putative P loop. PMI2 shares sequence and structural similarity with PMI15, another unknown protein in Arabidopsis that, when mutated, causes a defect in chloroplast avoidance under high-light intensities

A Plant-Specific Protein Essential for Blue-Light-Induced Chloroplast Movements

In Arabidopsis (Arabidopsis thaliana), light-dependent chloroplast movements are induced by blue light. When exposed to low fluence rates of light, chloroplasts accumulate in periclinal layers perpendicular to the direction of light, presumably to optimize light absorption by exposing more chloroplast area to the light. Under high light conditions, chloroplasts become positioned parallel to the incoming light in a response that can reduce exposure to light intensities that may damage the photosynthetic machinery. To identify components of the pathway downstream of the photoreceptors that mediate chloroplast movements (i.e. phototropins), we conducted a mutant screen that has led to the isolation of several Arabidopsis mutants displaying altered chloroplast movements. The plastid movement impaired1 (pmi1) mutant exhibits severely attenuated chloroplast movements under all tested fluence rates of light, suggesting that it is a necessary component for both the low- and high-light-dependant chloroplast movement responses. Analysis of pmi1 leaf cross sections revealed that regardless of the light condition, chloroplasts are more evenly distributed in leaf mesophyll cells than in the wild type. The pmi1-1 mutant was found to contain a single nonsense mutation within the open reading frame of At1g42550. This gene encodes a plant-specific protein of unknown function that appears to be conserved among angiosperms. Sequence analysis of the protein suggests that it may be involved in calcium-mediated signal transduction, possibly through protein–protein interactions

Insights in luteovirid structural biology guided by chemical cross-linking and high resolution mass spectrometry

Interactions among plant pathogenic viruses in the family Luteoviridae and their plant hosts and insect vectors are governed by the topology of the viral capsid, which is the sole vehicle for long distance movement of the viral genome. Previous application of a mass spectrometry-compatible cross-linker to preparations of the luteovirid Potato leafroll virus (PLRV; Luteoviridae: Polerovirus) revealed a detailed network of interactions between viral structural proteins and enabled generation of the first cross-linking guided coat protein models. In this study, we extended application of chemical cross-linking technology to the related Turnip yellows virus (TuYV; Luteoviridae: Polerovirus). Remarkably, all cross-links found between sites in the viral coat protein found for TuYV were also found in PLRV. Guided by these data, we present two models for the TuYV coat protein trimer, the basic structural unit of luteovirid virions. Additional cross-links found between the TuYV coat protein and a site in the viral protease domain suggest a possible role for the luteovirid protease in regulating the structural biology of these viruses

A Conserved Mechanism of Bract Suppression in the Grass Family[W][OA]

Bract suppression in maize, rice, and barley is regulated by a conserved genetic mechanism. Interestingly, the orthologous gene in Arabidopsis has no role in bract suppression, suggesting distinct bract suppression mechanisms have evolved in these two lineages

Profiling Plant Proteome and Transcriptome Changes during Grapevine Fanleaf Virus Infection

Viruses can elicit varying types and severities of symptoms during plant host infection. We investigated changes in the proteome and transcriptome of Nicotiana benthamiana plants infected by grapevine fanleaf virus (GFLV) with an emphasis on vein clearing symptom development. Comparative, time-course liquid chromatography tandem mass spectrometry and 3′ ribonucleic acid sequencing analyses of plants infected by two wildtype GFLV strains, one symptomatic and one asymptomatic, and their asymptomatic mutant strains carrying a single amino acid change in the RNA-dependent RNA polymerase (RdRP) were conducted to identify host biochemical pathways involved in viral symptom development. During peak vein clearing symptom display at 7 days post-inoculation (dpi), protein and gene ontologies related to immune response, gene regulation, and secondary metabolite production were overrepresented when contrasting wildtype GFLV strain GHu and mutant GHu-1EK802GPol. Prior to the onset of symptom development at 4 dpi and when symptoms faded away at 12 dpi, protein and gene ontologies related to chitinase activity, hypersensitive response, and transcriptional regulation were identified. This systems biology approach highlighted how a single amino acid of a plant viral RdRP mediates changes to the host proteome (∼1%) and transcriptome (∼8.5%) related to transient vein clearing symptoms and the network of pathways involved in the virus–host arms race

Profiling Plant Proteome and Transcriptome Changes during Grapevine Fanleaf Virus Infection

Viruses can elicit varying types and severities of symptoms during plant host infection. We investigated changes in the proteome and transcriptome of Nicotiana benthamiana plants infected by grapevine fanleaf virus (GFLV) with an emphasis on vein clearing symptom development. Comparative, time-course liquid chromatography tandem mass spectrometry and 3′ ribonucleic acid sequencing analyses of plants infected by two wildtype GFLV strains, one symptomatic and one asymptomatic, and their asymptomatic mutant strains carrying a single amino acid change in the RNA-dependent RNA polymerase (RdRP) were conducted to identify host biochemical pathways involved in viral symptom development. During peak vein clearing symptom display at 7 days post-inoculation (dpi), protein and gene ontologies related to immune response, gene regulation, and secondary metabolite production were overrepresented when contrasting wildtype GFLV strain GHu and mutant GHu-1EK802GPol. Prior to the onset of symptom development at 4 dpi and when symptoms faded away at 12 dpi, protein and gene ontologies related to chitinase activity, hypersensitive response, and transcriptional regulation were identified. This systems biology approach highlighted how a single amino acid of a plant viral RdRP mediates changes to the host proteome (∼1%) and transcriptome (∼8.5%) related to transient vein clearing symptoms and the network of pathways involved in the virus–host arms race