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Antagonistic actions of FPA and IBM2 regulate transcript processing from genes containing heterochromatin. Plant Physiol. 2019 Feb 27

Antagonistic actions of FPA and IBM2 regulate transcript processing from genes containing heterochromatin. Plant Physiol. 2019 Feb 27 | Publications @IJPB | Scoop.it

Repressive epigenetic marks, such as DNA and histone methylation, are sometimes located within introns. In Arabidopsis (Arabidopsis thaliana), INCREASE IN BONSAI METHYLATION2 (IBM2), an RNA-binding protein containing a BAH domain, is required to process functional transcript isoforms of genes carrying intronic heterochromatin. In a genetic screen for suppressors of the ibm2 mutation, we identified FPA, an RNA-binding protein which promotes use of proximal polyadenylation sites in genes targeted by IBM2, including IBM1 encoding an essential H3K9 histone demethylase and the disease resistance gene RECOGNITION OF PERONOSPORA PARASITICA7 (RPP7). Both IBM2 and FPA are involved in the processing of their common mRNA targets: transcription of IBM2 target genes is restored when FPA is mutated in ibm2 and impaired in transgenic plants over-expressing FPA. By contrast, transposons targeted by IBM2 and localised outside introns are not under this antagonistic control. The DNA methylation patterns of some genes and transposons are modified in fpa plants, including the large intron of IBM1, but these changes are rather limited and reversed when the mutant is complemented, indicating that FPA has a restricted role in mediating silencing. These data reveal a complex regulation by IBM2 and FPA pathways in processing mRNAs of genes bearing heterochromatic marks.

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VarEpi IJPB team

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Modelling Sex-Specific Crossover Patterning in Arabidopsis. Genetics. 2019 Jan 22

Modelling Sex-Specific Crossover Patterning in Arabidopsis. Genetics. 2019 Jan 22 | Publications @IJPB | Scoop.it

nterference is a major force governing the patterning of meiotic crossovers. A leading model describing how interference influences crossover-patterning is the beam film model, a mechanical model based on the accumulation and redistribution of crossover-promoting stress along the chromosome axis. We use the beam-film model in conjunction with a large Arabidopsis reciprocal back-cross data set to gain mechanistic insights into the differences between male and female meiosis and crossover patterning. Beam-film modelling suggests that the underlying mechanics of crossover patterning and interference are identical in the two sexes, with the large difference in recombination rates and distributions able to be entirely explained by the shorter chromosome axes in females. The modelling supports previous indications that fewer crossovers occur via the class II pathway in female meiosis and that this could be explained by reduced DNA double strand breaks in female meiosis, paralleling the observed reduction in synaptonemal complex length between the two sexes. We also demonstrate that changes in the strength of suppression of neighboring class I crossovers can have opposite effects on effective interference depending on the distance between two genetic intervals.

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POLYMEIO IJPB team

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Clone-Dependent Expression of Esca Disease Revealed by Leaf Metabolite Analysis. Front Plant Sci. 2019 Jan 9

Clone-Dependent Expression of Esca Disease Revealed by Leaf Metabolite Analysis. Front Plant Sci. 2019 Jan 9 | Publications @IJPB | Scoop.it

Grapevine trutk diseases, especially Esca, are of major concern since they gradually alter vineyards worldwide and cause heavy economic losses. The expression of Esca disease symptoms depends on several factors, including the grapevine cultivar. In this context, a possible clone-dependent expression of the Esca disease was studied. Two clones of 'Chardonnay' grown in the same plot were compared according to their developmental and physiological traits, metabolome, and foliar symptom expression. Analysis of their leaf metabolome highlighted differences related to symptom expression. Interestingly, the content of a few specific metabolites exhibited opposite variations in leaves of symptomatic shoots of clones 76 and 95. Altogether this study showed a clone-dependent expression of Esca disease in 'Chardonnay' and the relevance of GC-MS and 3D fluorescence methods to analyze the impact of the disease on the leaf metabolome.

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ACCI IJPB team

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Efficient and transgene-free gene targeting using Agrobacterium-mediated delivery of the CRISPR/Cas9 system in tomato. Plant Cell Rep. 2019 Jan 16

Efficient and transgene-free gene targeting using Agrobacterium-mediated delivery of the CRISPR/Cas9 system in tomato. Plant Cell Rep. 2019 Jan 16 | Publications @IJPB | Scoop.it
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DRAGON IJPB team

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Novel loci and a role for nitric oxide for seed dormancy and preharvest sprouting in barley. Plant Cell Environ. 2018 Nov 22.

Novel loci and a role for nitric oxide for seed dormancy and preharvest sprouting in barley. Plant Cell Environ. 2018 Nov 22. | Publications @IJPB | Scoop.it

Barley is used for food and feed, and brewing. Nondormant seeds are required for malting, but the lack of dormancy can lead to preharvest sprouting (PHS), which is also undesired. Here, we report several new loci that modulate barley seed dormancy and PHS. Using genome-wide association mapping of 184 spring barley genotypes, we identified four new, highly significant associations on chromosomes 1H, 3H, and 5H previously not associated with barley seed dormancy or PHS. A total of 71 responsible genes were found mostly related to flowering time and hormone signalling. A homolog of the well-known Arabidopsis Delay of Germination 1 (DOG1) gene was annotated on the barley chromosome 3H. Unexpectedly, DOG1 appears to play only a minor role in barley seed dormancy. However, the gibberellin oxidase gene HvGA20ox1 contributed to dormancy alleviation, and another seven important loci changed significantly during after-ripening. Furthermore, nitric oxide release correlated negatively with dormancy and shared 27 associations. Origin and growth environment affected seed dormancy and PHS more than did agronomic traits. Days to anthesis and maturity were shorter when seeds were produced under drier conditions, seeds were less dormant, and PHS increased, with a heritability of 0.57-0.80. The results are expected to be useful for crop improvement.

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PHYGERM IJPB team

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Assessing the response of small RNA populations to allopolyploidy using resynthesized Brassica napus allotetraploids. Mol Biol Evol. 2019 Jan 17

Assessing the response of small RNA populations to allopolyploidy using resynthesized Brassica napus allotetraploids. Mol Biol Evol. 2019 Jan 17 | Publications @IJPB | Scoop.it

Allopolyploidy, combining interspecific hybridization with whole genome duplication, has had significant impact on plant evolution. Its evolutionary success is related to the rapid and profound genome reorganizations that allow neo-allopolyploids to form and adapt. Nevertheless, how neo-allopolyploid genomes adapt to regulate their expression remains poorly understood. The hypothesis of a major role for small non-coding RNAs (sRNAs) in mediating the transcriptional response of neo-allopolyploid genomes has progressively emerged. Generally, 21-nt sRNAs mediate post-transcriptional gene silencing (PTGS) by mRNA cleavage whereas 24-nt sRNAs repress transcription (transcriptional gene silencing, TGS) through epigenetic modifications. Here, we characterize the global response of sRNAs to allopolyploidy in Brassica, using three independently resynthesized B. napus allotetraploids originating from crosses between diploid B. oleracea and B. rapa accessions, surveyed at two different generations in comparison with their diploid progenitors. Our results suggest an immediate but transient response of specific sRNA populations to allopolyploidy. These sRNA populations mainly target non-coding components of the genome but also target the transcriptional regulation of genes involved in response to stresses and in metabolism; this suggests a broad role in adapting to allopolyploidy. We finally identify the early accumulation of both 21- and 24-nt sRNAs involved in regulating the same targets, supporting a PTGS-to-TGS shift at the first stages of the neo-allopolyploid formation. We propose that reorganization of sRNA production is an early response to allopolyploidy in order to control the transcriptional reactivation of various non-coding elements and stress-related genes, thus ensuring genome stability during the first steps of neo-allopolyploid formation.

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POLYMEIO IJPB team

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POLQ plays a key role in repair of CRISPR/Cas9 induced double strand breaks in the moss Physcomitrella patens. - New Phytol. 2019 Jan 13

POLQ plays a key role in repair of CRISPR/Cas9 induced double strand breaks in the moss Physcomitrella patens. - New Phytol. 2019 Jan 13 | Publications @IJPB | Scoop.it

Double strand breaks can be repaired by different mechanisms such as homologous recombination (HR), classical non-homologous end joining (C-NHEJ) and alternative end joining (Alt-EJ). The polymerase Q (POLQ) has been proposed to be the main factor involved in Alt-EJ mediated DNA repair. •Here we describe the role of POLQ in DNA repair and gene targeting in Physcomitrella patens. The disruption of the POLQ gene does not influence the genetic stability of P. patens nor its development. •The polq mutant shows the same sensitivity as wild-type towards most of the genotoxic agents tested (UV, MMS and cisplatin) with the notable exception of bleomycin for which it shows less sensitivity than the wild-type. Furthermore, we show that POLQ is involved in the repair of CRISPR-Cas9 induced double strand breaks in P. patens. We also demonstrate that POLQ is a potential competitor and/or inhibitor of the HR repair pathway. •This has a consequence in terms of genetic engineering as in absence of POLQ the frequency of gene targeting is significantly increased and the number of clean two-sided HR-mediated insertions is enhanced. Thus, controlling POLQ activity in plants could be a useful strategy to optimize the tools of genome engineering for plant breeding.

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DRAGON IJPB team

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Clonal seeds from hybrid rice by simultaneous genome engineering of meiosis and fertilization genes. Nat Biotechnol. 2019 Jan 4.

Clonal seeds from hybrid rice by simultaneous genome engineering of meiosis and fertilization genes. Nat Biotechnol. 2019 Jan 4. | Publications @IJPB | Scoop.it

Heterosis, or hybrid vigor, is exploited by breeders to produce elite high-yielding crop lines, but beneficial phenotypes are lost in subsequent generations owing to genetic segregation. Clonal propagation through seeds would enable self-propagation of F1 hybrids. Here we report a strategy to enable clonal reproduction of F1 rice hybrids through seeds. We fixed the heterozygosity of F1 hybrid rice by multiplex CRISPR-Cas9 genome editing of the REC8, PAIR1 and OSD1 meiotic genes to produce clonal diploid gametes and tetraploid seeds. Next, we demonstrated that editing the MATRILINEAL (MTL) gene (involved in fertilization) could induce formation of haploid seeds in hybrid rice. Finally, we combined fixation of heterozygosity and haploid induction by simultaneous editing of all four genes (REC8, PAIR1, OSD1 and MTL) in hybrid rice and obtained plants that could propagate clonally through seeds. Application of our method may enable self-propagation of a broad range of elite F1 hybrid crops.

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MeioMe IJPB team

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A male-expressed rice embryogenic trigger redirected for asexual propagation through seeds. Nature. 2018 Dec 12

A male-expressed rice embryogenic trigger redirected for asexual propagation through seeds. Nature. 2018 Dec 12 | Publications @IJPB | Scoop.it

The molecular pathways that trigger the initiation of embryogenesis after fertilization in flowering plants, and prevent its occurrence without fertilization, are not well understood. Here we show in rice (Oryza sativa) that BABY BOOM1 (BBM1), a member of the AP2 family of transcription factors that is expressed in sperm cells, has a key role in this process. Ectopic expression of BBM1 in the egg cell is sufficient for parthenogenesis, which indicates that a single wild-type gene can bypass the fertilization checkpoint in the female gamete. Zygotic expression of BBM1 is initially specific to the male allele but is subsequently biparental, and this is consistent with its observed auto-activation. Triple knockout of the genes BBM1, BBM2 and BBM3 causes embryo arrest and abortion, which are fully rescued by male-transmitted BBM1. These findings suggest that the requirement for fertilization in embryogenesis is mediated by male-genome transmission of pluripotency factors. When genome editing to substitute mitosis for meiosis (MiMe) is combined with the expression of BBM1 in the egg cell, clonal progeny can be obtained that retain genome-wide parental heterozygosity. The synthetic asexual-propagation trait is heritable through multiple generations of clones. Hybrid crops provide increased yields that cannot be maintained by their progeny owing to genetic segregation. This work establishes the feasibility of asexual reproduction in crops, and could enable the maintenance of hybrids clonally through seed propagation.

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MeioMe IJPB team

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Europe's first and last field trial of gene-edited plants? - Elife. 2018 Dec 18

Europe's first and last field trial of gene-edited plants? - Elife. 2018 Dec 18 | Publications @IJPB | Scoop.it

On 5 June this year the first field trial of a CRISPR-Cas-9 gene-edited crop began at Rothamsted Research in the UK, having been approved by the UK Department for Environment, Food & Rural Affairs. However, in late July 2018, after the trial had started, the European Court of Justice ruled that techniques such as gene editing fall within the European Union's 2001 GMO directive, meaning that our gene-edited Camelina plants should be considered as genetically modified (GM). Here we describe our experience of running this trial and the legal transformation of our plants. We also consider the future of European plant research using gene-editing techniques, which now fall under the burden of GM regulation, and how this will likely impede translation of publicly funded basic research.

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DIPOL IJPB team

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The DFR locus: A smart landing pad for targeted transgene insertion in tomato. PLoS One. 2018 Dec 6

The DFR locus: A smart landing pad for targeted transgene insertion in tomato. PLoS One. 2018 Dec 6 | Publications @IJPB | Scoop.it

Targeted insertion of transgenes in plants is still challenging and requires further technical innovation. In the present study, we chose the tomato DFR gene involved in anthocyanin biosynthesis as a landing pad for targeted transgene insertion using CRISPR-Cas9 in a two-step strategy. First, a 1013 bp was deleted in the endogenous DFR gene. Hypocotyls and callus of in vitro regenerated plantlets homozygous for the deletion were green instead of the usual anthocyanin produced purple colour. Next, standard Agrobacterium-mediated transformation was used to target transgene insertion at the DFR landing pad in the dfr deletion line. The single binary vector carried two sgRNAs, a donor template containing two homology arms of 400 bp, the previously deleted DFR sequence, and a NptII expression cassette. Regenerating plantlets were screened for a purple-colour phenotype indicating that DFR function had been restored. Targeted insertions were identified in 1.29% of the transformed explants. Thus, we established an efficient method for selecting HDR-mediated transgene insertion using the CRISPR-Cas9 system in tomato. The visual screen used here facilitates selection of these rare gene targeting events, does not necessitate the systematic PCR screening of all the regenerating material and can be potentially applied to other crops.

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Macroscale fluorescence imaging against autofluorescence under ambient light. Light Sci Appl. 2018 Nov 28

Macroscale fluorescence imaging against autofluorescence under ambient light. Light Sci Appl. 2018 Nov 28 | Publications @IJPB | Scoop.it

Macroscale fluorescence imaging is increasingly used to observe biological samples. However, it may suffer from spectral interferences that originate from ambient light or autofluorescence of the sample or its support. In this manuscript, we built a simple and inexpensive fluorescence macroscope, which has been used to evaluate the performance of Speed OPIOM (Out of Phase Imaging after Optical Modulation), which is a reference-free dynamic contrast protocol, to selectively image reversibly photoswitchable fluorophores as labels against detrimental autofluorescence and ambient light. By tuning the intensity and radial frequency of the modulated illumination to the Speed OPIOM resonance and adopting a phase-sensitive detection scheme that ensures noise rejection, we enhanced the sensitivity and the signal-to-noise ratio for fluorescence detection in blot assays by factors of 50 and 10, respectively, over direct fluorescence observation under constant illumination. Then, we overcame the strong autofluorescence of growth media that are currently used in microbiology and realized multiplexed fluorescence observation of colonies of spectrally similar fluorescent bacteria with a unique configuration of excitation and emission wavelengths. Finally, we easily discriminated fluorescent labels from the autofluorescent and reflective background in labeled leaves, even under the interference of incident light at intensities that are comparable to sunlight. The proposed approach is expected to find multiple applications, from biological assays to outdoor observations, in fluorescence macroimaging.

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DIPOL IJPB team

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Arabidopsis thaliana DGAT3 is a [2Fe-2S] protein involved in TAG biosynthesis - Scientific Reports 22 Nov 2018

Arabidopsis thaliana DGAT3 is a [2Fe-2S] protein involved in TAG biosynthesis - Scientific Reports 22 Nov 2018 | Publications @IJPB | Scoop.it

Acyl-CoA:diacylglycerol acyltransferases 3 (DGAT3) are described as plant cytosolic enzymes synthesizing triacylglycerol. Their protein sequences exhibit a thioredoxin-like ferredoxin domain typical of a class of ferredoxins harboring a [2Fe-2S] cluster. The Arabidopsis thaliana DGAT3 (AtDGAT3; At1g48300) protein is detected in germinating seeds. The recombinant purified protein produced from Escherichia coli, although very unstable, exhibits DGAT activity in vitro. A shorter protein version devoid of its N-terminal putative chloroplast transit peptide, Δ46AtDGAT3, was more stable in vitro, allowing biochemical and spectroscopic characterization. The results obtained demonstrate the presence of a [2Fe-2S] cluster in the protein. To date, AtDGAT3 is the first metalloprotein described as a DGAT.

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DYSCOL IJPB team

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Transgene-Free Genome Editing in Tomato and Potato Plants Using Agrobacterium-Mediated Delivery of a CRISPR/Cas9 Cytidine Base Editor. Int J Mol Sci. 2019 Jan 18

Transgene-Free Genome Editing in Tomato and Potato Plants Using Agrobacterium-Mediated Delivery of a CRISPR/Cas9 Cytidine Base Editor. Int J Mol Sci. 2019 Jan 18 | Publications @IJPB | Scoop.it

Genome editing tools have rapidly been adopted by plant scientists for gene function discovery and crop improvement. The current technical challenge is to efficiently induce precise and predictable targeted point mutations valuable for crop breeding purposes. Cytidine base editors (CBEs) are CRISPR/Cas9 derived tools recently developed to direct a C-to-T base conversion. Stable genomic integration of CRISPR/Cas9 components through Agrobacterium-mediated transformation is the most widely used approach in dicotyledonous plants. However, elimination of foreign DNA may be difficult to achieve, especially in vegetatively propagated plants. In this study, we targeted the acetolactate synthase (ALS) gene in tomato and potato by a CBE using Agrobacterium-mediated transformation. We successfully and efficiently edited the targeted cytidine bases, leading to chlorsulfuron-resistant plants with precise base edition efficiency up to 71% in tomato. More importantly, we produced 12.9% and 10% edited but transgene-free plants in the first generation in tomato and potato, respectively. Such an approach is expected to decrease deleterious effects due to the random integration of transgene(s) into the host genome. Our successful approach opens up new perspectives for genome engineering by the co-edition of the ALS with other gene(s), leading to transgene-free plants harboring new traits of interest.

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DRAGON IJPB team

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Dissecting the pathways coordinating patterning and growth by plant boundary domains. PLoS Genet. 2019 Jan 24

Dissecting the pathways coordinating patterning and growth by plant boundary domains. PLoS Genet. 2019 Jan 24 | Publications @IJPB | Scoop.it

Boundary domains play important roles during morphogenesis in plants and animals, but how they contribute to patterning and growth coordination in plants is not understood. The CUC genes determine the boundary domains in the aerial part of the plants and, in particular, they have a conserved role in regulating leaf complexity across Angiosperms. Here, we used tooth formation at the Arabidopsis leaf margin controlled by the CUC2 transcription factor to untangle intertwined events during boundary-controlled morphogenesis in plants. Combining conditional restoration of CUC2 function with morphometrics as well as quantification of gene expression and hormone signaling, we first established that tooth morphogenesis involves a patterning phase and a growth phase. These phases can be separated, as patterning requires CUC2 while growth can occur independently of CUC2. Next, we show that CUC2 acts as a trigger to promote growth through the activation of three functional relays. In particular, we show that KLUH acts downstream of CUC2 to modulate auxin response and that expressing KLUH can compensate for deficient CUC2 expression during tooth growth. Together, we reveal a genetic and molecular network that allows coordination of patterning and growth by CUC2-defined boundaries during morphogenesis at the leaf margin.

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FTA IJPB team

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A New Role for SAG12 Cysteine Protease in Roots of Arabidopsis thaliana. Front Plant Sci. 2019 Jan 11

A New Role for SAG12 Cysteine Protease in Roots of Arabidopsis thaliana. Front Plant Sci. 2019 Jan 11 | Publications @IJPB | Scoop.it

Senescence associated gene (SAG) 12, which encodes a cysteine protease is considered to be important in nitrogen (N) allocation to Arabidopsis thaliana seeds. A decrease in the yield and N content of the seeds was observed in the Arabidopsis SAG12 knockout mutants (sag12) relative to the wild type (Col0) under limited nitrogen nutrition. However, leaf senescence was similar in both lines. To test whether SAG12 is involved in N remobilization from organs other than the leaves, we tested whether root N could be used in N mobilization to the seeds. Root architecture, N uptake capacity and 15N partitioning were compared in the wild type and sag12 under either high nitrogen (HN) or low nitrogen (LN) conditions. No differences in root architecture or root N uptake capacity were observed between the lines under HN or LN. However, under LN conditions, there was an accumulation of 15N in the sag12 roots compared to the wild type with lower allocation of 15N to the seeds. This was accompanied by an increase in root N protein contents and a significant decrease in root cysteine protease activity. SAG12 is expressed in the root stele of the plants at the reproductive stage, particularly under conditions of LN nutrition. Taken together, these results suggest a new role for SAG12. This cysteine protease plays a crucial role in root N remobilization that ensures seed filling and sustains yields when nitrogen availability is low.

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SATURNE IJPB team

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Lateral Transport of Organic and Inorganic Solutes. Plants (Basel). 2019 Jan 15

Lateral Transport of Organic and Inorganic Solutes. Plants (Basel). 2019 Jan 15 | Publications @IJPB | Scoop.it

Organic (e.g., sugars and amino acids) and inorganic (e.g., K⁺, Na⁺, PO₄2-, and SO₄2-) solutes are transported long-distance throughout plants. Lateral movement of these compounds between the xylem and the phloem, and vice versa, has also been reported in several plant species since the 1930s, and is believed to be important in the overall resource allocation. Studies of Arabidopsis thaliana have provided us with a better knowledge of the anatomical framework in which the lateral transport takes place, and have highlighted the role of specialized vascular and perivascular cells as an interface for solute exchanges. Important breakthroughs have also been made, mainly in Arabidopsis, in identifying some of the proteins involved in the cell-to-cell translocation of solutes, most notably a range of plasma membrane transporters that act in different cell types. Finally, in the future, state-of-art imaging techniques should help to better characterize the lateral transport of these compounds on a cellular level. This review brings the lateral transport of sugars and inorganic solutes back into focus and highlights its importance in terms of our overall understanding of plant resource allocation.

 
 

 

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PATS IJPB team

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Combined Proteomic and Metabolomic Profiling of the Arabidopsis thaliana vps29 Mutant Reveals Pleiotropic Functions of the Retromer in Seed Development. Int J Mol Sci. 2019 Jan

Combined Proteomic and Metabolomic Profiling of the Arabidopsis thaliana vps29 Mutant Reveals Pleiotropic Functions of the Retromer in Seed Development. Int J Mol Sci. 2019 Jan | Publications @IJPB | Scoop.it

The retromer is a multiprotein complex conserved from yeast to humans, which is involved in intracellular protein trafficking and protein recycling. Selection of cargo proteins transported by the retromer depends on the core retromer subunit composed of the three vacuolar protein sorting (VPS) proteins, namely VPS26, VPS29, and VPS35. To gain a better knowledge of the importance of the plant retromer in protein sorting, we carried out a comparative proteomic and metabolomic analysis of Arabidopsis thaliana seeds from the wild-type and the null-retromer mutant vps29. Here, we report that the retromer mutant displays major alterations in the maturation of seed storage proteins and synthesis of lipid reserves, which are accompanied by severely impaired seed vigor and longevity. We also show that the lack of retromer components is counterbalanced by an increase in proteins involved in intracellular trafficking, notably members of the Ras-related proteins in brain (RAB) family proteins. Our study suggests that loss of the retromer stimulates energy metabolism, affects many metabolic pathways, including that of cell wall biogenesis, and triggers an osmotic stress response, underlining the importance of retromer function in seed biology.

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PHYGERM IJPB teal

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Isoflavone production in hairy root cultures and plantlets of Trifolium pratense. Biotechnol Lett. 2019 Jan 19.

Isoflavone production in hairy root cultures and plantlets of Trifolium pratense. Biotechnol Lett. 2019 Jan 19. | Publications @IJPB | Scoop.it

OBJECTIVES:

The aim of this study was to develop a Trifolium pratense hairy root (HR) production protocol and select HR lines with high isoflavone yield following elicitor treatments.

RESULTS:

We obtained 13 independent HR lines, producing approximately three times more isoflavonoids than seedlings (3.3 mg/g dry weight) and in which 27 isoflavonoids were detected. Each HR line had its own isoflavonoid profile. These lines produced as major components daidzein, genistein, formononetin and biochanin A. Sucrose, salicylic acid (SA), yeast extract (YE) and flagellin 22 (flg22) were tested as elicitors. Using SA 140 mg/L, allowed the maximum isoflavonoid production in plantlets (11.9 mg/g dry weight) but reduced root growth, possibly as a result of its toxicity. The highest isoflavone production in HR (27.9 mg/g dry weight) was obtained using sucrose 60 g/L, for 3.5 days.

CONCLUSION:

This work reports the high production of various isoflavonoids with T. pratense elicited HR cultures.

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IJPB Plant Observatory

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Small is big in Arabidopsis mitochondrial ribosome - Nat Plants. 2019 Jan 9

Small is big in Arabidopsis mitochondrial ribosome - Nat Plants. 2019 Jan 9 | Publications @IJPB | Scoop.it

Mitochondria are responsible for energy production through aerobic respiration, and represent the powerhouse of eukaryotic cells. Their metabolism and gene expression processes combine bacterial-like features and traits that evolved in eukaryotes. Among mitochondrial gene expression processes, translation remains the most elusive. In plants, while numerous pentatricopeptide repeat (PPR) proteins are involved in all steps of gene expression, their function in mitochondrial translation remains unclear. Here we present the biochemical characterization of Arabidopsis mitochondrial ribosomes and identify their protein subunit composition. Complementary biochemical approaches identified 19 plant-specific mitoribosome proteins, of which ten are PPR proteins. The knockout mutations of ribosomal PPR (rPPR) genes result in distinct macroscopic phenotypes, including lethality and severe growth delay. The molecular analysis of rppr1 mutants using ribosome profiling, as well as the analysis of mitochondrial protein levels, demonstrate rPPR1 to be a generic translation factor that is a novel function for PPR proteins. Finally, single-particle cryo-electron microscopy (cryo-EM) reveals the unique structural architecture of Arabidopsis mitoribosomes, characterized by a very large small ribosomal subunit, larger than the large subunit, bearing an additional RNA domain grafted onto the head. Overall, our results show that Arabidopsis mitoribosomes are substantially divergent from bacterial and other eukaryote mitoribosomes, in terms of both structure and protein content.

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OrganRepro IJPB team

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Pivotal roles of cryptochromes 1a and 2 in tomato development and physiology. Plant Physiol. 2018 Dec 12

Pivotal roles of cryptochromes 1a and 2 in tomato development and physiology. Plant Physiol. 2018 Dec 12 | Publications @IJPB | Scoop.it

Cryptochromes are flavin-containing blue/UV-A light photoreceptors that regulate various plant light-induced physiological processes. In Arabidopsis thaliana (Brassicaceae), cryptochromes mediate de-etiolation, photoperiodic control of flowering, entrainment of the circadian clock, cotyledon opening and expansion, anthocyanin accumulation and root growth. In tomato (Solanum lycopersicum; Solanaceae), cryptochromes are encoded by a multi-gene family, comprising CRY1a, CRY1b, CRY2 and CRY3. We have previously reported the phenotypes of tomato cry1a mutants and CRY2 overexpressing plants. Here, we report the isolation, by TILLING, of a tomato cry2 knock-out mutant, its introgression in the indeterminate Moneymaker background, and the phenotypes of cry1a/cry2 single and double mutants. The cry1a/cry2 mutant showed phenotypes similar to its Arabidopsis counterpart (long hypocotyls in white and blue light), but also several further features such as increased seed weight and internode length, enhanced hypocotyl length in red light, inhibited primary root growth under different light conditions, anticipation of flowering under long-day conditions and alteration of the phase of circadian leaf movements. Both cry1a and cry2 control the levels of photosynthetic pigments in leaves, whereas cry2 has a predominant role in fruit pigmentation. Metabolites of the sterol, tocopherol, quinone and sugar classes are differentially accumulated in cry1a and cry2 leaves and fruits. These results demonstrate a pivotal role of cryptochromes in controlling tomato development and physiology. The manipulation of these photoreceptors represents a powerful tool to influence important agronomic traits such as flowering time and fruit quality.

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AGG IJPB Team

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Design and visualization of second generation cyanoisoindole based fluorescent strigolactone analogs. Plant J. 2018 Dec 15

Design and visualization of second generation cyanoisoindole based fluorescent strigolactone analogs. Plant J. 2018 Dec 15 | Publications @IJPB | Scoop.it

Strigolactones (SLs) are a family of terpenoid allelochemicals that were recognized as plant hormones only a decade ago. They influence a myriad of both above- and belowground developmental processes, and are an important survival strategy for plants in nutrient-deprived soils. A rapidly emerging approach to gain knowledge on hormone signaling is the use of traceable analogs. A unique class of labeled SL analogs was constructed, where the original tricyclic lactone moiety of natural SLs is replaced by a fluorescent cyanoisoindole ring system. Biological evaluation as parasitic seed germination stimulant and hypocotyl elongation repressor proved the potency of the cyanoisoindole strigolactone analogs (CISA) to be comparable to the commonly accepted standard GR24. Additionally, via a SMXL6 protein degradation assay, we provided molecular evidence that the compounds elicit SL-like responses through the natural signaling cascade. All CISA analogs were shown to exhibit fluorescent properties, and the high quantum yield and Stokes shift of the pyrroloindole derivative CISA-7 also enabled in vivo visualization in plants. In contrast to the previously reported fluorescent analogs, CISA-7 displays a large similarity in shape and structure with natural SLs, which renders the analog a promising tracer to investigate the spatiotemporal distribution of SLs in plants and fungi.

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CORAM IJPB team

www-ijpb.versailles.inra.fr/en/bc/equipes/pois/index.html

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The Tonoplastic Inositol Transporter INT1 From Arabidopsis thaliana Impacts Cell Elongation in a Sucrose-Dependent Way. Front Plant Sci. 2018 Nov 16

The Tonoplastic Inositol Transporter INT1 From Arabidopsis thaliana Impacts Cell Elongation in a Sucrose-Dependent Way. Front Plant Sci. 2018 Nov 16 | Publications @IJPB | Scoop.it

The tonoplastic inositol transporter INT1 is the only known transport protein in Arabidopsis that facilitates myo-inositol import from the vacuole into the cytoplasm. Impairment of the release of vacuolar inositol by knockout of INT1 results in a severe inhibition of cell elongation in roots as well as in etiolated hypocotyls. Importantly, a more strongly reduced cell elongation was observed when sucrose was supplied in the growth medium, and this sucrose-dependent effect can be complemented by the addition of exogenous myo-inositol. Comparing int1 mutants (defective in transport) with mutants defective in myo-inositol biosynthesis (mips1 mutants) revealed that the sucrose-induced inhibition in cell elongation does not just depend on inositol depletion. Secondary effects as observed for altered availability of inositol in biosynthesis mutants, as disturbed membrane turnover, alterations in PIN protein localization or alterations in inositol-derived signaling molecules could be ruled out to be responsible for impairing the cell elongation in int1 mutants. Although the molecular mechanism remains to be elucidated, our data implicate a crucial role of INT1-transported myo-inositol in regulating cell elongation in a sucrose-dependent manner and underline recent reports of regulatory roles for sucrose and other carbohydrate intermediates as metabolic semaphores.

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ACCI IJPB team

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Photoinactivation of the S aureus Lactose-Specific EIICB Phosphotransferase Component with p-azidophenyl-β-D-Galactoside and Phosphorylation of the Covalently Bound Substrate - J Mol Microbiol Biot...

The phosphoenolpyruvate (PEP):lactose phosphotransferase system of Staphylococcus aureus transports and phosphorylates lactose and various phenylgalactosides. Their phosphorylation is catalyzed by the Cys476-phosphorylated EIIB domain of the lactose-specific permease enzyme IICB (EIICBLac). Phosphorylation causes the release of galactosides bound to the EIIC domain into the cytoplasm by a mechanism not yet understood. Results: Irradiation of a reaction mixture containing the photoactivatable p-azidophenyl-β-D-galactopyranoside and EIICBLac with UV light caused a loss of EIICBLac activity. Nevertheless, photoinactivated EIICBLac could still be phosphorylated with [32P]PEP. Proteolysis of photoinactivated [32P]P-EIICBLac with subtilisin provided an 11-kDa radioactive peptide. Only the sequence of its first three amino acids (-H-G-P-, position 245–247) could be determined. They are part of the substrate binding pocket in EIICs of the lactose/cellobiose PTS family. Surprisingly, while acid treatment caused hydrolysis of the phosphoryl group in active [32P]P∼EIICBLac, photoinactivated [32P]P-EIICBLac remained strongly phosphorylated. Conclusion: Phosphorylation of the –OH group at C6 of p-nitrenephenyl-β-D-galactopyranoside covalently bound to EIICLac by the histidyl-phosphorylated [32P]P∼EIIBLac domain is a likely explanation for the observed acid resistance. Placing p-nitrenephenyl-β-D-galactopyranoside into the active site of modelled EIICLac suggested that the nitrene binds to the -NH- group of Ser248, which would explain why no sequence data beyond Pro247could be obtained.

 

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DYSCOL IJPB team

www-ijpb.versailles.inra.fr/en/bs/equipes/biostructurale/index.htm

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Unleashing meiotic crossovers in crops. Nat Plants. 2018 Dec 4

Unleashing meiotic crossovers in crops. Nat Plants. 2018 Dec 4 | Publications @IJPB | Scoop.it

Improved plant varieties are important in our attempts to face the challenges of a growing human population and limited planet resources. Plant breeding relies on meiotic crossovers to combine favourable alleles into elite varieties1. However, meiotic crossovers are relatively rare, typically one to three per chromosome2, limiting the efficiency of the breeding process and related activities such as genetic mapping. Several genes that limit meiotic recombination were identified in the model species Arabidopsis thaliana2. Mutation of these genes in Arabidopsis induces a large increase in crossover frequency. However, it remained to be demonstrated whether crossovers could also be increased in crop species hybrids. We explored the effects of mutating the orthologues of FANCM3, RECQ44 or FIGL15 on recombination in three distant crop species, rice (Oryza sativa), pea (Pisum sativum) and tomato (Solanum lycopersicum). We found that the single recq4 mutation increases crossovers about three-fold in these crops, suggesting that manipulating RECQ4 may be a universal tool for increasing recombination in plants. Enhanced recombination could be used with other state-of-the-art technologies such as genomic selection, genome editing or speed breeding6 to enhance the pace and efficiency of plant improvement.

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MEIOME IJPB team

www-ijpb.versailles.inra.fr/en/sgap/equipes/meiose/mma.htm

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