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Transposable elements (TEs) are major contributors to genome complexity in eukaryotes. TE mobilization may cause genome instability, although it can also drive genome diversity throughout evolution. TE transposition may influence the transcriptional activity of neighboring genes by modulating the epigenomic profile of the region or by altering the relative position of regulatory elements. Notably, TEs have emerged in the last few years as an important source of functional long and small non-coding RNAs. A plethora of small RNAs derived from TEs have been linked to the trans regulation of gene activity at the transcriptional and post-transcriptional levels. Furthermore, TE-derived long non-coding RNAs have been shown to modulate gene expression by interacting with protein partners, sequestering active small RNAs, and forming duplexes with DNA or other RNA molecules. In this review, we summarize our current knowledge of the functional and mechanistic paradigms of TE-derived long and small non-coding RNAs and discuss their role in plant development and evolution.

The extraordinary maturation in high‐throughput sequencing technologies has revealed the existence of a complex network of transcripts in eukaryotic organisms, including thousands of long noncoding (lnc) RNAs with little or no protein‐coding capacity. Subsequent discoveries have shown that lncRNAs participate in a wide range of molecular processes, controlling gene expression and protein activity though direct interactions with proteins, DNA, or other RNA molecules. While significant advances have been achieved in the understanding of lncRNA biology in the animal kingdom, the functional characterization of plant lncRNAs is still in its infancy and remains a major challenge. In this review, we report emerging functional and mechanistic paradigms of plant lncRNAs and partner molecules and discuss how cutting‐edge technologies may help to identify and classify yet uncharacterized transcripts into functional groups. Fil: Lucero, Leandro Exequiel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Ferrero, Lucia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Fonouni-farde, Camille Audrey. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Ariel, Federico Damian. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina

Two class I TCP transcription factors are required for an efficient elongation of hypocotyls in response to auxin and for the correct expression of a subset of auxin-inducible genes In this work, we analyzed the response to auxin of plants with altered function of the class I TEOSINTE BRANCHED 1, CYCLOIDEA, PCF (TCP) transcription factors TCP14 and TCP15. Several SMALL AUXIN UP RNA (SAUR) genes showed decreased expression in mutant plants defective in these TCPs after an increase in ambient temperature to 29 °C, a condition that causes an increase in endogenous auxin levels. Overexpression of SAUR63 caused a more pronounced elongation response in the mutant than in the wild-type at 29 °C, suggesting that the decreased expression of SAUR genes is partly responsible for the defective elongation at warm temperature. Notably, several SAUR genes and the auxin response gene IAA19 also showed reduced expression in the mutant after auxin treatment, while the expression of other SAUR genes and of IAA29 was not affected or was even higher. Expression of the auxin reporter DR5::GUS was also higher in a tcp15 mutant than in a wild-type background after auxin treatment. However, the elongation of hypocotyls in response to auxin was impaired in the mutant. Remarkably, a significant proportion of auxin inducible genes and of targets of the AUXIN RESPONSE FACTOR 6 are regulated by TCP15 and often contain putative TCP recognition motifs in their promoters. Furthermore, we demonstrated that several among them are recognized by TCP15 in vivo. Our results indicate that TCP14 and TCP15 are required for an efficient elongation response to auxin, most likely by regulating a subset of auxin inducible genes related to cell expansion.

CURLY LEAF (CLF) encodes the methyl-transferase sub-unit of the PolycombRepressor Complex 2 (PRC2), which regulates the expression of target genesthrough H3K27 tri-methylation. We isolated a new CLF mutant allele (clf-78)using a genetic screening designed to identify micro RNAs (miRNA) deficientmutants. CLF mutant plants showed impaired miRNA activity caused byincreased AGO1 ubiquitination and enhanced degradation in specific tissues.Such CLF-mediated AGO1 regulation was evidenced when plants wereexposed to UV radiation, causing increased susceptibility of clf mutants to someUV-induced responses. Furthermore, we showed that CLF directly regulatesFBW2, which in turn triggers AGO1 degradation in the mutants. Interestingly,AGO1 bound to a target appeared particularly prone to degradation in themutant plants, a process that is exacerbated when the complex bound a noncleavable target. Thus, a prolonged AGO1-target interaction seems to favorAGO1 degradation, suggesting that non-cleavable miRNA targets mayovercome translation inhibition by modulating AGO1 stability in plants. Fil: Ré, Delfina Adela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Cambiagno, Damián Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Arce, Agustín Lucas. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Tomassi, Ariel Hernán. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Giustozzi, Marisol. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Centro de Estudios Fotosintéticos y Bioquímicos. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Centro de Estudios Fotosintéticos y Bioquímicos; Argentina Fil: Casati, Paula. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Centro de Estudios Fotosintéticos y Bioquímicos. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Centro de Estudios Fotosintéticos y Bioquímicos; Argentina Fil: Ariel, Federico Damian. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Manavella, Pablo Andrés. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina

SUMMARYRNA-DNA hybrid (R-loop)-associated long noncoding RNAs (lncRNAs), including the Arabidopsis lncRNAAUXIN-REGULATED PROMOTER LOOP(APOLO), are emerging as important regulators of three-dimensional chromatin conformation and gene transcriptional activity. Here, we showed that in addition to the PRC1-component LIKE-HETEROCHROMATIN PROTEIN 1 (LHP1),APOLOinteracts with the methylcytosine-binding protein VARIANT IN METHYLATION 1 (VIM1), a conserved homolog of the mammalian DNA methylation regulator UBIQUITIN-LIKE CONTAINING PHD AND RING FINGER DOMAINS 1 (UHRF1). TheAPOLO-VIM1-LHP1 complex directly regulates the transcription of the auxin biosynthesis geneYUCCA2by dynamically determining DNA methylation and H3K27me3 deposition over its promoter during the plant thermomorphogenic response. Strikingly, we demonstrated that the lncRNAUHRF1 Protein Associated Transcript(UPAT), a direct interactor of UHRF1 in humans, can be recognized by VIM1 and LHP1 in plant cells, despite the lack of sequence homology betweenUPATandAPOLO. In addition, we showed that increased levels ofAPOLOorUPAThamper VIM1 and LHP1 binding toYUCCA2promoter. Collectively, our results uncover a new mechanism in which a plant lncRNA coordinates Polycomb action and DNA methylation, and reveal that evolutionary unrelated lncRNAs may exert similar functions across kingdoms.

Plant long noncoding RNAs (lncRNAs) are key chromatin dynamics regulators, directing the transcriptional programs driving a wide variety of developmental outputs. Recently, we uncovered how the lncRNA AUXIN REGULATED PROMOTER LOOP (APOLO) directly recognizes the locus encoding the root hair (RH) master regulator ROOT HAIR DEFECTIVE 6 (RHD6) modulating its transcriptional activation and leading to low temperature-induced RH elongation. We further demonstrated that APOLO interacts with the transcription factor WRKY42 in a novel ribonucleoprotein complex shaping RHD6 epigenetic environment and integrating signals governing RH growth and development. In this work, we expand this model showing that APOLO is able to bind and positively control the expression of several cell wall EXTENSIN (EXT) encoding genes, including EXT3, a key regulator for RH growth. Interestingly, EXT3 emerged as a novel common target of APOLO and WRKY42. Furthermore, we showed that the ROS homeostasis-related gene NADPH OXIDASE C (NOXC) is deregulated upon APOLO overexpression, likely through the RHD6-RSL4 pathway, and that NOXC is required for low temperature-dependent enhancement of RH growth. Collectively, our results uncover an intricate regulatory network involving the APOLO/WRKY42 hub in the control of master and effector genes during RH development.

In Arabidopsis, lateral root (LR) development is mainly controlled by several known auxin-regulated transcription factors (TFs). Here, we show that AtHB23 (a homeodomain-leucine zipper I TF) participates in this intricate network. Our study of the expression pattern of AtHB23 revealed that it is transcriptionally activated in the early stages of secondary LR primordium. We found that AtHB23 directly limits the expression of LBD16, a key factor in LR initiation, and also directly induces the auxin transporter gene LAX3. We propose that this HD-Zip I mediates the regulation of LAX3 by ARF7/19. Furthermore, AtHB23 plays distinct roles during the formation of secondary and tertiary roots, exhibiting differential expression patterns. ATHB23 is expressed throughout the tertiary root primordium, whereas it is restricted to early stages in secondary primordia, likely later repressing LBD16 in tertiary LR development and further inhibiting root emergence. Our results suggest that different genetic programs govern the formation of LR primordia from the main or secondary roots, thereby shaping the global dynamic architecture of the root system. This article is protected by copyright. All rights reserved. Fil: Perotti, María Florencia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Ribone, Pamela Anahí. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Cabello, Julieta Virginia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Ariel, Federico Damian. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Chan, Raquel Lia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina

International audience; The first reference to the “C-value paradox” reported an apparent imbalance between organismal genome size and morphological complexity. Since then, next-generation sequencing has revolutionized genomic research and revealed that eukaryotic transcriptomes contain a large fraction of non-protein-coding components. Eukaryotic genomes are pervasively transcribed and noncoding regions give rise to a plethora of noncoding RNAs with undeniable biological functions. Among them, long noncoding RNAs (lncRNAs) seem to represent a new layer of gene expression regulation, participating in a wide range of molecular mechanisms at the transcriptional and post-transcriptional levels. In addition to their role in epigenetic regulation, plant lncRNAs have been associated with the degradation of complementary RNAs, the regulation of alternative splicing, protein sub-cellular localization, the promotion of translation and protein post-translational modifications. In this review, we report and integrate numerous and complex mechanisms through which long noncoding transcripts regulate post-transcriptional gene expression in plants.

Clustered organization of biosynthetic non-homologous genes is emerging as a characteristic feature of plant genomes. The co-regulation of clustered genes seems to largely depend on epigenetic reprogramming and three-dimensional chromatin conformation. Here we identified the long noncoding RNA (lncRNA) MARneral Silencing (MARS), localized inside the Arabidopsis marneral cluster, which controls the local epigenetic activation of its surrounding region in response to ABA. MARS modulates the POLYCOMB REPRESSIVE COMPLEX 1 (PRC1) component LIKE-HETEROCHROMATIN PROTEIN 1 (LHP1) binding throughout the cluster in a dose-dependent manner, determining H3K27me3 deposition and chromatin condensation. In response to ABA, MARS decoys LHP1 away from the cluster and promotes the formation of a chromatin loop bringing together the MARNERAL SYNTHASE 1 (MRN1) locus and a distal ABA-responsive enhancer. The enrichment of co-regulated lncRNAs in clustered metabolic genes in Arabidopsis suggests that the acquisition of novel noncoding transcriptional units may constitute an additional regulatory layer driving the evolution of biosynthetic pathways.

Deep learning methods have outperformed previous techniques in most computer vision tasks, including image-based plant phenotyping. However, massive data collection of root traits and the development of associated artificial intelligence approaches have been hampered by the inaccessibility of the rhizosphere. Here we present ChronoRoot, a system which combines 3D printed open-hardware with deep segmentation networks for high temporal resolution phenotyping of plant roots in agarized medium. We developed a novel deep learning based root extraction method which leverages the latest advances in convolutional neural networks for image segmentation, and incorporates temporal consistency into the root system architecture reconstruction process. Automatic extraction of phenotypic parameters from sequences of images allowed a comprehensive characterization of the root system growth dynamics. Furthermore, novel time-associated parameters emerged from the analysis of spectral features derived from temporal signals. Altogether, our work shows that the combination of machine intelligence methods and a 3D-printed device expands the possibilities of root high-throughput phenotyping for genetics and natural variation studies as well as the screening of clock-related mutants, revealing novel root traits.; Deep learning methods have outperformed previous techniques in most computer vision tasks, including image-based plant phenotyping. However, massive data collection of root traits and the development of associated artificial intelligence approaches have been hampered by the inaccessibility of the rhizosphere. Here we present ChronoRoot, a system which combines 3D printed open-hardware with deep segmentation networks for high temporal resolution phenotyping of plant roots in agarized medium. We developed a novel deep learning based root extraction method which leverages the latest advances in convolutional neural networks for image segmentation, and incorporates temporal consistency into the root system architecture reconstruction process. Automatic extraction of phenotypic parameters from sequences of images allowed a comprehensive characterization of the root system growth dynamics. Furthermore, novel time-associated parameters emerged from the analysis of spectral features derived from temporal signals. Altogether, our work shows that the combination of machine intelligence methods and a 3D-printed device expands the possibilities of root high-throughput phenotyping for genetics and natural variation studies as well as the screening of clock-related mutants, revealing novel root traits.

Trichomes and the cuticle are two specialized structures of the aerial epidermis that are important for plant organ development and interaction with the environment. In this study, we report that Arabidopsis thaliana plants affected in the function of the class I TEOSINTE BRANCHED 1, CYCLOIDEA, PCF (TCP) transcription factors TCP14 and TCP15 show overbranched trichomes in leaves and stems and increased cuticle permeability. We found that TCP15 regulates the expression of MYB106, a MIXTA-like transcription factor involved in epidermal cell and cuticle development, and overexpression of MYB106 in a tcp14 tcp15 mutant reduces trichome branch number. TCP14 and TCP15 are also required for the expression of the cuticle biosynthesis genes CYP86A4, GPAT6, and CUS2, and of SHN1 and SHN2, two AP2/EREBP transcription factors required for cutin and wax biosynthesis. SHN1 and CUS2 are also targets of TCP15, indicating that class I TCPs influence cuticle formation acting at different levels, through the regulation of MIXTA-like and SHN transcription factors and of cuticle biosynthesis genes. Our study indicates that class I TCPs are coordinators of the regulatory network involved in trichome and cuticle development. Fil: Camoirano, Alejandra. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Arce, Agustín Lucas. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Ariel, Federico Damian. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Alem, Antonela Lucía. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Gonzalez, Daniel Hector. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Viola, Ivana Lorena. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina

International audience; Plant long noncoding RNAs (lncRNAs) have emerged as important regulators of chromatin dynamics, impacting on transcriptional programs leading to different developmental outputs. The lncRNA AUXIN-REGULATED PROMOTER LOOP (APOLO) directly recognizes multiple independent loci across the Arabidopsis genome and modulates their three-dimensional chromatin conformation, leading to transcriptional shifts. Here, we show that APOLO recognizes the locus encoding the root hair (RH) master regulator ROOT HAIR DEFECTIVE 6 (RHD6) and controls RHD6 transcriptional activity, leading to cold-enhanced RH elongation through the consequent activation of the transcription factor gene RHD6-like RSL4. Furthermore, we demonstrate that APOLO interacts with the transcription factor WRKY42 and modulates its binding to the RHD6 promoter. WRKY42 is required for the activation of RHD6 by low temperatures and WRKY42 deregulation impairs cold-induced RH expansion. Collectively, our results indicate that a novel ribonucleoprotein complex with APOLO and WRKY42 forms a regulatory hub to activate RHD6 by shaping its epigenetic environment and integrate signals governing RH growth and development.

International audience; The advent of novel high-throughput sequencing techniques has revealed that eukaryotic genomes are massively transcribed although only a small fraction of RNAs exhibits protein-coding capacity. In the last years, long noncoding RNAs (lncRNAs) have emerged as regulators of eukaryotic gene expression in a wide range of molecular mechanisms. Plant lncRNAs can be transcribed by alternative RNA polymerases, acting directly as long transcripts or can be processed into active small RNAs. Several lncRNAs have been recently shown to interact with chromatin, DNA or nuclear proteins to condition the epigenetic environment of target genes or modulate the activity of transcriptional complexes. In this review, we will summarize the recent discoveries about the actions of plant lncRNAs in the regulation of gene expression at the transcriptional level.

Plant long noncoding RNAs (lncRNAs) have emerged as important regulators of chromatin dynamics, impacting on transcriptional programs leading to different developmental outputs. The lncRNAAUXIN REGULATED PROMOTER LOOP(APOLO) directly recognizes multiple independent loci across theArabidopsisgenome and modulates their three-dimensional chromatin conformation, leading to transcriptional shifts. Here, we show thatAPOLOrecognizes the locus encoding the root hair (RH) master regulator ROOT HAIR DEFECTIVE 6 (RHD6) and controlsRHD6transcriptional activity leading to cold-enhanced RH elongation through the consequent activation of the transcription factor gene RHD6-likeRSL4. Furthermore, we demonstrate thatAPOLOinteracts with the transcription factor WRKY42 and modulates its binding to theRHD6promoter. WRKY42 is required for the activation ofRHD6by low temperatures andWRKY42deregulation impairs cold-induced RH expansion. Collectively, our results indicate that a novel ribonucleoprotein complex involvingAPOLOand WRKY42 forms a regulatory hub which activatesRHD6by shaping its epigenetic environment and integrates signals governing RH growth and development.SUMMARYThe lncRNAAPOLOdirectly regulates the transcription of the root hair-master geneRHD6. In response to cold,APOLOis induced and it decoys the H3K27me3-binding protein LHP1 away fromRHD6. In addition,APOLOmodulates the binding of the transcription factor WRKY42 to theRHD6promoter at low temperatures.

Root architecture depends on the development of the main root and also on the number and density of lateral roots. Most molecular knowledge about the development of lateral roots was acquired studying primary roots, and it was implied that high order roots follow the same pattern. Recently, we informed that AtHB23 is differentially regulated in primary and secondary roots. Here we show that LBD16, a target of AtHB23, also is differentially regulated; it is expressed in the tip of secondary and tertiary roots but not in primary ones. Moreover, the key hormone auxin exhibits a different distribution pattern in secondary and tertiary roots, according to the reporter DR5. Finally, we show that in Col 0 and Ler ecotypes development of secondary and tertiary roots exhibits significant variations. Altogether, we can conclude that different genetic programs govern secondary and tertiary roots development and such processes are dependent on the Arabidopsis genotype. Fil: Perotti, María Florencia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Ariel, Federico Damian. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Chan, Raquel Lia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina

RNA-Binding Protein 1 (RBP1) was first identified as a protein partner of the long noncoding RNA (lncRNA) ENOD40 in Medicago truncatula, involved in symbiotic nodule development. RBP1 is localized in nuclear speckles and can be relocalized to the cytoplasm by the interaction with ENOD40. The two closest homologs to RBP1 in Arabidopsis thaliana were called Nuclear Speckle RNA-binding proteins (NSRs) and characterized as alternative splicing modulators of specific mRNAs. They can recognize in vivo the lncRNA ALTERNATIVE SPLICING COMPETITOR (ASCO) among other lncRNAs, regulating lateral root formation. Here, we performed a phylogenetic analysis of NSR/RBP proteins tracking the roots of the family to the Embryophytes. Strikingly, eudicots faced a reductive trend of NSR/RBP proteins in comparison with other groups of flowering plants. In Medicago truncatula and Lotus japonicus, their expression profile during nodulation and in specific regions of the symbiotic nodule was compared to that of the lncRNA ENOD40, as well as to changes in alternative splicing. This hinted at distinct and specific roles of each member during nodulation, likely modulating the population of alternatively spliced transcripts. Our results establish the basis to guide future exploration of NSR/RBP function in alternative splicing regulation in different developmental contexts along the plant lineage. Fil: Lucero, Leandro Exequiel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Bazin, Jeremie. Université Paris-sud; Francia. Centre National de la Recherche Scientifique; Francia. Institut National de la Recherche Agronomique; Francia Fil: Rodriguez Melo, Johan Stiben. Universidad Nacional de Rio Cuarto. Facultad de Ciencias Exactas Fisicoquímicas y Naturales. Instituto de Investigaciones Agrobiotecnológicas - Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Agrobiotecnológicas; Argentina Fil: Ibañez, Fernando Julio. Universidad Nacional de Rio Cuarto. Facultad de Ciencias Exactas Fisicoquímicas y Naturales. Instituto de Investigaciones Agrobiotecnológicas - Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Agrobiotecnológicas; Argentina Fil: Crespi, Leandro Martín. Centre National de la Recherche Scientifique; Francia. Institut National de la Recherche Agronomique; Francia. Université Paris-sud; Francia Fil: Ariel, Federico Damian. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina

In autogamous plants like Arabidopsis (Arabidopsis thaliana), stamen filament elongation must be finely regulated to ensure that anthers reach the pistil at the correct developmental stage. In this work, we studied the roles of Arabidopsis TEOSINTE BRANCHED1, CYCLOIDEA, PCF15 (TCP15), and related class-I TCP transcription factors in stamen filament elongation. Plants with decreased expression of class-I TCPs and plants that express a fusion of TCP15 to a repressor domain (pTCP15::TCP15-EAR) had shorter stamens, indicating that class-I TCPs stimulate filament growth. These plants also showed reduced expression of several SMALL AUXIN UP RNA (SAUR)63 subfamily genes, which contain TCP target motifs in their promoters. Mutational analysis indicated that the TCP target motif in the SAUR63 promoter is required for expression of SAUR63 in stamen filaments. Moreover, TCP15 directly binds to the SAUR63 promoter region that contains the TCP target motif in vivo, highlighting the role of the TCPs in this process. Class-I TCPs are also required for the induction of SAUR63 subfamily genes by gibberellins (GAs). In addition, overexpression of SAUR63 restores filament growth in pTCP15::TCP15-EAR plants, whereas overexpression of TCP15 rescues the short stamen phenotype of GA-deficient plants. The results indicate that TCP15 and related class-I TCPs modulate GA-dependent stamen filament elongation by direct activation of SAUR63 subfamily genes through conserved target sites in their promoters. This work provides insight into GA-dependent stamen filament elongation. Fil: Gastaldi, Victoria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Lucero, Leandro Exequiel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Ferrero, Lucia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Ariel, Federico Damian. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Gonzalez, Daniel Hector. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina

Nuclear speckle RNA binding proteins (NSRs) act as regulators of alternative splicing (AS) and auxin-regulated developmental processes such as lateral root formation in Arabidopsis thaliana. These proteins were shown to interact with specific alternatively spliced mRNA targets and at least with one structured lncRNA, named Alternative Splicing Competitor RNA. Here, we used genome-wide analysis of RNAseq to monitor the NSR global role on multiple tiers of gene expression, including RNA processing and AS. NSRs affect AS of 100s of genes as well as the abundance of lncRNAs particularly in response to auxin. Among them, the FPA floral regulator displayed alternative polyadenylation and differential expression of antisense COOLAIR lncRNAs in nsra/b mutants. This may explains the early flowering phenotype observed in nsra and nsra/b mutants. GO enrichment analysis of affected lines revealed a novel link of NSRs with the immune response pathway. A RIP-seq approach on an NSRa fusion protein in mutant background identified that lncRNAs are privileged direct targets of NSRs in addition to specific AS mRNAs. The interplay of lncRNAs and AS mRNAs in NSR-containing complexes may control the crosstalk between auxin and the immune response pathway. Fil: Bazin, Jérémie. Universite Paris-sud Xi; Fil: Romero, Natali. Universite Paris-sud Xi; Fil: Rigo, Richard. Universite Paris-sud Xi; Fil: Charon, Celine. Universite Paris-sud Xi; Fil: Blein, Thomas. Universite Paris-sud Xi; Fil: Ariel, Federico Damian. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Crespi, Martin. Universite Paris-sud Xi

Fil: Carlotto, Nicolas. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Oficina de Coordinacion Administrativa Ciudad Universitaria. Instituto de Biodiversidad y Biologia Experimental y Aplicada. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Biodiversidad y Biologia Experimental y Aplicada; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Fisiologia, Biologia Molecular y Celular. Laboratorio de Agrobiotecnologia; Argentina

The term non-coding RNA (ncRNA) refers to functional RNA molecules that, despite being transcribed from DNA, are not translated into proteins. These molecules can play an important role in the regulation of gene expression in the eukaryotic cell, and they can act either as long ncRNAs or being processed intosmall RNAs, being globally classified by their size, function or genomic origin. In recent years, it has been found that diverse ncRNAs participate directly or indirectly in several epigenetic phenomena controlling different phenotypes within clonal cells, and in the specificity determination of various hysiological processes. Although some of their mechanisms of action have been characterized, much remains to be known to understand the highly complex processes in which most of these molecules are involved. In this chapter we discuss and illustrate examples of different ncRNAs that can interact with the plant epigenomic machinery or intervene in its function, leading to specific epigenetic, transcriptional and physiological states. We explore the link between chromatin compaction, histone modifications, DNA methylation, gene silencing and these molecules, which represent a high proportion of the cellular transcriptome. Fil: Ramirez Prado, Juan. Institute of Plant Sciences Paris-Saclay; Francia. King Abdullah University of Science and Technology; Arabia Saudita Fil: Ariel, Federico Damian. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Benhamed, Moussa. King Abdullah University of Science and Technology; Arabia Saudita. Institute of Plant Sciences Paris-Saclay; Francia Fil: Crespi, Martin. Institute of Plant Sciences Paris-Saclay; Francia

Summary Legume crops related to the model plant Medicago truncatula can adapt their root architecture to environmental conditions, both by branching and by establishing a symbiosis with rhizobial bacteria to form nitrogen-fixing nodules. Soil salinity is a major abiotic stress affecting plant yield and root growth. Previous transcriptomic analyses identified several transcription factors linked to the M. truncatula response to salt stress in roots, including NAC (NAM/ATAF/CUC)-encoding genes. Over-expression of one of these transcription factors, MtNAC969, induced formation of a shorter and less-branched root system, whereas RNAi-mediated MtNAC969 inactivation promoted lateral root formation. The altered root system of over-expressing plants was able to maintain its growth under high salinity, and roots in which MtNAC969 was down-regulated showed improved growth under salt stress. Accordingly, expression of salt stress markers was decreased or induced in MtNAC969 over-expressing or RNAi roots, respectively, suggesting a repressive function for this transcription factor in the salt-stress response. Expression of MtNAC969 in central symbiotic nodule tissues was induced by nitrate treatment, and antagonistically affected by salt in roots and nodules, similarly to senescence markers. MtNAC969 RNAi nodules accumulated amyloplasts in the nitrogen-fixing zone, and were prematurely senescent. Therefore, the MtNAC969 transcription factor, which is differentially affected by environmental cues in root and nodules, participates in several pathways controlling adaptation of the M. truncatula root system to the environment.

Phytohormonal interactions are essential to regulate plant organogenesis. In response to the presence of signals from symbiotic bacteria, the Nod factors, legume roots generate a new organ: the nitrogen-fixing nodule. Analysis of mutants in the Medicago truncatula CRE1 cytokinin receptor and of the MtRR4 cytokinin primary response gene expression pattern revealed that cytokinin acts in initial cortical cell divisions and later in the transition between meristematic and differentiation zones of the mature nodule. MtCRE1 signaling is required for activation of the downstream nodulation-related transcription factors MtERN1, MtNSP2 and MtNIN, as well as to regulate expression and accumulation of PIN auxin efflux carriers. Whereas the MtCRE1 pathway is required to allow the inhibition of polar auxin transport in response to rhizobia, nodulation is still negatively regulated by the MtEIN2/SICKLE-dependent ethylene pathway in cre1 mutants. Hence, MtCRE1 signaling acts as a regulatory knob, integrating positive plant and bacterial cues to control legume nodule organogenesis.

The endosymbiotic association between legumes and rhizobia leads to the formation of root nodules in which differentiated bacteria convert atmospheric nitrogen into a form that can be assimilated by the host plant. Successful root infection by rhizobia and nodule organogenesis require the activation of symbiotic genes that are controlled by a set of early transcription factors (TFs). MtNF-YA1 and MtNF-YA2 are two TFs playing partially redundant functions during several steps of the symbiotic interaction between Medicago truncatula and Sinorhizobium meliloti. NF-Y proteins are part of a transcriptional complex composed of three proteins (NF-YA, NF-YB and NF-YC) which bind DNA at CCAAT-boxes, a motif present in most eukaryotic promoters. In plants, each subunit is encoded by small gene families, potentially leading to a multitude of heterotrimeric NF-Y complexes. Here, using yeast two hybrid screenings, we identified the MtNF-YB and MtNF-YC subunits that interact with MtNF-YA1 and A2. Further, we confirmed, both in yeast and in planta, the formation of trimeric NF-Y complexes and showed that these complexes are functional during nodulation using reverse genetic approaches and ChIP-PCR. Finally, as orthologs of the characterized NF-Y subunits also control nodulation in other legumes, we showed in common bean that similar NF-Y trimers could form in planta. Our results suggest that we have identified a group of evolutionary conserved NF-Y proteins that interact to control nodulation in leguminous plants. Fil: Baudin, Maël. Centre National de la Recherche Scientifique; Francia. Centre de Recherche de Nantes. Institut National de la Recherche Agronomique; Francia Fil: Laloum, Tom. Centre National de la Recherche Scientifique; Francia. Centre de Recherche de Nantes. Institut National de la Recherche Agronomique; Francia Fil: Lepage, Agnes. Centre National de la Recherche Scientifique; Francia. Centre de Recherche de Nantes. Institut National de la Recherche Agronomique; Francia Fil: Rípodas, Carolina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; Argentina Fil: Ariel, Federico Damian. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Centre D'etudes de Saclay; Francia Fil: Frances, Lisa. Centre National de la Recherche Scientifique; Francia. Centre de Recherche de Nantes. Institut National de la Recherche Agronomique; Francia Fil: Crespi, Martin. Centre D'etudes de Saclay; Francia Fil: Gamas, Pascal. Centre National de la Recherche Scientifique; Francia Fil: Blanco, Flavio Antonio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; Argentina Fil: Zanetti, María Eugenia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; Argentina Fil: de Carvalho-Niebel, Fernanda. Centre National de la Recherche Scientifique; Francia. Centre de Recherche de Nantes. Institut National de la Recherche Agronomique; Francia Fil: Niebel, Andreas. Centre National de la Recherche Scientifique; Francia. Centre de Recherche de Nantes. Institut National de la Recherche Agronomique; Francia

International audience; Noncoding RNAs have emerged as major components of the eukaryotic transcriptome. Genome-wide analyses revealed the existence of thousands of long noncoding RNAs (IncRNAs) in several plant species. Plant IncRNAs are transcribed by the plant-specific RNA polymerases Pol IV and Pol V, leading to transcriptional gene silencing, as well as by Pol II. They are involved in a wide range of regulatory mechanisms impacting on gene expression, including chromatin remodeling, modulation of alternative splicing, fine-tuning of miRNA activity, and the control of mRNA translation or accumulation. Recently, dual noncoding transcription by alternative RNA polymerases was implicated in epigenetic and chromatin conformation dynamics. This review integrates the current knowledge on the regulatory mechanisms acting through plant noncoding transcription.

SWI/SNF complexes mediate ATP-dependent chromatin remodeling to regulate gene expression. Many components of these complexes are evolutionarily conserved, and several subunits of Arabidopsis thaliana SWI/SNF complexes are involved in the control of flowering, a process that depends on the floral repressor FLOWERING LOCUS C (FLC). BAF60 is a SWI/SNF subunit, and in this work, we show that BAF60, via a direct targeting of the floral repressor FLC, induces a change at the high-order chromatin level and represses the photoperiod flowering pathway in Arabidopsis. BAF60 accumulates in the nucleus and controls the formation of the FLC gene loop by modulation of histone density, composition, and posttranslational modification. Physiological analysis of BAF60 RNA interference mutant lines allowed us to propose that this chromatin-remodeling protein creates a repressive chromatin configuration at the FLC locus.

Cytokinin regulates many aspects of plant development, and in legume crops, this phytohormone is necessary and sufficient for symbiotic nodule organogenesis, allowing them to fix atmospheric nitrogen. To identify direct links between cytokinins and nodule organogenesis, we determined a consensus sequence bound in vitro by a transcription factor (TF) acting in cytokinin signaling, the nodule-enhanced Medicago truncatula Mt RR1 response regulator (RR). Among genes rapidly regulated by cytokinins and containing this so-called RR binding site (RRBS) in their promoters, we found the nodulation-related Type-A RR Mt RR4 and the Nodulation Signaling Pathway 2 (NSP2) TF. Site-directed mutagenesis revealed that RRBS cis-elements in the RR4 and NSP2 promoters are essential for expression during nodule development and for cytokinin induction. Furthermore, a microRNA targeting NSP2 (miR171 h) is also rapidly induced by cytokinins and then shows an expression pattern anticorrelated with NSP2. Other primary targets regulated by cytokinins depending on the Cytokinin Response1 (CRE1) receptor were a cytokinin oxidase/dehydrogenase (CKX1) and a basic Helix-Loop-Helix TF (bHLH476). RNA interference constructs as well as insertion of a Tnt1 retrotransposon in the bHLH gene led to reduced nodulation. Hence, we identified two TFs, NSP2 and bHLH476, as direct cytokinin targets acting at the convergence of phytohormonal and symbiotic cues.

International audience; Plants have remarkable developmental plasticity, and the same genotype can result in different phenotypes depending on environmental variation. Indeed, abiotic stresses or biotic interactions affect organogenesis and post-embryonic growth and significantly affect gene regulation. The large diversity of non-protein-coding RNAs (npcRNAs) and genes containing only short open reading frames that are expressed during plant growth and development, contribute to the regulation of gene expression. Certain npcRNAs code for oligopeptides and may possess additional biological activity linked to the RNA moiety. The ENOD40 gene is a dual RNA that is activated during a symbiotic interaction leading to root nodule organogenesis. Both the oligopeptides encoded by ENOD40 and the structured regions of the ENOD40 RNA have been shown to interact with different proteins in the cell to control enzymatic activities or induce the relocalisation of ribonucleoproteins, respectively. Other npcRNAs encode for small signalling peptides or are the precursors of small RNAs involved in post-transcriptional or transcriptional gene silencing. They may have RNA-related activities or encode peptides (or even larger proteins), and therefore act as dual RNAs. In addition, long natural antisense RNAs with a coding function and a regulatory RNA-mediated action that are expressed in response to abiotic stress in plants have been identified. In certain cases, these RNAs lead to the synthesis of nat-siRNAs, that are small RNAs derived from the overlapping double-stranded RNA region of natural antisense RNAs, which facilitates the silencing of complementary mRNAs. Finally, the advent of deep sequencing technologies has identified a large number of non-protein-coding RNAs in plants, which could be a large reservoir for dual RNAs.

The adaptation of root architecture to environmental constraints is a major agricultural trait, notably in legumes, the third main crop worldwide. This root developmental plasticity depends on the formation of lateral roots (LRs) emerging from primary roots. In the model legume Medicago truncatula, the HD-Zip I transcription factor HB1 is expressed in primary and lateral root meristems and induced by salt stress. Constitutive expression of HB1 in M. truncatula roots alters their architecture, whereas hb1 TILLING mutants showed increased lateral root emergence. Electrophoretic mobility shift assay, promoter mutagenesis, and chromatin immunoprecipitation-PCR assays revealed that HB1 directly recognizes a CAATAATTG cis-element present in the promoter of a LOB-like (for Lateral Organ Boundaries) gene, LBD1, transcriptionally regulated by auxin. Expression of these genes in response to abscisic acid and auxin and their behavior in hb1 mutants revealed an HB1-mediated repression of LBD1 acting during LR emergence. M. truncatula HB1 regulates an adaptive developmental response to minimize the root surface exposed to adverse environmental stresses. Fil: Ariel, Federico Damian. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Diet, Anouck. Centre National de la Recherche Scientifique; Francia Fil: Verdenaud, Marion. Institut National de la Recherche Agronomique; Francia Fil: Gruber, Véronique. Centre National de la Recherche Scientifique; Francia Fil: Frugier, Florian. Centre National de la Recherche Scientifique; Francia Fil: Chan, Raquel Lia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina Fil: Crespi, Martín. Centre National de la Recherche Scientifique; Francia