RESEARCH
Plants are very interesting living beings! Unlike animals, plants are extremely plastic in responding to changes in the environment and they can shape their own development! Plants are capable to generate new tissues/organs throughout their entire life cycle. The establishment and development of such tissues/organs depend upon several physiological and molecular processes that go on inside the cells.
Our research group is interested in studying the formation of vegetative and reproductive organs as well as how plants shape these organs in response to the environment. More specifically, we are interested in understanding the molecular mechanisms that underline the formation of these organs. Several genetic “players” are involved in such mechanisms, including transcription factors, epigenetic factors (such as chromatin remodelers) and small non-coding RNAs (sRNAs).
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sRNAs of 19-25 nt in size regulate transcriptionally and posttranscriptionally gene expression, shaping the transcriptome and the proteome of the cells. Amongst them, small interfering RNAs (siRNAs) and microRNAs (miRNAs) play crucial roles in plant development. Although siRNAs and miRNAs are both generated by long non-coding RNAs, their biogenesis differs in crucial points. Deep sequencing approaches helped us to identify hundreds of distinct sRNAs that may have important roles in development. For example, our group identified sRNAs associated with sugarcane (an important Brazilian biofuel crop) axillary bud dormancy and development.
We identified for example an emerging new class of sRNAs called tRNA-derived fragments or tRFs that may have roles in cell differentiation.
We are studying the molecular and genetic interactions between microRNAs and phytohormones during some aspects of plant development, such as flowering and fruit development.
We also are studying the roles of specific miRNAs in flower and fruit development and rooting. Finally, we are interested in understanding the possible roles of epigenetic factors in the plant response to the environment. To do so, we are using Arabidopsis and tomato as models.
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To study these molecular mechanisms, we are using a variety of tools such as: cloning, next-generation deep sequencing, generation of transgenic and mutant plants, protein interactions, ChIP, luciferase assays, and in situ hybridization.
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You can have a look in our main projects going on in the lab.
MAIN PROJECTS
Interactions between light, microRNAs and hormones in distinct aspects of plant development
The light perception in seedlings modulates the post-embryonic development, integrating phytohormone and other endogenous responses with environmental cues. A proper hypocotyl elongation at the first hours/days of development is critical for plant establishment and photosynthetic success. The precise coordination of hypocotyl elongation is under parallel and redundant regulatory pathways. Gene expression, at specific cells, modulated by photoreceptors and phytohormones, is the main regulatory mechanism at this phase. However, the integrative pathways controlling hypocotyl development are not completely elucidated. Our previous results showed that the miRNA156/SPL9 module is associated with hypocotyl elongation control. We hypothesized that this module acts as a regulatory hub in gibberellin (GA), brassinosteroid (BR), and light crosstalk. We already generated several data indicating that SPL9 expression is modulated by GA and BR, and perhaps by light during hypocotyl elongation. This project proposes to evaluate in greater detail the hypocotyl phenotype and transcriptome alterations in Arabidopsis miR156/SPL9 mutants and transgenic lines under distinct light regimes. The hypocotyl length from seedlings growing in high red/far-red, and low blue/UV-A light will show whether the phenotype observed in SPL9 gain or loss of function is dependent on the light quality. Moreover, transcriptome analyses of specific cells in the hypocotyl expansion zone under distinct light regimes will provide important insights into the SPL9 function as an activator and/or repressor of light responses. The use of SPL9-GFP transgenic plants will allow the discovery of direct targets by ChIP-qPCR. Finally, the combination of RNA-seq and ChIP-qPCR will allow the discovery of SPL9 direct targets, which may be modulated by BR, GA, and light in the hypocotyl.
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Distinct environments and latitudes limit human agricultural activities due to the photoperiodic response and its strong impact on flowering time and crop yield. The CONSTANS (CO) gene is the central regulatory hub in the photoperiodic pathway in several species. In the model plant
Arabidopsis thaliana, CO was identified as a transcriptional activator of the phloem-mobile florigen protein FLOWERING LOCUS T (FT). Although cultivated tomato (Solanum lycopersicum) is considered a neutral day plant, its wild relative species show a drastic delay in flowering under long day conditions, which can be associated with the activity of SELF-PRUNING 5G (SP5G) protein. SP5G represses the expression of the tomato FT homolog, SINGLE FLOWER TRUSS (SFT). We then hypothesized that a tomato CO homologue (SlCONSTANS1, SlCO1) positively regulates SP5G transcription under long days (LD), which was confirmed by our molecular and functional data. However, it is still unknown whether the activation of SP5G by SlCO1 is direct or indirect, as well as the role and post-translational regulation of SlCO1 protein. Our experiments suggest that photoperiodic pathway is negatively and positively regulated by DELLA and PHYTOCHROME B1 (PHYB1) proteins, respectively. In this context, this proposal aims to evaluate the direct activation of the SP5G gene by SlCO1, and the possible SlCO1 interaction with PROCERA and PHYB1 proteins, since both proteins post-translationally regulate CO activity in Arabidopsis. Additionally, the role of SlCO1 in tomato will be analyzed through characterization of slco1 mutant (e.g., co1CRISPR) and overexpression transgenic plants (e.g., p35S::SlCO1), already generated by our research group. Another goal is to identify common target genes among photoperiodic, gibberellin (GA) and age (miR156/SPLs) flowering pathways. For this, we will analyze transcriptomes from vegetative meristems in wild type and mutants in key genotypes of each pathway (photoperiodic, gibberellin and age). Understanding the photoperiodic responses and its interaction with gibberellin and age flowering pathways may be an important tool to guide tomato and other crops breeding programs.
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Molecular control of the plant architecture: interplay between microRNAs, transcription factors and phytohormones
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Fundamental questions in plant biology include how cells and tissues maintain their identity over time and how they switch from one stable state to the next. Understanding such basic questions may help us to better predict how plants modulate their shoot architecture in response to endogenous cues and the environment. Shoot architecture is one of the main developmental factors affecting plant productivity. Predictable, plants have evolved intricate regulatory networks to module their shoot architecture, which include phytohormones, transcription factor-regulated transcriptional programming, and epigenetic factors like microRNAs. However, how these different factors integrate to control shoot architecture at molecular and cellular levels is still unclear in most species. Following our findings that microRNAs (miRNAs) can regulate different aspects of sugarcane and tomato development by interacting with phytohormones, the characterization of miRNA-based patterning mechanisms and their association with phytohormones is a central aspect of our work. For example, we are studying the interaction between the gibberellin-negative regulator DELLA with miRNA-controlled pathways during flower and shoot development. Moreover, we are evaluating the interactions between phytohormones and microRNA modules in the control of axillary shoot branching, which directly influence shoot architecture. For each of the research areas, we intend to employ developmental genetics, genome editing (via CRISPR/Cas9-based technologies), next generation sequencing (NGS), imaging, and bioinformatics approaches. Moreover, we use Arabidopsis thaliana and tomato (Solanum lycopersicum) in our studies, as each model organism offers unique experimental advantages and because comparative studies can provide an evolutionary perspective on key genetic pathways associated with the establishment of the shoot architecture.
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Effects of chromatin remodelers on tomato development
Cells utilize diverse ATP-dependent nucleosome-remodelling complexes to carry out histone sliding, ejection or the incorporation of histone variants. Evidence accumulated over recent years has revealed that flowering time, flower development, gamete formation, and early seed development are under epigenetic regulatory control. Covalent modifications of the core components of chromatin, DNA, and histones provide a heritable mechanism for the regulation of gene expression. More specifically, SWI/SNF2 type ATP-dependent chromatin remodelers have an important role in flower development and reproduction, although most of the work thus far has been done in Arabidopsis. We are interested in understanding how these chromatin remodelers modulate reproductive development in crops. Once reproductive-associated genes which are targets for chromatin remodelers are identified, the knowledge acquired might help to design novel strategies for breeding programs intending to improve plant reproduction.
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MicroRNAs and early fruit and seed development
Parthenocarpy is the developmental process in which fruits develop in the absence of fertilization. It was shown by our research group that the miR159 module is crucial for ovule and seed development in tomato. The miR159 module comprises the microRNA159 and its targets, SlGAMYB1 and SlGAMYB2, which belong to the R2R3 MYB domain transcription factor family. Disruption of the miR159 module leads to defects in the embryo sac development and production of seedless fruits in tomato. We are continuing this work by expanding our findings in order to identify novel genetic and molecular networks regulated by the miR159 module during fruit set. Although this work was initially be done in tomato, we would like to expand the research to other important fruit-bearing crops, as well as to orphan crops.
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