RESUMO
Plants undergo rapid developmental transitions, which occur contemporaneously with gradual changes in physiology. Moreover, individual plants within a population undergo developmental transitions asynchronously. Single-plant-omics has the potential to distinguish between transcriptional events that are associated with these binary and continuous processes. Furthermore, we can use single-plant-omics to order individual plants by their intrinsic biological age, providing a high-resolution transcriptional time series. We performed RNA-seq on leaves from a large population of wild-type Arabidopsis (Arabidopsis thaliana) during the vegetative-to-reproductive transition. Though most transcripts were differentially expressed between bolted and unbolted plants, some regulators were more closely associated with leaf size and biomass. Using a pseudotime inference algorithm, we determined that some senescence-associated processes, such as the reduction in ribosome biogenesis, were evident in the transcriptome before a bolt was visible. Even in this near-isogenic population, some variants are associated with developmental traits. These results support the use of single-plant-omics to uncover rapid transcriptional dynamics by exploiting developmental asynchrony.
RESUMO
Mature plant leaves are a composite of distinct cell types, including epidermal, mesophyll, and vascular cells. Notably, the proportion of these cells and the relative transcript concentrations within different cell types may change over time. While gene expression data at a single-cell level can provide cell-type-specific expression values, it is often too expensive to obtain these data for high-resolution time series. Although bulk RNA-seq can be performed in a high-resolution time series, RNA-seq using whole leaves measures average gene expression values across all cell types in each sample. In this study, we combined single-cell RNA-seq data with time-series data from whole leaves to assemble an atlas of cell-type-specific changes in gene expression over time for Arabidopsis (Arabidopsis thaliana). We inferred how the relative transcript concentrations of different cell types vary across diurnal and developmental timescales. Importantly, this analysis revealed 3 subgroups of mesophyll cells with distinct temporal profiles of expression. Finally, we developed tissue-specific gene networks that form a community resource: an Arabidopsis Leaf Time-dependent Atlas (AraLeTa). This allows users to extract gene networks that are confirmed by transcription factor-binding data and specific to certain cell types at certain times of day and at certain developmental stages. AraLeTa is available at https://regulatorynet.shinyapps.io/araleta/.
Assuntos
Arabidopsis , Regulação da Expressão Gênica de Plantas , Folhas de Planta , Arabidopsis/genética , Arabidopsis/metabolismo , Folhas de Planta/genética , Folhas de Planta/metabolismo , Folhas de Planta/crescimento & desenvolvimento , Ritmo Circadiano/genética , Redes Reguladoras de Genes , Perfilação da Expressão Gênica , Transcriptoma/genéticaRESUMO
Circadian clocks are endogenous timekeeping mechanisms that coordinate internal physiological responses with the external environment. EARLY FLOWERING3 (ELF3), PSEUDO RESPONSE REGULATOR (PRR9), and PRR7 are essential components of the plant circadian clock and facilitate entrainment of the clock to internal and external stimuli. Previous studies have highlighted a critical role for ELF3 in repressing the expression of PRR9 and PRR7. However, the functional significance of activity in regulating circadian clock dynamics and plant development is unknown. To explore this regulatory dynamic further, we first employed mathematical modeling to simulate the effect of the prr9/prr7 mutation on the elf3 circadian phenotype. These simulations suggested that simultaneous mutations in prr9/prr7 could rescue the elf3 circadian arrhythmia. Following these simulations, we generated all Arabidopsis elf3/prr9/prr7 mutant combinations and investigated their circadian and developmental phenotypes. Although these assays could not replicate the results from the mathematical modeling, our results have revealed a complex epistatic relationship between ELF3 and PRR9/7 in regulating different aspects of plant development. ELF3 was essential for hypocotyl development under ambient and warm temperatures, while PRR9 was critical for root thermomorphogenesis. Finally, mutations in prr9 and prr7 rescued the photoperiod-insensitive flowering phenotype of the elf3 mutant. Together, our results highlight the importance of investigating the genetic relationship among plant circadian genes.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Relógios Circadianos , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Relógios Circadianos/genética , Ritmo Circadiano/genética , Regulação da Expressão Gênica de Plantas , Fenômenos Fisiológicos Vegetais , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
BACKGROUND: Historically, disabled people have been marginalised in research that traditionally adopted a medical model perspective. Since the 1970's, there has been a shift from research on disabled people to research with disabled people with a strong emphasis on co-produced participatory research. Co-production involves disabled people working with academics to produce research and outcomes which are informed by the end user. This paper reflects on the role and experiences of peer researchers in co-producing a recent UK-wide research project called 'Getting our Voices Heard'. This project sought to identify the best approaches for people with a learning disability and their supporting organisations to influence adult safeguarding policies, across the four jurisdictions of the UK. METHODS: A co-produced participatory design was used to address the project aims; achieved through the establishment of a collaborative research team comprising academic researchers, key stakeholders and six peer researchers, each of whom had a learning disability. Semi-structured interviews were completed with senior policy makers. Following this, in each of the four Nations, an organisational case-study was completed (four in total). Organisations were purposively sampled to identify one organisation in each country which was recognised as being successful in influencing adult safeguarding policy. Data were gathered through focus groups discussions and semi-structured interviews with key stakeholders. Findings were developed into an Impact Strategy and Action Plan. Co-production methods were used throughout. RESULTS: Six individuals with a learning disability were recruited and trained to work as peer researchers, involved at key stages of the project, working alongside a wider research team. The role and experiences of the peer researchers in the context of policy are presented. Peer researchers provided largely positive first-hand accounts of their experiences. The importance of collaboration, the invaluable role of Learning Disability support organisations, and the need for additional time and resources to facilitate co-production, was noted. CONCLUSION: Whilst peer researchers were positive about their experiences, some success in promoting co-produced research and areas for improvement were evident. Collaboration at all stages would have been strengthened with research funding which enabled involvement of all team members in all research activities.
Since the 1970's, there has been a shift from research on disabled people to research with disabled people. This is often referred to as co-produced research. Co-production has a wide definition but includes disabled people working with academics to produce research and outcomes which neither group could achieve by working in isolation. This paper presents the co-production methodology used to conduct a research project called 'Getting our Voices Heard'. This project, sought to find the best way to get the voices of people with a learning disability heard inpolicy development in the UK.We explain how this research was carried out, using a co-produced participatory design. We established a research team with professional researchers from a university, who worked with peer researchers. Peer researchers are people who have lived experience of the issue being studied. In this project, we worked with six peer researchers who all had a learning disability.The experiences of the peer researchers, and ways in which the peer researchers were involved at each stage of the project are discussed. The peer researchers described feeling positive about their role and felt involved. We show that working together is important and recommend that additional time and resources are essential for this joint working.