Synthetic hypomorphic alleles to study MUTE-dependent molecular transitions in stomatal development.
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PROVIDER: PRJNA1137830 | ENA |
REPOSITORIES: ENA
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Project description:Stomatal abundance sets the plant potential for gas exchange, impacting photosynthesis and transpiration, and ultimately, plant survival and growth. Stomata originate from cell lineages initiated by asymmetric divisions of protodermal cells, producing meristemoids that develop into guard cell pairs. The transcription factors SPEECHLESS, MUTE, and FAMA are essential for stomatal lineage development, sequentially driving cell division and differentiation events. Their absence produces stomataless epidermis, hindering analysis of their roles during lineage development. MUTE drives the transition from proliferating meristemoids to stomata commitment. We aim to explore the molecular mechanisms underlying MUTE activity, using partial loss-of-function alleles predicted to impair DNA-binding and potentially alter the expression of MUTE transcriptional targets. We engineered mutant allele coding sequences, generated Arabidopsis lines carrying these alleles and analyzed their epidermal and transcriptional phenotypes using microscopy and RNAseq. Synthetic alleles driven by the MUTE promoter rescued the stomataless phenotype of the seedling-lethal mute-3 mutant, enabling stomata differentiation and resulting in viable, fertile plants. Further examination of the developmental consequences of MUTE partial loss-of-function revealed arrested lineages, reduced stomatal abundance and altered stomatal spacing. Transcriptomic analysis of very young cotyledons from complemented lines indicated that some MUTE targets, but not others, require an intact MUTE bHLH domain. Comparison with existing lineage cell-specific transcriptional profiles showed that lineage development in the mutant lines was delayed compared to the wild-type but followed similar gene networks. These synthetic alleles provide new insight into MUTE ability to accurately and timely specify stomata formation.
2025-06-24 | GSE272620 | GEO
Project description:We have previously identified genes that are induced by MUTE, key transcription factor for stomatal lineage fate commitment and symmetric cell division that create stomata surrounded by a pair of guard cells in Arabidopsis. To test whether these genes are indeed direct MUTE targets, we performed genome-wide MUTE ChIP-sequencing.
2022-02-12 | GSE173338 | GEO
Project description:We initiated a study to investigate the transcriptional profiles associated with cell states of the stomatal lineage. A stem-cell like precursor of stomata, a meristemoid. reiterates asymmetric divisions and renews itself before differentiating into guard cells. The transient and asynchronous nature of the meristemoid has made it difficult to study its molecular characteristics. Through combinatorial use of genetic resources that either arrest or constitutively drive stomatal cell-state progressions due to loss- or gain-of-function mutations in the key transcription factor genes, SPEECHLESS, MUTE, and SCRM, we obtained seedlings highly enriched in pavement cells, meristemoids, or stomata. Here we present transcriptome and genome-wide trends in gene regulation associated with each cell state and identify molecular signatures associated with meristemoids. 12 samples are included in this study. Three biological replicates of 5-dag seedlings of speechless, scrm-D and scrm-D;mute were compared wild type seedlings for changes in gene expression.
2011-11-08 | E-GEOD-29814 | biostudies-arrayexpress
Project description:Stomata are pores in the epidermis of plants that can open and close and allow for gas exchange vital for photosynthesis and regulate transpiration. Stomatal development is driven by a set of conserved bHLH transcription factors (SPCH, MUTE, FAMA, and their heterodimerization partners ICE1, SCRM2), that initiate and promote progression of cell fates in the stomatal lineage. Due to the shared ancestry of SPCH, MUTE and FAMA (subgroup Ia) and ICE1 and SCRM2 (subgroup IIIb) their DNA binding specificity is similar and there is some functional redundancy. However, individual bHLHs also have unique functions. For example, SPCH is required for initiation of the stomatal lineage, while FAMA is responsible for terminal differentiation of the guard cell pair. In grasses, the stomatal complex comprises the guard cell pair, and two flanking subsidiary cells. Recruitment of the latter from neighboring cell filed during development requires expression of MUTE. Remarkably, while MUTE is absolutely required for the promotion of guard mother cell fate in maize and rice, this is not the case in Brachypodium. This suggests that another TF can at least partially substitute for MUTE in this function. While different expression profiles of the bHLH dimers within the stomatal lineage may partially responsible for distinct functions of each pair, it is likely that each pair forms different transcriptional complexes and that interaction with other transcriptional regulators affects the dimer’s binding DNA binding and gene regulation properties. Given the differences in bHLH function between dicots and grasses and even within the grass family, we were interested in elucidating the protein interaction networks of the stomatal lineage regulators in Brachypodium. To this end, we performed co-immunoprecipitation coupled to LC-MS/MS of BdSPCH2-YFP, YFP-BdMUTE, YFP-BdFAMA, YFP-ICE1 and YFP-SCRM2 from the developmental zone of young B. distachyon leaves using GFP-Trap beads. All YFP-fusion proteins were expressed under the endogenous promoter in the Bd21-3 background. The only exception was BdICE1, which was expressed under the ZmUBI promoter, but was nevertheless mostly restricted to the stomatal lineage. As control lines we use the Bd21-3 wild type and a line expressing 3x YFPnls (nuclear YFP) under the MUTE promoter. Comparison of proteins enriched with the bHLH fusion proteins vs. the controls revealed overlapping and distinct putative interactors, which is in agreement with the assumption that these transcription factors have both shared and unique functions. Notably, in addition to the presumed hetero-dimerization partners, we found a number of other bHLH transcription factors that were identified with one or more of the bait proteins. This suggests the presence of a larger bHLH network acting in the stomatal lineage.
2023-03-11 | PXD035582 | Pride
Project description:Purpose: Identify SLGC cell fate by transcriptomes Methods: Cells expressing SLGC markers (BASLp:BRX-YFP, BASLp:myrBRX-YFP and myrBRXp:myrBRX-YFP) and stomatal commitment stage marker (MUTEp:MUTE-YFP) were collected by FACS. 9dpg plants were used for protoplasting and subjected to cell sorting. Around 20,000/sample were cellected and used for library preparation. Results: SLGCs are renriched in mitotic genes. In addition, top candidates, DEK and MYB16 proteins function in SLGC cell divisions. Conclusions: SLGCs are division competent cells and the coordination of cell cycle and transcriptional regulation though DEK is important for maintaining the transition (SLGC) state during stomatal development.
2021-04-05 | GSE129938 | GEO
Project description:We initiated a study to investigate the transcriptional profiles associated with cell states of the stomatal lineage. A stem-cell like precursor of stomata, a meristemoid. reiterates asymmetric divisions and renews itself before differentiating into guard cells. The transient and asynchronous nature of the meristemoid has made it difficult to study its molecular characteristics. Through combinatorial use of genetic resources that either arrest or constitutively drive stomatal cell-state progressions due to loss- or gain-of-function mutations in the key transcription factor genes, SPEECHLESS, MUTE, and SCRM, we obtained seedlings highly enriched in pavement cells, meristemoids, or stomata. Here we present transcriptome and genome-wide trends in gene regulation associated with each cell state and identify molecular signatures associated with meristemoids.
2011-11-08 | GSE29814 | GEO
Project description:Adaptive evolution in response to cellular stress is a critical process implicated in a wide range of core biological and clinical phenomena. Two major routes of adaptation have been identified: non-genetic cellular plasticity, which allows expression of different phenotypes in novel environments, and genetic variation achieved by selection of fitter phenotypes. While these processes are now broadly accepted, their temporal and epistatic features in the context of cellular evolution and emerging drug resistance are contentious. In this project, we generated hypomorphic alleles of the essential nuclear pore complex (NPC) gene NUP58. By dissecting both early and long-term mechanisms of adaptation in independent clones, we observed that early physiological adaptation correlated with transcriptome rewiring and upregulation of genes known to interact with the NPC. In contrast, long-term adaptation and fitness recovery occurred via focal amplification of NUP58 and restoration of mutant protein expression. These data support the concept that plasticity-mediated and genetic routes can co-exist to enable cellular evolution, with early flexibility of phenotype allowing later acquisition of genetic adaptations to a specific impairment. We propose this approach as a genetic model to mimic targeted drug therapy in human cells and to dissect early and late mechanisms of adaptation. Targeting both mechanisms in parallel may reduce the emergence of drug resistance.
2021-11-02 | PXD022255 | Pride
Project description:Stomata in the plant epidermis play a vital role in growth and survival by controlling gas exchange and immunity to pathogens. A genetic frame of key transcriptional factors and cellular communication has been established, by which plants modulate stomatal cell fate and patterning. miRNAs contribute to functional and developmental plasticity in multicellular organisms. However, it remains very elusive as to whether miRNAs pitch in stomatal development. Here, we reveal dynamic miRNA expression profiles from stomatal lineage cells in a development stage-specific manner and show that stomatal lineage miRNAs positively and negatively regulate stomatal formation and pattern to avoid clustered and paired stomata. Target prediction of stomatal lineage miRNAs suggests potential cellular processes involved in stomatal development. Furthermore, dysregulation of stomatal lineage miRNAs and their target mRNAs disclose unexpected genetic pathways modulating stomatal development. Our study demonstrates that miRNAs constitute an additional layer in the complex regulatory mechanism of stomatal development.
2020-02-03 | GSE140918 | GEO
Project description:Precise cell division control is critical for developmental patterning. For the differentiation of a functional stoma, a cellular valve for efficient gas exchange, the single symmetric division of an immediate precursor is absolutely essential. Yet, the mechanism governing the single division event remains unclear. Here we report the complete inventories of gene expression by the Arabidopsis bHLH protein MUTE, a potent inducer of stomatal differentiation. We found that MUTE switches the gene expression program initiated by its sister bHLH, SPEECHLESS and directly induces a suite of cell-cycle genes and their transcriptional repressors. The architecture of the regulatory network initiated by MUTE represents an Incoherent Type 1 Feed-Forward Loop. Our study delineates the role for MUTE in orchestrating a transcriptional cascade leading to stomatal differentiation, and unravels a core regulatory circuit.
2019-11-16 | GSE107018 | GEO
Project description:Regulation of Stomatal Development by Stomatal Lineage miRNAs
| PRJNA591466 | ENA