Project description:The precise onset of flowering is crucial to ensure successful plant reproduction. The gene FLOWERING LOCUS T (FT) encodes florigen, a mobile signal that is produced in leaves and initiates flowering in the shoot apical meristem. FT is highly expressed in phloem companion cells that reside in distal leaf regions. Thus far, a detailed molecular characterization of the FT-expressing phloem companion cells has been lacking. Here, we used single-nuclei RNA-seq to investigate gene expression in FT-expressing cells and other phloem companion cells. We discovered that companion cells with high FT expression form a unique cluster and express other genes encoding small proteins, including the anti-florigen BROTHER OF FT (BFT). The promoters of FT and the genes co-expressed with FT were enriched for the consensus binding motif of NITRATE-INDUCIBLE GARP-TYPE TRANSCRIPTIONAL REPRESSOR1 (NIGT1). Overexpression of the paralogs NIGT1.2 and NIGT1.4 in transgenic Arabidopsis lines significantly delayed flowering under nitrogen-rich conditions, consistent with NIGT1s acting as nitrogen-dependent FT repressors. Taken together, our results demonstrate that FT-expressing cells show a distinct expression profile that suggests that these cells produce multiple systemic signals to regulate plant growth and development.
Project description:Leaves are asymmetric, with differential functionalization of abaxial and adaxial tissues. The bundle sheath (BS) surrounding the vasculature of the C3 crop barley is dorsoventrally differentiated into three domains: adaxial structural, lateral S-type, and abaxial L-type. S-type cells seem to transfer assimilates towards the phloem. Here we used single-cell RNA sequencing to investigate BS differentiation in C4 maize. Abaxial BS (abBS) cells of rank-2 intermediate veins specifically expressed three SWEET sucrose uniporters (SWEET13a, b, and c) and UmamiT amino acid efflux transporters. SWEET13a, b, c were also identified in the phloem parenchyma (PP). Thus maize acquired a unique mechanism for phloem loading in which abBS cells provide the main pathway for apoplasmic sucrose transfer towards the phloem. This pathway predominates in veins responsible for phloem loading (rank-2 intermediate), while rank-1 intermediate and major veins export sucrose from the phloem parenchyma (PP) adjacent to the sieve element companion cell (SE/CC) complex, as in Arabidopsis. We surmise that abBS identity is subject to dorsoventral patterning and has components of PP identity. The observations provide first insights into the unique properties of abBS cells, cells previously considered to fulfill the same functions as other bundle sheath cells (BSCs), and a basis for understanding the C4 syndrome.
Project description:Leaves are asymmetric, with differential functionalization of abaxial and adaxial tissues. The bundle sheath (BS) surrounding the vasculature of the C3 crop barley is dorsoventrally differentiated into three domains: adaxial structural, lateral S-type, and abaxial L-type. S-type cells seem to transfer assimilates towards the phloem. Here we used single-cell RNA sequencing to investigate BS differentiation in C4 maize. Abaxial BS (abBS) cells of rank-2 intermediate veins specifically expressed three SWEET sucrose uniporters (SWEET13a, b, and c) and UmamiT amino acid efflux transporters. SWEET13a, b, c were also identified in the phloem parenchyma (PP). Thus maize acquired a unique mechanism for phloem loading in which abBS cells provide the main pathway for apoplasmic sucrose transfer towards the phloem. This pathway predominates in veins responsible for phloem loading (rank-2 intermediate), while rank-1 intermediate and major veins export sucrose from the phloem parenchyma (PP) adjacent to the sieve element companion cell (SE/CC) complex, as in Arabidopsis. We surmise that abBS identity is subject to dorsoventral patterning and has components of PP identity. The observations provide first insights into the unique properties of abBS cells, cells previously considered to fulfill the same functions as other bundle sheath cells (BSCs), and a basis for understanding the C4 syndrome.
Project description:The vascular system of plants consists of xylem, phloem and procambium in a specific pattern. The phloem consists of sieve elements, the apparatus for bulk flow of photo-assimilates, and companion cells, which mediate transport of photo-assimilates between the sieve elements and surrounding cells and support the biological activities of the sieve element cells. The regulatory mechanisms of vascular development are being uncovered. Here we show that PHLOEM EARLY DOFs (PEARs) and related genes (collectively phloem-Dofs) not only regulate the number of procambium cell files, but also positively regulate phloem differentiation. Overexpression of phloem-Dofs induced cells that expressed either sieve element or companion cell marker genes, which are mutually exclusive. Conversely, disruption of phloem-Dofs caused loss of phloem. Phloem-Dofs induce CLAVATA3/EMBRYO SURROUNDING REGION-RELATED25 (CLE25) and CLE26 peptides, which in turn inhibit expression of phloem-Dofs and phloem formation, forming a negative feedback loop. Disruption of multiple genes for either phloem-expressed CLEs, BARELY ANY MERISTEM (BAM)-class receptors, or their coreceptors, CLAVATA3 INSENSITIVE RECEPTOR KINASEs (CIKs), caused excess formation of phloem cell files. We further show that phloem-Dofs are under positive self and mutual regulation. These positive and negative feedback loops create the proper phloem pattern.
Project description:The leaf vasculature plays a key role in solute translocation. Veins consist of at least seven distinct cell types, with specific roles in transport, metabolism, and signaling. Little is known about the vascular cells in leaves, in particular the phloem parenchyma (PP). PP effluxes sucrose into the apoplasm as a basis for phloem loading; yet PP has only been characterized microscopically. Here, we enriched vascular cells from Arabidopsis leaves to generate a single-cell transcriptome atlas of leaf vasculature. We identified ?19 cell clusters, encompassing epidermis, guard cells, hydathodes, mesophyll, and all vascular cell types, and used metabolic pathway analysis to define their roles. Clusters comprising PP cells were enriched for transporters, including SWEET11 and SWEET12 sucrose and UmamiT amino acid efflux carriers. PP development occurs independently from APL, a transcription factor required for phloem differentiation. PP cells have a unique pattern of amino acid metabolism activity distinct from companion cells (CC), explaining differential distribution/metabolism of amino acids in veins. The kinship relation of the vascular clusters is strikingly similar to the vein morphology, except for a clear separation of CC from the other vascular cells including PP. In summary, our scRNA-seq analysis provides a wide range of information into the leaf vasculature and the role and relationship of the leaf cell types.
Project description:The leaf vasculature plays a key role in solute translocation. Veins consist of at least seven distinct cell types, with specific roles in transport, metabolism, and signaling. Little is known about the vascular cells in leaves, in particular the phloem parenchyma (PP). PP effluxes sucrose into the apoplasm as a basis for phloem loading; yet PP has only been characterized microscopically. Here, we enriched vascular cells from Arabidopsis leaves to generate a single-cell transcriptome atlas of leaf vasculature. We identified ?19 cell clusters, encompassing epidermis, guard cells, hydathodes, mesophyll, and all vascular cell types, and used metabolic pathway analysis to define their roles. Clusters comprising PP cells were enriched for transporters, including SWEET11 and SWEET12 sucrose and UmamiT amino acid efflux carriers. PP development occurs independently from APL, a transcription factor required for phloem differentiation. PP cells have a unique pattern of amino acid metabolism activity distinct from companion cells (CC), explaining differential distribution/metabolism of amino acids in veins. The kinship relation of the vascular clusters is strikingly similar to the vein morphology, except for a clear separation of CC from the other vascular cells including PP. In summary, our scRNA-seq analysis provides a wide range of information into the leaf vasculature and the role and relationship of the leaf cell types.
Project description:A florigen-expressing subpopulation of phloem companion cells expresses other small proteins and reveals a nitrogen-sensitive FT repressor
Project description:Plant organs are comprised of distinct cell types with unique assemblages of mRNAs. This is a collection of CEL files of mRNA profiles of the total steady-state mRNAs and polysomal mRNAs of distinct cell types of the whole root and shoot of 7-d-old Arabidopsis thaliana seedlings. The cell type specific mRNA populations are those present in ribosome-mRNA complexes. This sub-population of mRNAs was obtained by first establishing a collection of Arabidopsis lines that express a FLAG-epitope tagged ribosomal protein L18 (RPL18) directed by promoters expressed in specific cell types and regions. Thirteen different promoter:FLAG-RPL18 lines were used. The targeted cell types and promoters included root atrichoblast (non-hair) epidermal cells (pGL2), root endodermis (pSCR), root stelar xylem and pericycle (pWOL, pSHR), root phloem companion cells (phloem CC) (pSUC2, pSultr2;2), root proliferating cells (pRPL11C), root cortex meristematic cells (pCO2), root cortex elongation/maturation cells (pPEP), shoot mesophyll (pRBCS), shoot epidermis (pCER5), shoot guard cells (pKAT1), shoot bundle sheath (pSultr2;2), shoot phloem CC (pSUC2) and shoot trichomes (pGL2). A CaMV 35S promoter:FLAG-RPL18 line was used to obtain the polysomal mRNA of multiple cell types. The immunopurification of ribosome-mRNA complexes of specific cell types/regions was accomplished by the method described in Zanetti et al. (Plant Physiology, 138, 624-635; 2005). Hybridization of the immunopurified mRNAs to the Affymetrix ATH1 DNA microarray platform and subsequent data analysis permitted the identification of transcripts that are enriched or depleted in specific cell types/regions of roots and shoots. The dataset includes samples from cell types/regions from seedlings grown under control conditions and cell types/regions of seedlings exposed to low oxygen stress (hypoxia) for 2 h. Keywords: cell-type specific expression, hypoxic stress, polysomal mRNA, abiotic stress, atrichoblasts, epidermis, cortex, endodermis, stele, phloem companion cells, guard cells, mesophyll
Project description:We investigated the chromatin modifications H3K4me3 and H3K27me3 in the A. thaliana shoot phloem companion cells using INTACT reporter lines. Samples were collected in two biological replications.