The CLAVATA pathway is a key regulator of stem cell function in the multicellular shoot tips of Arabidopsis, where it acts via the WUSCHEL transcription factor to modulate hormone homeostasis. Broad-scale evolutionary comparisons have shown that CLAVATA is a conserved regulator of land plant stem cell function, but CLAVATA acts independently of WUSCHEL-like (WOX) proteins in bryophytes. The relationship between CLAVATA, hormone homeostasis and the evolution of land plant stem cell functions is unknown. Here we show that in the moss, Physcomitrella (Physcomitrium patens), CLAVATA affects stem cell activity by modulating hormone homeostasis. CLAVATA pathway genes are expressed in the tip cells of filamentous tissues, regulating cell identity, filament branching, plant spread and auxin synthesis. The receptor-like kinase PpRPK2 plays the major role, and Pprpk2 mutants have abnormal responses to cytokinin, auxin and auxin transport inhibition, and show reduced expression of PIN auxin transporters. We propose a model whereby PpRPK2 modulates auxin gradients in filaments to determine stem cell identity and overall plant form. Our data indicate that CLAVATA-mediated auxin homeostasis is a fundamental property of plant stem cell function, probably exhibited by the last shared common ancestor of land plants.

• Keywords
• CLAVATA, CLV-WUS, evo-devo, moss filament identity, physcomitrella, plant stem cell,
• MeSH
• Bryophyta * metabolism   MeSH
• Homeostasis MeSH
• Stem Cells metabolism   MeSH
• Indoleacetic Acids metabolism   MeSH
• Bryopsida * genetics   metabolism   MeSH
• Arabidopsis Proteins * genetics   metabolism   MeSH
• Gene Expression Regulation, Plant MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Indoleacetic Acids MeSH
• Arabidopsis Proteins * MeSH

The proper distribution of the hormone auxin is essential for plant development. It is channeled by auxin efflux carriers of the PIN family, typically asymmetrically located on the plasma membrane (PM). Several studies demonstrated that some PIN transporters are also located at the endoplasmic reticulum (ER). From the PM-PINs, they differ in a shorter internal hydrophilic loop, which carries the most important structural features required for their subcellular localization, but their biological role is otherwise relatively poorly known. We discuss how ER-PINs take part in maintaining intracellular auxin homeostasis, possibly by modulating the internal levels of IAA; it seems that the exact identity of the metabolites downstream of ER-PINs is not entirely clear as well. We further review the current knowledge about their predicted structure, evolution and localization. Finally, we also summarize their role in plant development.

• Keywords
• ER-PINs, PIN proteins, PIN5, PIN8, auxin metabolism, auxin transport,
• Publication type
• Journal Article MeSH
• Review MeSH

The trafficking of subcellular cargos in eukaryotic cells crucially depends on vesicle budding, a process mediated by ARF-GEFs (ADP-ribosylation factor guanine nucleotide exchange factors). In plants, ARF-GEFs play essential roles in endocytosis, vacuolar trafficking, recycling, secretion, and polar trafficking. Moreover, they are important for plant development, mainly through controlling the polar subcellular localization of PIN-FORMED transporters of the plant hormone auxin. Here, using a chemical genetics screen in Arabidopsis thaliana, we identified Endosidin 4 (ES4), an inhibitor of eukaryotic ARF-GEFs. ES4 acts similarly to and synergistically with the established ARF-GEF inhibitor Brefeldin A and has broad effects on intracellular trafficking, including endocytosis, exocytosis, and vacuolar targeting. Additionally, Arabidopsis and yeast (Saccharomyces cerevisiae) mutants defective in ARF-GEF show altered sensitivity to ES4. ES4 interferes with the activation-based membrane association of the ARF1 GTPases, but not of their mutant variants that are activated independently of ARF-GEF activity. Biochemical approaches and docking simulations confirmed that ES4 specifically targets the SEC7 domain-containing ARF-GEFs. These observations collectively identify ES4 as a chemical tool enabling the study of ARF-GEF-mediated processes, including ARF-GEF-mediated plant development.

• MeSH
• Arabidopsis drug effects   genetics   metabolism   MeSH
• Brefeldin A pharmacology   MeSH
• Cell Membrane drug effects   metabolism   MeSH
• Chromones chemistry   pharmacology   MeSH
• DNA-Binding Proteins genetics   metabolism   MeSH
• Endocytosis drug effects   MeSH
• Plants, Genetically Modified MeSH
• Membrane Glycoproteins genetics   metabolism   MeSH
• Membrane Transport Proteins genetics   metabolism   MeSH
• Mutation MeSH
• Protein Domains MeSH
• Arabidopsis Proteins genetics   metabolism   MeSH
• Saccharomyces cerevisiae Proteins genetics   metabolism   MeSH
• Saccharomyces cerevisiae drug effects   metabolism   MeSH
• Molecular Docking Simulation MeSH
• Transcription Factors genetics   metabolism   MeSH
• Protein Transport drug effects   MeSH
• Guanine Nucleotide Exchange Factors chemistry   genetics   metabolism   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• ARF1 protein, Arabidopsis MeSH Browser
• Brefeldin A MeSH
• Chromones MeSH
• DNA-Binding Proteins MeSH
• GNL1 protein, Arabidopsis MeSH Browser
• GNOM protein, Arabidopsis MeSH Browser
• Membrane Glycoproteins MeSH
• Membrane Transport Proteins MeSH
• PIN1 protein, Arabidopsis MeSH Browser
• Arabidopsis Proteins MeSH
• Saccharomyces cerevisiae Proteins MeSH
• SEC12 protein, S cerevisiae MeSH Browser
• Transcription Factors MeSH
• Guanine Nucleotide Exchange Factors MeSH

A comparative approach in biology is needed to assess the universality of rules governing this discipline. In plant telomere research, most of the key principles were established based on studies in only single model plant, Arabidopsis thaliana. These principles include the absence of telomere shortening during plant development and the corresponding activity of telomerase in dividing (meristem) plant cells. Here we examine these principles in Physcomitrella patens as a representative of lower plants. To follow telomerase expression, we first characterize the gene coding for the telomerase reverse transcriptase subunit PpTERT in P. patens, for which only incomplete prediction has been available so far. In protonema cultures of P. patens, growing by filament apical cell division, the proportion of apical (dividing) cells was quantified and telomere length, telomerase expression and activity were determined. Our results show telomere stability and demonstrate proportionality of telomerase activity and expression with the number of apical cells. In addition, we analyze telomere maintenance in mre11, rad50, nbs1, ku70 and lig4 mutants of P. patens and compare the impact of these mutations in double-strand-break (DSB) repair pathways with earlier observations in corresponding A. thaliana mutants. Telomere phenotypes are absent and DSB repair kinetics is not affected in P. patens mutants for DSB factors involved in non-homologous end joining (NHEJ). This is compliant with the overall dominance of homologous recombination over NHEJ pathways in the moss, contrary to the inverse situation in flowering plants.

• MeSH
• Arabidopsis genetics   MeSH
• Chromosomes, Plant genetics   MeSH
• DNA, Plant genetics   MeSH
• DNA Breaks, Double-Stranded MeSH
• Phenotype MeSH
• Phylogeny MeSH
• Telomere Homeostasis genetics   MeSH
• Homologous Recombination MeSH
• Bryopsida genetics   metabolism   MeSH
• Molecular Sequence Data MeSH
• Mutation MeSH
• DNA Repair * MeSH
• Plant Proteins genetics   metabolism   MeSH
• Amino Acid Sequence MeSH
• Base Sequence MeSH
• Sequence Analysis, DNA MeSH
• Sequence Alignment MeSH
• Telomerase genetics   metabolism   MeSH
• Telomere genetics   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• DNA, Plant MeSH
• Plant Proteins MeSH
• Telomerase MeSH

Shoot branching is a primary contributor to plant architecture, evolving independently in flowering plant sporophytes and moss gametophytes. Mechanistic understanding of branching is largely limited to flowering plants such as Arabidopsis, which have a recent evolutionary origin. We show that in gametophytic shoots of Physcomitrella, lateral branches arise by re-specification of epidermal cells into branch initials. A simple model co-ordinating the activity of leafy shoot tips can account for branching patterns, and three known and ancient hormonal regulators of sporophytic branching interact to generate the branching pattern- auxin, cytokinin and strigolactone. The mode of auxin transport required in branch patterning is a key divergence point from known sporophytic pathways. Although PIN-mediated basipetal auxin transport regulates branching patterns in flowering plants, this is not so in Physcomitrella, where bi-directional transport is required to generate realistic branching patterns. Experiments with callose synthesis inhibitors suggest plasmodesmal connectivity as a potential mechanism for transport.

• Keywords
• Physcomitrella, apical dominance, branching, developmental biology, gametophyte, plant biology, stem cells,
• MeSH
• Models, Biological MeSH
• Biological Transport drug effects   MeSH
• Cytokinins biosynthesis   MeSH
• Plant Epidermis cytology   growth & development   MeSH
• Plants, Genetically Modified MeSH
• Indoleacetic Acids metabolism   pharmacology   MeSH
• Lactones pharmacology   MeSH
• Bryopsida drug effects   growth & development   MeSH
• Morphogenesis drug effects   MeSH
• Mutation genetics   MeSH
• Gene Expression Regulation, Plant drug effects   MeSH
• Plant Growth Regulators pharmacology   MeSH
• Plant Proteins metabolism   MeSH
• Body Patterning drug effects   MeSH
• Plant Shoots drug effects   growth & development   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Cytokinins MeSH
• Indoleacetic Acids MeSH
• Lactones MeSH
• Plant Growth Regulators MeSH
• Plant Proteins MeSH