To compensate for their sessile lifestyle, plants developed several responses to exogenous changes. One of the previously investigated and not yet fully understood adaptations occurs at the level of early subcellular trafficking, which needs to be rapidly adjusted to maintain cellular homeostasis and membrane integrity under osmotic stress conditions. To form a vesicle, the membrane needs to be deformed, which is ensured by multiple factors, including the activity of specific membrane proteins, such as flippases from the family of P4-ATPases. The membrane pumps actively translocate phospholipids from the exoplasmic/luminal to the cytoplasmic membrane leaflet to generate curvature, which might be coupled with recruitment of proteins involved in vesicle formation at specific sites of the donor membrane. We show that lack of the AMINOPHOSPHOLIPID ATPASE3 (ALA3) flippase activity caused defects at the plasma membrane and trans-Golgi network, resulting in altered endocytosis and secretion, processes relying on vesicle formation and movement. The mentioned cellular defects were translated into decreased intracellular trafficking flexibility failing to adjust the root growth on osmotic stress-eliciting media. In conclusion, we show that ALA3 cooperates with ARF-GEF BIG5/BEN1 and ARF1A1C/BEX1 in a similar regulatory pathway to vesicle formation, and together they are important for plant adaptation to osmotic stress.

• Klíčová slova
• Arabidopsis thaliana, ARF, GEF, endocytosis, flippase, osmotic stress, protein trafficking, secretion,
• MeSH
• Arabidopsis * metabolismus   MeSH
• biologický transport MeSH
• buněčná membrána metabolismus   MeSH
• membránové proteiny metabolismus   MeSH
• osmotický tlak MeSH
• proteiny huseníčku * genetika   metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• membránové proteiny MeSH
• proteiny huseníčku * MeSH

Phosphatidylinositol 4-kinases (PI4Ks) are the first enzymes that commit phosphatidylinositol into the phosphoinositide pathway. Here, we show that Arabidopsis thaliana seedlings deficient in PI4Kβ1 and β2 have several developmental defects including shorter roots and unfinished cytokinesis. The pi4kβ1β2 double mutant was insensitive to exogenous auxin concerning inhibition of root length and cell elongation; it also responded more slowly to gravistimulation. The pi4kß1ß2 root transcriptome displayed some similarities to a wild type plant response to auxin. Yet, not all the genes displayed such a constitutive auxin-like response. Besides, most assessed genes did not respond to exogenous auxin. This is consistent with data with the transcriptional reporter DR5-GUS. The content of bioactive auxin in the pi4kß1ß2 roots was similar to that in wild-type ones. Yet, an enhanced auxin-conjugating activity was detected and the auxin level reporter DII-VENUS did not respond to exogenous auxin in pi4kß1ß2 mutant. The mutant exhibited altered subcellular trafficking behavior including the trapping of PIN-FORMED 2 protein in rapidly moving vesicles. Bigger and less fragmented vacuoles were observed in pi4kß1ß2 roots when compared to the wild type. Furthermore, the actin filament web of the pi4kß1ß2 double mutant was less dense than in wild-type seedling roots, and less prone to rebuilding after treatment with latrunculin B. A mechanistic model is proposed in which an altered PI4K activity leads to actin filament disorganization, changes in vesicle trafficking, and altered auxin homeostasis and response resulting in a pleiotropic root phenotypes.

• MeSH
• Arabidopsis * metabolismus   MeSH
• fosfatidylinositolfosfáty MeSH
• fosfatidylinositoly metabolismus   MeSH
• kořeny rostlin genetika   metabolismus   MeSH
• kyseliny indoloctové metabolismus   MeSH
• proteiny huseníčku * genetika   metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• fosfatidylinositolfosfáty MeSH
• fosfatidylinositoly MeSH
• kyseliny indoloctové MeSH
• phosphatidylinositol 4-phosphate MeSH Prohlížeč
• proteiny huseníčku * MeSH

The root is the below-ground organ of a plant, and it has evolved multiple signaling pathways that allow adaptation of architecture, growth rate, and direction to an ever-changing environment. Roots grow along the gravitropic vector towards beneficial areas in the soil to provide the plant with proper nutrients to ensure its survival and productivity. In addition, roots have developed escape mechanisms to avoid adverse environments, which include direct illumination. Standard laboratory growth conditions for basic research of plant development and stress adaptation include growing seedlings in Petri dishes on medium with roots exposed to light. Several studies have shown that direct illumination of roots alters their morphology, cellular and biochemical responses, which results in reduced nutrient uptake and adaptability upon additive stress stimuli. In this review, we summarize recent methods that allow the study of shaded roots under controlled laboratory conditions and discuss the observed changes in the results depending on the root illumination status.

• Klíčová slova
• D-rootsystem, abiotic stress, auxin, cytokinin, dark-grown roots, direct root illumination, flavonols, light escape mechanism, reactive oxygen species, root growth,
• MeSH
• fyziologická adaptace * MeSH
• kořeny rostlin metabolismus   účinky záření   MeSH
• regulace genové exprese u rostlin účinky záření   MeSH
• rostlinné proteiny genetika   metabolismus   MeSH
• rostliny metabolismus   účinky záření   MeSH
• semenáček metabolismus   účinky záření   MeSH
• světlo * MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Názvy látek
• rostlinné proteiny MeSH

Eukaryotic cells rely on the accuracy and efficiency of vesicular traffic. In plants, disturbances in vesicular trafficking are well studied in quickly dividing root meristem cells or polar growing root hairs and pollen tubes. The development of the female gametophyte, a unique haploid reproductive structure located in the ovule, has received far less attention in studies of vesicular transport. Key molecules providing the specificity of vesicle formation and its subsequent recognition and fusion with the acceptor membrane are Rab proteins. Rabs are anchored to membranes by covalently linked geranylgeranyl group(s) that are added by the Rab geranylgeranyl transferase (RGT) enzyme. Here we show that Arabidopsis plants carrying mutations in the gene encoding the β-subunit of RGT (rgtb1) exhibit severely disrupted female gametogenesis and this effect is of sporophytic origin. Mutations in rgtb1 lead to internalization of the PIN1 and PIN3 proteins from the basal membranes to vesicles in provascular cells of the funiculus. Decreased transport of auxin out of the ovule is accompanied by auxin accumulation in tissue surrounding the growing gametophyte. In addition, female gametophyte development arrests at the uni- or binuclear stage in a significant portion of the rgtb1 ovules. These observations suggest that communication between the sporophyte and the developing female gametophyte relies on Rab-dependent vesicular traffic of the PIN1 and PIN3 transporters and auxin efflux out of the ovule.

• Klíčová slova
• Arabidopsis, PIN1, PIN3, Rab, auxin transport, female gametophyte, funiculus, ovule, rab geranylgeranyl transferase,
• MeSH
• Arabidopsis * genetika   MeSH
• kyseliny indoloctové MeSH
• proteiny huseníčku * genetika   MeSH
• pylová láčka MeSH
• vajíčko rostlin genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• kyseliny indoloctové MeSH
• proteiny huseníčku * MeSH

Plant growth and productivity are orchestrated by a network of signaling cascades involved in balancing responses to perceived environmental changes with resource availability. Vascular plants are divided into the shoot, an aboveground organ where sugar is synthesized, and the underground located root. Continuous growth requires the generation of energy in the form of carbohydrates in the leaves upon photosynthesis and uptake of nutrients and water through root hairs. Root hair outgrowth depends on the overall condition of the plant and its energy level must be high enough to maintain root growth. TARGET OF RAPAMYCIN (TOR)-mediated signaling cascades serve as a hub to evaluate which resources are needed to respond to external stimuli and which are available to maintain proper plant adaptation. Root hair growth further requires appropriate distribution of the phytohormone auxin, which primes root hair cell fate and triggers root hair elongation. Auxin is transported in an active, directed manner by a plasma membrane located carrier. The auxin efflux carrier PIN-FORMED 2 is necessary to transport auxin to root hair cells, followed by subcellular rearrangements involved in root hair outgrowth. This review presents an overview of events upstream and downstream of PIN2 action, which are involved in root hair growth control.

• Klíčová slova
• PIN-FORMED 2, ROP2, ROS, TOR signaling, auxin, plant adaptation, polar cell elongation, root growth, root hair growth,
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

Cell polarity is a fundamental feature of all multicellular organisms. PIN auxin transporters are important cell polarity markers that play crucial roles in a plethora of developmental processes in plants. Here, to identify components involved in cell polarity establishment and maintenance in plants, we performed a forward genetic screening of PIN2:PIN1-HA;pin2 Arabidopsis (Arabidopsis thaliana) plants, which ectopically express predominantly basally localized PIN1 in root epidermal cells, leading to agravitropic root growth. We identified the regulator of PIN polarity 12 (repp12) mutation, which restored gravitropic root growth and caused a switch in PIN1-HA polarity from the basal to apical side of root epidermal cells. Next Generation Sequencing and complementation experiments established the causative mutation of repp12 as a single amino acid exchange in Aminophospholipid ATPase3 (ALA3), a phospholipid flippase predicted to function in vesicle formation. repp12 and ala3 T-DNA mutants show defects in many auxin-regulated processes, asymmetric auxin distribution, and PIN trafficking. Analysis of quintuple and sextuple mutants confirmed the crucial roles of ALA proteins in regulating plant development as well as PIN trafficking and polarity. Genetic and physical interaction studies revealed that ALA3 functions together with the ADP ribosylation factor GTPase exchange factors GNOM and BIG3 in regulating PIN polarity, trafficking, and auxin-mediated development.

• MeSH
• ADP-ribosylační faktory metabolismus   MeSH
• Arabidopsis účinky léků   metabolismus   MeSH
• biologický transport účinky léků   MeSH
• brefeldin A farmakologie   MeSH
• buněčná membrána účinky léků   metabolismus   MeSH
• genetická epistáze účinky léků   MeSH
• GTP-fosfohydrolasy metabolismus   MeSH
• kyseliny indoloctové metabolismus   MeSH
• mutace genetika   MeSH
• proteiny huseníčku metabolismus   MeSH
• proteiny přenášející fosfolipidy metabolismus   MeSH
• tabák metabolismus   MeSH
• trans-Golgiho síť účinky léků   metabolismus   MeSH
• vazba proteinů účinky léků   MeSH
• výměnné faktory guaninnukleotidů metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• ADP-ribosylační faktory MeSH
• brefeldin A MeSH
• GTP-fosfohydrolasy MeSH
• kyseliny indoloctové MeSH
• proteiny huseníčku MeSH
• proteiny přenášející fosfolipidy MeSH
• výměnné faktory guaninnukleotidů MeSH

The fine tuning of hormone (e.g., auxin and gibberellin) levels and hormone signaling is required for maintaining normal embryogenesis. Embryo polarity, for example, is ensured by the directional movement of auxin that is controlled by various types of auxin transporters. Here, we present pieces of evidence for the auxin-gibberellic acid (GA) hormonal crosstalk during embryo development and the regulatory role of the Arabidopsis thaliana Calcium-Dependent Protein Kinase-Related Kinase 5 (AtCRK5) in this regard. It is pointed out that the embryogenesis of the Atcrk5-1 mutant is delayed in comparison to the wild type. This delay is accompanied with a decrease in the levels of GA and auxin, as well as the abundance of the polar auxin transport (PAT) proteins PIN1, PIN4, and PIN7 in the mutant embryos. We have previously showed that AtCRK5 can regulate the PIN2 and PIN3 proteins either directly by phosphorylation or indirectly affecting the GA level during the root gravitropic and hypocotyl hook bending responses. In this manuscript, we provide evidence that the AtCRK5 protein kinase can in vitro phosphorylate the hydrophilic loops of additional PIN proteins that are important for embryogenesis. We propose that AtCRK5 can govern embryo development in Arabidopsis through the fine tuning of auxin-GA level and the accumulation of certain polar auxin transport proteins.

• Klíčová slova
• Arabidopsis thaliana, Calcium-Dependent Protein Kinase-Related Kinase (CRK), GA3, auxin gradient, embryogenesis, polar auxin transport (PAT) proteins,
• MeSH
• Arabidopsis růst a vývoj   metabolismus   MeSH
• ELISA MeSH
• gibereliny analýza   metabolismus   MeSH
• klíčení * MeSH
• kyseliny indoloctové metabolismus   MeSH
• membránové transportní proteiny genetika   metabolismus   MeSH
• protein-serin-threoninkinasy metabolismus   MeSH
• proteiny huseníčku genetika   metabolismus   MeSH
• receptory buněčného povrchu metabolismus   MeSH
• regulace genové exprese u rostlin MeSH
• semena rostlinná anatomie a histologie   růst a vývoj   metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• CRK5 protein, Arabidopsis MeSH Prohlížeč
• gibberellic acid MeSH Prohlížeč
• gibereliny MeSH
• kyseliny indoloctové MeSH
• membránové transportní proteiny MeSH
• protein-serin-threoninkinasy MeSH
• proteiny huseníčku MeSH
• receptory buněčného povrchu MeSH

Arabidopsis PIN2 protein directs transport of the phytohormone auxin from the root tip into the root elongation zone. Variation in hormone transport, which depends on a delicate interplay between PIN2 sorting to and from polar plasma membrane domains, determines root growth. By employing a constitutively degraded version of PIN2, we identify brassinolides as antagonists of PIN2 endocytosis. This response does not require de novo protein synthesis, but involves early events in canonical brassinolide signaling. Brassinolide-controlled adjustments in PIN2 sorting and intracellular distribution governs formation of a lateral PIN2 gradient in gravistimulated roots, coinciding with adjustments in auxin signaling and directional root growth. Strikingly, simulations indicate that PIN2 gradient formation is no prerequisite for root bending but rather dampens asymmetric auxin flow and signaling. Crosstalk between brassinolide signaling and endocytic PIN2 sorting, thus, appears essential for determining the rate of gravity-induced root curvature via attenuation of differential cell elongation.

• MeSH
• Arabidopsis účinky léků   metabolismus   MeSH
• biologický transport účinky léků   MeSH
• brassinosteroidy metabolismus   farmakologie   MeSH
• endocytóza účinky léků   MeSH
• gravitropismus účinky léků   fyziologie   MeSH
• kořeny rostlin účinky léků   metabolismus   MeSH
• kyseliny indoloctové metabolismus   MeSH
• meristém účinky léků   metabolismus   MeSH
• proteiny huseníčku metabolismus   MeSH
• regulátory růstu rostlin metabolismus   farmakologie   MeSH
• signální transdukce MeSH
• steroidy heterocyklické metabolismus   farmakologie   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• brassinolide MeSH Prohlížeč
• brassinosteroidy MeSH
• kyseliny indoloctové MeSH
• PIN2 protein, Arabidopsis MeSH Prohlížeč
• proteiny huseníčku MeSH
• regulátory růstu rostlin MeSH
• steroidy heterocyklické MeSH

The plant-specific proteins named PIN-FORMED (PIN) efflux carriers facilitate the direction of auxin flow and thus play a vital role in the establishment of local auxin maxima within plant tissues that subsequently guide plant ontogenesis. They are membrane integral proteins with two hydrophobic regions consisting of alpha-helices linked with a hydrophilic loop, which is usually longer for the plasma membrane-localized PINs. The hydrophilic loop harbors molecular cues important for the subcellular localization and thus auxin efflux function of those transporters. The three-dimensional structure of PIN has not been solved yet. However, there are scattered but substantial data concerning the functional characterization of amino acid strings that constitute these carriers. These sequences include motifs vital for vesicular trafficking, residues regulating membrane diffusion, cellular polar localization, and activity of PINs. Here, we summarize those bits of information striving to provide a reference to structural motifs that have been investigated experimentally hoping to stimulate the efforts toward unraveling of PIN structure-function connections.

• Klíčová slova
• PIN efflux carriers, auxin transport, protein domains, sequence motifs, subcellular trafficking,
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

Coordination of plant development requires modulation of growth responses that are under control of the phytohormone auxin. PIN-FORMED plasma membrane proteins, involved in intercellular transport of the growth regulator, are key to the transmission of such auxin signals and subject to multilevel surveillance mechanisms, including reversible post-translational modifications. Apart from well-studied PIN protein modifications, namely phosphorylation and ubiquitylation, no further post-translational modifications have been described so far. Here, we focused on root-specific Arabidopsis PIN2 and explored functional implications of two evolutionary conserved cysteines, by a combination of in silico and molecular approaches. PIN2 sequence alignments and modeling predictions indicated that both cysteines are facing the cytoplasm and therefore would be accessible to redox status-controlled modifications. Notably, mutant pin2C-A alleles retained functionality, demonstrated by their ability to almost completely rescue defects of a pin2 null allele, whereas high resolution analysis of pin2C-A localization revealed increased intracellular accumulation, and altered protein distribution within plasma membrane micro-domains. The observed effects of cysteine replacements on root growth and PIN2 localization are consistent with a model in which redox status-dependent cysteine modifications participate in the regulation of PIN2 mobility, thereby fine-tuning polar auxin transport.

• Klíčová slova
• Arabidopsis, Auxin, PIN proteins, SRRF, intracellular distribution, plasma membrane protein sorting, protein mobility, protein modeling, root phenotype,
• MeSH
• Arabidopsis genetika   metabolismus   MeSH
• cystein genetika   MeSH
• konzervovaná sekvence * MeSH
• kořeny rostlin růst a vývoj   metabolismus   MeSH
• kyseliny indoloctové metabolismus   MeSH
• membránové mikrodomény metabolismus   MeSH
• proteiny huseníčku chemie   genetika   metabolismus   MeSH
• transport proteinů MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• cystein MeSH
• kyseliny indoloctové MeSH
• PIN2 protein, Arabidopsis MeSH Prohlížeč
• proteiny huseníčku MeSH