The growth of Arabidopsis primary root is repressed by several and diverse amino acids through auxin-dependent and independent mechanisms and MPK6 kinase activity.

Amino acids serve as structural monomers for protein synthesis and are considered important biostimulants for plants. In this report, the effects of all 20-L amino acids in Arabidopsis primary root growth were evaluated. 15 amino acids inhibited growth, being l-leucine (l-Leu), l-lysine (l-Lys), l-tryptophan (l-Trp), and l-glutamate (l-Glu) the most active, which repressed both cell division and elongation in primary roots. Comparisons of DR5:GFP expression and growth of WT Arabidopsis seedlings and several auxin response mutants including slr, axr1 and axr2 single mutants, arf7/arf19 double mutant and tir1/afb2/afb3 triple mutant, treated with inhibitory concentrations of l-Glu, l-Leu, l-Lys and l-Trp revealed gene-dependent, specific changes in auxin response. In addition, l- isomers of Glu, Leu and Lys, but not l-Trp diminished the GFP fluorescence of pPIN1::PIN1:GFP, pPIN2::PIN2:GFP, pPIN3::PIN3:GFP and pPIN7::PIN7:GFP constructs in root tips. MPK6 activity in roots was enhanced by amino acid treatment, being greater in response to l-Trp while mpk6 mutants supported cell division and elongation at high doses of l-Glu, l-Leu, l-Lys and l-Trp. We conclude that independently of their auxin modulating properties, amino acids signals converge in MPK6 to alter the Arabidopsis primary root growth.

Stem cell ageing of the root apical meristem of Arabidopsis thaliana.

Plants form new organs from pluripotent stem cells throughout their lives and under changing environmental conditions. In the Arabidopsis root meristem, a pool of stem cells surrounding a stem cell organizer, named Quiescent Center (QC), gives rise to the specific root tissues. Among them, the columella stem cell niche that gives rise to the gravity-sensing columella cells has been used as a model system to study stem cell regulation at the young seedling stage. However, little is known about the changes of the stem cell niche during later development. Here, we report that the columella stem cell niche undergoes pronounced histological and molecular reorganization as the plant progresses towards the adult stage. Commonly-used reporters for cellular states undergo re-patterning after an initial juvenile meristem phase. Furthermore, the responsiveness to the plant hormone abscisic acid, an integrator of stress response, strongly decreases. Many ageing effects are reminiscent of the loss-of-function phenotype of the central stem cell regulator WOX5 and can be explained by gradually decreasing WOX5 expression levels during ageing. Our results show that the architecture and central regulatory components of the root stem cell niche are already highly dynamic within the first weeks of development.

Augmentation of root gravitropism by hypocotyl curvature in Brassica rapa seedlings.

Main Conclusion Root gravitropism of primary roots is assisted by curvature of the hypocotyl base. Root gravitropism is typically described as the sequence of signal perception, signal processing, and response that causes differential elongation and the establishment of a new gravitropic set-point angle. We describe two components of the graviresponse of Brassica seedlings that comprise a primary curvature of the root tip and a later onset but stronger curvature that occurs at the base of the hypocotyl. This second curvature is preceded by straightening of the tip region but leads to the completion of the alignment of the root axis. Curvature in both regions require a minimum displacement of 20 deg. The rate of tip curvature is a function of root length. After horizontal reorientation, tip curvature of five mm long roots curved twice as fast as 10 mm long roots (33.6 ±â€¯3.3 vs. 14.3 ±â€¯1.5 deg hr-1). The onset of curvature at the hypocotyl base is correlated with root length, but the rate of this curvature is independent of seedling length. Decapping of roots prevented tip curvature but the curvature at base of hypocotyl was unaffected. Endodermal cells at the root shoot junction show numerous, large and sedimenting amyloplasts, which likely serve as gravity sensors (statoliths). The amyloplasts at the hypocotyl were 3-4 µm in diameter, similar in size to those in the root cap, and twice the size of starch grains in the cortical layers of hypocotyl or elsewhere in the root. These data indicate that the root shoot reorientation of young seedlings is not limited to the root tip but includes more than one gravitropically responsive region.

Root extracellular traps versus neutrophil extracellular traps in host defence, a case of functional convergence?

The root cap releases cells that produce massive amounts of mucilage containing polysaccharides, proteoglycans, extracellular DNA (exDNA) and a variety of antimicrobial compounds. The released cells - known as border cells or border-like cells - and mucilage secretions form networks that are defined as root extracellular traps (RETs). RETs are important players in root immunity. In animals, phagocytes are some of the most abundant white blood cells in circulation and are very important for immunity. These cells combat pathogens through multiple defence mechanisms, including the release of exDNA-containing extracellular traps (ETs). Traps of neutrophil origin are abbreviated herein as NETs. Similar to phagocytes, plant root cap-originating cells actively contribute to frontline defence against pathogens. RETs and NETs are thus components of the plant and animal immune systems, respectively, that exhibit similar compositional and functional properties. Herein, we describe and discuss the formation, molecular composition and functional similarities of these similar but different extracellular traps.

The Root Cap Cuticle: A Cell Wall Structure for Seedling Establishment and Lateral Root Formation.

The root cap surrounding the tip of plant roots is thought to protect the delicate stem cells in the root meristem. We discovered that the first layer of root cap cells is covered by an electron-opaque cell wall modification resembling a plant cuticle. Cuticles are polyester-based protective structures considered exclusive to aerial plant organs. Mutations in cutin biosynthesis genes affect the composition and ultrastructure of this cuticular structure, confirming its cutin-like characteristics. Strikingly, targeted degradation of the root cap cuticle causes a hypersensitivity to abiotic stresses during seedling establishment. Furthermore, lateral root primordia also display a cuticle that, when defective, causes delayed outgrowth and organ deformations, suggesting that it facilitates lateral root emergence. Our results show that the previously unrecognized root cap cuticle protects the root meristem during the critical phase of seedling establishment and promotes the efficient formation of lateral roots.

The build-up of osmotic stress responses within the growing root apex using kinematics and RNA-sequencing.

Molecular regulation of growth must include spatial and temporal coupling of cell production and cell expansion. The underlying mechanisms, especially under environmental challenge, remain obscure. Spatial patterns of cell processes make the root apex well suited to deciphering stress signaling pathways, and to investigating both processes. Kinematics and RNA-sequencing were used to analyze the immediate growth response of hydroponically grown Populus nigra cuttings submitted to osmotic stress. About 7400 genes and unannotated transcriptionally active regions were differentially expressed between the division and elongation zones. Following the onset of stress, growth decreased sharply, probably due to mechanical effects, before recovering partially. Stress impaired cell expansion over the apex, progressively shortened the elongation zone, and reduced the cell production rate. Changes in gene expression revealed that growth reduction was mediated by a shift in hormone homeostasis. Osmotic stress rapidly elicited auxin, ethylene, and abscisic acid. When growth restabilized, transcriptome remodeling became complex and zone specific, with the deployment of hormone signaling cascades, transcriptional regulators, and stress-responsive genes. Most transcriptional regulations fit growth reduction, but stress also promoted expression of some growth effectors, including aquaporins and expansins Together, osmotic stress interfered with growth by activating regulatory proteins rather than by repressing the machinery of expansive growth.

Root cap-dependent gravitropic U-turn of maize root requires light-induced auxin biosynthesis via the YUC pathway in the root apex.

Gravitropism refers to the growth or movement of plants that is influenced by gravity. Roots exhibit positive gravitropism, and the root cap is thought to be the gravity-sensing site. In some plants, the root cap requires light irradiation for positive gravitropic responses. However, the mechanisms regulating this phenomenon are unknown. We herein report that maize roots exposed to white light continuously for ≥1-2h show increased indole-3-acetic acid (IAA) levels in the root tips, especially in the transition zone (1-3mm from the tip). Treatment with IAA biosynthesis inhibitors yucasin and l-kynurenine prevented any increases in IAA content and root curvature under light conditions. Analyses of the incorporation of a stable isotope label from tryptophan into IAA revealed that some of the IAA in roots was synthesized in the root apex. Furthermore, Zmvt2 and Zmyuc gene transcripts were detected in the root apex. One of the Zmyuc genes (ZM2G141383) was up-regulated by light irradiation in the 0-1mm tip region. Our findings suggest that IAA accumulation in the transition zone is due to light-induced activation of Zmyuc gene expression in the 0-1mm root apex region. Light-induced changes in IAA levels and distributions mediate the maize root gravitropic U-turn.

The root cap: a short story of life and death.

Over 130 years ago, Charles Darwin recognized that sensory functions in the root tip influence directional root growth. Modern plant biology has unravelled that many of the functions that Darwin attributed to the root tip are actually accomplished by a particular organ-the root cap. The root cap surrounds and protects the meristematic stem cells at the growing root tip. Due to this vanguard position, the root cap is predisposed to receive and transmit environmental information to the root proper. In contrast to other plant organs, the root cap shows a rapid turnover of short-lived cells regulated by an intricate balance of cell generation, differentiation, and degeneration. Thanks to these particular features, the root cap is an excellent developmental model system, in which generation, differentiation, and degeneration of cells can be investigated in a conveniently compact spatial and temporal frame. In this review, we give an overview of the current knowledge and concepts of root cap biology, focusing on the model plant Arabidopsis thaliana.

PCaP2 regulates nuclear positioning in growing Arabidopsis thaliana root hairs by modulating filamentous actin organization.

KEY MESSAGE: PCaP2 plays a key role in maintaining the nucleus at a relatively fixed distance from the apex during root hair growth by modulating actin filaments. During root hair growth, the nucleus localizes at a relatively fixed distance from the apex. In Arabidopsis thaliana, the position of the nucleus is mainly dependent on the configuration of microfilaments (filamentous actin). However, the mechanisms underlying the regulation of actin dynamics and organization for nuclear positioning are largely unknown. In the present study, we demonstrated that plasma membrane-associated Ca(2+) binding protein 2 (PCaP2) influences the position of the nucleus during root hair growth. Abnormal expression of PCaP2 in pcap2 and PCaP2 over-expression plants led to the disorganization of actin filaments, rather than microtubules, in the apex and sub-apical regions of root hairs, which resulted in aberrant root hair growth patterns and misplaced nuclei. Analyses using a PCaP2 mutant protein revealed that actin-severing activity is essential for the function of PCaP2 in root hairs. We demonstrated that PCaP2 plays a key role in maintaining nuclear position in growing root hairs by modulating actin filaments.

Novel roles of hydrogen peroxide (H2O2) in regulating pectin synthesis and demethylesterification in the cell wall of rice (Oryza sativa) root tips.

Hydrogen peroxide (H2O2) has been reported to increase lignin formation, enhance cell wall rigidification, restrict cell expansion and inhibit root elongation. However, our results showed that it not only inhibited rice (Oryza sativa) root elongation, but also increased root diameter. No study has reported how and why H2O2 increases cell expansion and root diameter. Exogenous H2O2 and its scavenger 4-hydroxy-Tempo were applied to confirm the roles of H2O2. Immunofluorescence, fluorescence probe, ruthenium red staining, histological section and spectrophotometry were used to monitor changes in the degree of pectin methylesterification, pectin content, pectin methylesterase (PME) activity and H2O2 content. Exogenous H2O2 inhibited root elongation, but increased cell expansion and root diameter significantly. H2O2 not only increased the region of pectin synthesis and pectin content in root tips, but also increased PME activity and pectin demethylesterification. The scavenger 4-hydroxy-Tempo reduced root H2O2 content and recovered H2O2-induced increases in cell expansion and root diameter by inhibiting pectin synthesis, PME activity and pectin demethylesterification. H2O2 plays a novel role in the regulation of pectin synthesis, PME activity and pectin demethylesterification. H2O2 increases cell expansion and root diameter by increasing pectin content and demethylesterification.

Programmed cell death controlled by ANAC033/SOMBRERO determines root cap organ size in Arabidopsis.

BACKGROUND: The root cap is a plant organ that ensheathes the meristematic stem cells at the root tip. Unlike other plant organs, the root cap shows a rapid cellular turnover, balancing constant cell generation by specific stem cells with the disposal of differentiated cells at the root cap edge. This cellular turnover is critical for the maintenance of root cap size and its position around the growing root tip, but how this is achieved and controlled in the model plant Arabidopsis thaliana remains subject to contradictory hypotheses. RESULTS: Here, we show that a highly organized cell death program is the final step of lateral root cap differentiation and that preparation for cell death is transcriptionally controlled by ANAC033/SOMBRERO. Precise timing of cell death is critical for the elimination of root cap cells before they fully enter the root elongation zone, which in turn is important in order to allow optimal root growth. Root cap cell death is followed by a rapid cell-autonomous corpse clearance and DNA fragmentation dependent on the S1-P1 type nuclease BFN1. CONCLUSIONS: Based on these results, we propose a novel concept in plant development that recognizes programmed cell death as a mechanism for maintaining organ size and tissue homeostasis in the Arabidopsis root cap.

Role of HSP101 in the stimulation of nodal root development from the coleoptilar node by light and temperature in maize (Zea mays L.) seedlings.

Nodal roots (NRs) constitute the prevalent root system of adult maize plants. NRs emerge from stem nodes located below or above ground, and little is known about their inducing factors. Here, it is shown that precocious development of NRs at the coleoptilar node (NRCNs) occurred in maize seedlings when: (i) dark grown and stimulated by the concurrent action of a single light shock of low intensity white light (2 µmol m(-2) s(-1)) and a single heat shock; (ii) grown under a photoperiod of low intensity light (0.1 µmol m(-2) s(-1)); or (iii) grown in the dark under a thermoperiod (28 °C/34 °C). The light shock effects were synergistic with heat shock and with the photoperiod, whereas the thermoperiodical and photoperiodical effects were additive. Dissection of the primary root or the root cap, to mimic the fatal consequences of severe heat shock, caused negligible effects on NRCN formation, indicating that the shoot is directly involved in perception of the heat shock-inducible signal that triggered NRCN formation. A comparison between hsp101-m5::Mu1/hsp101-m5::Mu1 and Hsp101/Hsp101 seedlings indicated that the heat shock protein 101 (HSP101) chaperone inhibited NRCN formation in the light and in the dark. Stimulation of precocious NRCN formation by light and heat shocks was affected by genetic background and by the stage of seedling development. HSP101 protein levels increased in the coleoptilar node of induced wild-type plants, particularly in the procambial region, where NRCN formation originated. The adaptive relevance of development of NRCNs in response to these environmental cues and hypothetical mechanisms of regulation by HSP101 are discussed.

Root cap angle and gravitropic response rate are uncoupled in the Arabidopsis pgm-1 mutant.

The sedimentation of starch-filled plastids is thought to be the primary mechanism by which gravity is perceived in roots. Following gravity perception, auxin redistribution toward the lower flank of roots, initiated in the root cap, is believed to play a role in regulation of the gravity response. Amyloplast sedimentation and auxin flux, however, have never been directly linked. The overall aim of this study was to investigate the relationship among plastid sedimentation, gravitropism and auxin flux. Our data show that pgm-1 roots respond to gravity at one-third the rate of wild-type (WT) roots. Maintaining the root tip at a constant angle using image analysis coupled to a rotating stage resulted in a constant rate of response regardless of the angle of tip orientation in pgm-1 mutants, in contrast to the responses of WT and pin3-1 mutants, which showed increasing response rates as the tip was constrained at greater angles. To indirectly visualize auxin flux following reorientation, we generated a pgm-1 mutant line expressing the DR5::GFPm reporter gene. In WT roots a GFP gradient was observed with a maximum along the lower flank, whereas pgm-1 roots formed a GFP maximum in the central columella but lacked any observable gradient up to 6 h following reorientation. Our study suggests that the relationship between root cap angle and gravitropic response depends upon plastid sedimentation-based gravity sensing and supports the idea that there are multiple, overlapping sensory response networks involved in gravitropism.

Involvement of Arabidopsis thaliana phospholipase Dzeta2 in root hydrotropism through the suppression of root gravitropism.

Root hydrotropism is the phenomenon of directional root growth toward moisture under water-deficient conditions. Although physiological and genetic studies have revealed the involvement of the root cap in the sensing of moisture gradients, and those of auxin and abscisic acid (ABA) in the signal transduction for asymmetric root elongation, the overall mechanism of root hydrotropism is still unclear. We found that the promoter activity of the Arabidopsis phospholipase Dzeta2 gene (PLDzeta2) was localized to epidermal cells in the distal root elongation zone and lateral root cap cells adjacent to them, and that exogenous ABA enhanced the activity and extended its area to the entire root cap. Although pldzeta2 mutant root caps did not exhibit a morphological phenotype in either the absence or presence of exogenous ABA, the inhibitory effect of ABA on gravitropism, which was significant in wild-type roots, was not observed in pldzeta2 mutant roots. In root hydrotropism experiments, pldzeta2 mutations significantly retarded or disturbed root hydrotropic responses. A drought condition similar to that used in a hydrotropism experiment enhanced the PLDzeta2 promoter activity in the root cap, as did exogenous ABA. These results suggest that PLDzeta2 responds to drought through ABA signaling in the root cap and accelerates root hydrotropism through the suppression of root gravitropism.

[Micromorphometric analysis of root cap cells in Allium cepa].

A two-dimensional micromorphometric analysis of a root cap in Allium cepa included the measurement of the areas of about six thousand cell profiles on both longitudinal and transversal sections. Basing on the results of this analysis, quantitative descriptions of two cell populations from the columella and periferic part of the root cap have been created. The scheme of arrangement of root cap cells, located at a distance of 250 microm or less from a root tip, has been developed.

D'orenone blocks polarized tip growth of root hairs by interfering with the PIN2-mediated auxin transport network in the root apex.

SUMMARY: The C(18) ketone (5E,7E)-6-methyl-8-(2,6,6-trimethylcyclohex-1-enyl)octa-5,7-dien-2-one (D'orenone) has been postulated to be an early cleavage product of beta-carotene en route to trisporic acids; these act as morphogenetic factors during the sexual reproduction of zygomycetes. Here we report that D'orenone blocks the highly polarized tip growth of root hairs, causing tip growth to stop completely within a few minutes. Importantly, external auxin reverses the effects of D'orenone on root hairs. Further analysis revealed that D'orenone lowers the auxin concentration in trichoblasts via PIN2-mediated auxin efflux to below the critical levels essential for root hair growth. D'orenone specifically increases PIN2 protein abundance without affecting PIN2 transcripts, and the PIN2 expression domain enlarges and shifts basipetally, resulting in more active auxin transport. The observation that D'orenone does not interfere with the root hair growth in roots of null mutant lines provides additional evidence that PIN2 is its specific target.

The root cap determines ethylene-dependent growth and development in maize roots.

Besides providing protection against mechanical damage to the root tip, the root cap is involved in the perception and processing of diverse external and internal stimuli resulting in altered growth and development. The transduction of these stimuli includes hormonal signaling pathways such as those of auxin, ethylene and cytokinin. Here, we show that the root cap is essential for the ethylene-induced regulation of elongation growth and root hair formation in maize. Exogenously applied ethylene is no longer able to inhibit elongation growth when the root cap has been surgically removed prior to hormone treatment. Reconstitution of the cap positively correlates with the developing capacity of the roots to respond to ethylene again. In contrast, the removal of the root cap does not per se affect growth inhibition controlled by auxin and cytokinin. Furthermore, our semi-quantitative RT-PCR results support earlier findings that the maize root cap is a site of high gene expression activity with respect to sensing and responding to hormones such as ethylene. From these data, we propose a novel function of the root cap which is the establishment of competence to respond to ethylene in the distal zones of the root.

[The structural characteristics and symplastic transport function of the Ectodesmata-like of root cap tissue in Zea mays].

The ultrastructure and symplastic transport function of Ectodesmata-like (ED-like) of the root cap cells of Zea mays, during the detaching stage, were reported by using fluorescence and electron microscopy. It was described the process that plasmodesmata (PD) were gradually stretched and changed into ED-like. It was discovered that the diameter of appressed endoplasmic reticulum (AER) in PD became thinner while the ED-like still remained some structures of PD. By using fluorescence probe incubating, 457Da Lucifer Yellow (LYCH) which was impermeable to the membrane, could enter the root cap cells through ED-like. The results proved that ED-like still retained physiological activity and kept the symplastic transport function during a period time. When the root tissues were pre-treated by cytochalasin D (CD), Phalloidin and 2,3-butanedione, 2-monoxime (BDM) and then combined with fluorescence probe detecting, the results showed that F-actin and myosin might take part in the regulation of the substance translocation of the ED-like.

Comparative study of cellular structures implicated in gravisensing in statocytes of primary and lateral roots of Vigna angularis.

The cellular structures of statocytes implicated in gravisensing in primary and lateral roots of Vigna angularis were compared. The statocytes of lateral roots already had small amyloplasts immediately after they emerged from the primary root. Although these amyloplasts sedimented, the lateral roots showed much weaker gravitropism than primary roots, at least until they reached a length of about 30 mm. The nuclei were usually positioned in the upper end of the statocytes in both types of roots. Electron microscopic surveys showed that many tubular elements of endoplasmic reticulum (ER) were frequently localized in the lower end of the statocyte and they sometimes diverged or curved, suggesting that the ER forms a large reticulate complex. It is worth noting that statocytes with a large ER complex were found much more frequently in primary roots than in lateral roots. The amyloplasts were not always settled on this complex but were very frequently under it, especially in the primary roots. In lateral roots, they were usually localized under the ER complex when they were present. Thus, it is suggested that the differential development and organization of the amyloplast-ER complex system is involved in the differential gravitropism of the two types of roots.

The production and release of living root cap border cells is a function of root apical meristem type in dicotyledonous angiosperm plants.

BACKGROUND AND AIMS: The root apical meristems (RAM) of flowering plant roots are organized into recognizable pattern types. At present, there are no known ecological or physiological benefits to having one RAM organization type over another. Although there are phylogenetic distribution patterns in plant groups, the possible evolutionary advantages of different RAM organization patterns are not understood. Root caps of many flowering plant roots are known to release living border cells into the rhizosphere, where the cells are believed to have the capacity to alter conditions in the soil and to interact with soil micro-organisms. Consequently, high rates of border cell production may have the potential to benefit plant growth and development greatly, and to provide a selective advantage in certain soil environments. This study reports the use of several approaches to elucidate the anatomical and developmental relationships between RAM organization and border cell production. METHODS: RAM types from many species were compared with numbers of border cells released in those species. In addition, other species were grown, fixed and sectioned to verify their organization type and capacity to produce border cells. Root tips were examined microscopically to characterize their pattern and some were stained to determine the viability of root cap cells. KEY RESULTS: The first report of a correlation between RAM organization type and the production and release of border cells is provided: species exhibiting open RAM organization produce significantly more border cells than species exhibiting closed apical organization. Roots with closed apical organization release peripheral root cap cells in sheets or large groups of dead cells, whereas root caps with open organization release individual living border cells. CONCLUSIONS: This study, the first to document a relationship between RAM organization, root cap behaviour and a possible ecological benefit to the plant, may yield a framework to examine the evolutionary causes for the diversification of RAM organization types across taxa.