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Keith Roberts

Publications and source records attributed to Keith Roberts.

At least 19 recordsLinked to original sources
PubMed2007

The biology of arabinogalactan proteins.

Arabinogalactan proteins is an umbrella term applied to a highly diverse class of cell surface glycoproteins, many of which contain glycosylphosphatidylinositol lipid anchors. The structures of protein and glycan moieties of arabinogalactan proteins are overwhelmingly diverse while the "hydroxproline contiguity hypothesis" predicts arabinogalactan modification of members of many families of extracellular proteins. Descriptive studies using monoclonal antibodies reacting with carbohydrate epitopes on arabinogalactan proteins and experimental work using beta-Yariv reagent implicate arabinogalactan proteins in many biological processes of cell proliferation and survival, pattern formation and growth, and in plant microbe interaction. Advanced structural understanding of arabinogalactan proteins and an emerging molecular genetic definition of biological roles of individual arabinogalactan protein species, in conjunction with potentially analogous extracellular matrix components of animals, stimulate hypotheses about their mode of action. Arabinogalactan proteins might be soluble signals, or might act as modulators and coreceptors of apoplastic morphogens; their amphiphilic molecular nature makes them prime candidates of mediators between the cell wall, the plasma membrane, and the cytoplasm.

Apoptosis↗
PubMed2006

Interactions between MUR10/CesA7-dependent secondary cellulose biosynthesis and primary cell wall structure.

Primary cell walls are deposited and remodeled during cell division and expansion. Secondary cell walls are deposited in specialized cells after the expansion phase. It is presently unknown whether and how these processes are interrelated. The Arabidopsis (Arabidopsis thaliana) MUR10 gene is required for normal primary cell wall carbohydrate composition in mature leaves as well as for normal plant growth, hypocotyl strength, and fertility. The overall sugar composition of young mur10 seedlings is not significantly altered; however, the relative proportion of pectin side chains is shifted toward an increase in 1 --> 5-alpha-arabinan relative to 1 --> 4-beta-galactan. mur10 seedlings display reduced fucogalactosylation of tightly cell wall-bound xyloglucan. Expression levels of genes encoding either nucleotide sugar interconversion enzymes or glycosyl transferases, known to be involved in primary and secondary cell wall biosynthesis, are generally unaffected; however, the CesA7 transcript is specifically suppressed in the mur10-1 allele. The MUR10 locus is identical with the CesA7 gene, which encodes a cellulose catalytic subunit previously thought to be specifically involved in secondary cell wall formation. The xylem vessels in young mur10 hypocotyls are collapsed and their birefringence is lost. Moreover, a fucogalactosylated xyloglucan epitope is reduced and a 1 --> 5-alpha-arabinan epitope increased in every cell type in mur10 hypocotyls, including cells that do not deposit secondary walls. mur10 also displays altered distribution of an arabinogalactan-protein epitope previously associated with xylem differentiation and secondary wall thickening. This work indicates the existence of a mechanism that senses secondary cell wall integrity and controls biosynthesis or structural remodeling of primary cell walls and cellular differentiation.

Alleles↗
PubMed2006

Distinct properties of the five UDP-D-glucose/UDP-D-galactose 4-epimerase isoforms of Arabidopsis thaliana.

Plant genomes contain genetically encoded isoforms of most nucleotide sugar interconversion enzymes. Here we show that Arabidopsis thaliana has five genes encoding functional UDP-D-glucose/UDP-D-galactose 4-epimerase (named UGE1 to UGE5). All A. thaliana UDP-d-glucose 4-epimerase isoforms are dimeric in solution, maximally active in vitro at 30-40 degrees C, and show good activity between pH 7 and pH 9. In vitro, UGE1, -3, and -5 act independently of externally added NAD+, whereas cofactor addition stimulates the activity of UGE2 and is particularly important for UGE4 activity. UGE1 and UGE3 are most efficiently inhibited by UDP. The five isoforms display kcatUDP-Gal values between 23 and 128 s(-1) and KmUDP-Gal values between 0.1 and 0.3 mm. This results in enzymatic efficiencies ranging between 97 and 890 mm(-1) s(-1) for UGE4 = UGE1 < UGE3 < UGE5 < UGE2. The KmUDP-Glc values, derived from the Haldane relationship, were 0.76 mm for UGE1, 0.56 mm for UGE4, and between 0.13 and 0.23 mm for UGE2, -3, and -5. The expression of UGE isoforms is ubiquitous and displays developmental and cell type-dependent variations. UGE1 and -3 expression patterns globally resemble enzymes involved in carbohydrate catabolism, and UGE2, -4, and -5 expression is more related to carbohydrate biosynthesis. UGE1, -2, and -4 are present in the cytoplasm, whereasUGE4 is additionally enriched close to Golgi stacks. All UGE genes tested complement the UGE4rhd1 phenotype, confer increased galactose tolerance in planta, and complement the galactose metabolization deficiency in the Saccharomyces cerevisiae gal10 mutant. We suggest that plant UGE isoforms function in different metabolic situations and that enzymatic properties, gene expression pattern, and subcellular localization contribute to the differentiation of isoform function.

Arabidopsis↗
PubMed2006

Arabidopsis SPO11-2 functions with SPO11-1 in meiotic recombination.

The Spo11 protein is a eukaryotic homologue of the archaeal DNA topoisomerase VIA subunit (topo VIA). In archaea it is involved, together with its B subunit (topo VIB), in DNA replication. However, most eukaryotes, including yeasts, insects and vertebrates, instead have a single gene for Spo11/topo VIA and no homologues for topo VIB. In these organisms, Spo11 mediates DNA double-strand breaks that initiate meiotic recombination. Many plant species, in contrast to other eukaryotes, have three homologues for Spo11/topo VIA and one for topo VIB. The homologues in Arabidopsis, AtSPO11-1, AtSPO11-2 and AtSPO11-3, all share 20-30% sequence similarity with other Spo11/topo VIA proteins, but their functional relationship during meiosis or other processes is not well understood. Previous genetic evidence suggests that AtSPO11-1 is a true orthologue of Spo11 in other eukaryotes and is required for meiotic recombination, whereas AtSPO11-3 is involved in DNA endo-reduplication as a part of the topo VI complex. In this study, we show that plants homozygous for atspo11-2 exhibit a severe sterility phenotype. Both male and female meiosis are severely disrupted in the atspo11-2 mutant, and this is associated with severe defects in synapsis during the first meiotic division and reduced meiotic recombination. Further genetic analysis revealed that AtSPO11-1 and AtSPO11-2 genetically interact, i.e. plants heterozygous for both atspo11-1 and atspo11-2 are also sterile, suggesting that AtSPO11-1 and AtSPO11-2 have largely overlapping functions. Thus, the three Arabidopsis Spo11 homologues appear to function in two discrete processes, i.e. AtSPO11-1 and AtSPO11-2 in meiotic recombination and AtSPO11-3 in DNA replication.

Alleles↗
PubMed2005

RHL1 is an essential component of the plant DNA topoisomerase VI complex and is required for ploidy-dependent cell growth.

How cells achieve their final sizes is a pervasive biological question. One strategy to increase cell size is for the cell to amplify its chromosomal DNA content through endoreduplication cycles. Although endoreduplication is widespread in eukaryotes, we know very little about its molecular mechanisms. Successful progression of the endoreduplication cycle in Arabidopsis requires a plant homologue of archaeal DNA topoisomerase (topo) VI. To further understand how DNA is endoreduplicated and how this process is regulated, we isolated a dwarf Arabidopsis mutant, hyp7 (hypocotyl 7), in which various large cell types that in the wild type normally endoreduplicate multiple times complete only the first two rounds of endoreduplication and stall at 8C. HYP7 encodes the RHL1 (ROOT HAIRLESS 1) protein, and sequence analysis reveals that RHL1 has similarity to the C-terminal domain of mammalian DNA topo IIalpha, another type II topo that shares little sequence homology with topo VI. RHL1 shows DNA binding activity in vitro, and we present both genetic and in vivo evidence that RHL1 forms a multiprotein complex with plant topo VI. We propose that RHL1 plays an essential role in the topo VI complex to modulate its function and that the two distantly related topos, topo II and topo VI, have evolved a common domain that extends their function. Our data suggest that plant topo II and topo VI play distinct but overlapping roles during the mitotic cell cycle and endoreduplication cycle.

Amino Acid Sequence↗
PubMed2005

A fasciclin-domain containing gene, ZeFLA11, is expressed exclusively in xylem elements that have reticulate wall thickenings in the stem vascular system of Zinnia elegans cv Envy.

The vascular cylinder of the mature stem of Zinnia elegans cv Envy contains two anatomically distinct sets of vascular bundles, stem bundles and leaf-trace bundles. We isolated a full-length cDNA of ZeFLA11, a fasciclin-domain-containing gene, from a zinnia cDNA library derived from in vitro cultures of mesophyll cells induced to form tracheary elements. Using RNA in situ hybridization, we show that ZeFLA11 is expressed in the differentiating xylem vessels with reticulate type wall thickenings and adjacent parenchyma cells of zinnia stem bundles, but not in the leaf-trace bundles that deposit spiral thickenings. Our results suggest a function for this cell-surface GPI-anchored glycoprotein in secondary wall deposition during differentiation of metaxylem tissue with reticulate vessels.

Amino Acid Sequence↗
PubMed2005

A RING domain gene is expressed in different cell types of leaf trace, stem, and juvenile bundles in the stem vascular system of zinnia.

The in vitro zinnia (Zinnia elegans) mesophyll cell system, in which leaf mesophyll cells are induced to transdifferentiate into tracheary elements with high synchrony, has become an established model for studying xylogenesis. The architecture of the stem vascular system of zinnia cv Envy contains three anatomically distinct vascular bundles at different stages of development. Juvenile vascular strands of the subapical region develop into mature vascular strands with leaf trace segments and stem segments. Characteristic patterns of gene expression in juvenile, leaf trace, and stem bundles are revealed by a molecular marker, a RING domain-encoding gene, ZeRH2.1, originally isolated from a zinnia cDNA library derived from differentiating in vitro cultures. Using RNA in situ hybridization, we show that ZeRH2.1 is expressed preferentially in two specific cell types in mature zinnia stems. In leaf trace bundles, ZeRH2.1 transcript is abundant in xylem parenchyma cells, while in stem bundles it is abundant in phloem companion cells. Both of these cell types show wall ingrowths characteristic of transfer cells. In addition, ZeRH2.1 transcript is abundant in some phloem cells of juvenile bundles and in leaf palisade parenchyma. The complex and developmentally regulated expression pattern of ZeRH2.1 reveals heterogeneity in the vascular anatomy of the zinnia stem. We discuss a potential function for this gene in intercellular transport processes.

Amino Acid Sequence↗
PubMed2004

Growth regulators and the control of nucleotide sugar flux.

A small number of plant growth regulators are involved in the control of cell expansion. Despite knowledge of some of their signal transduction cascades, surprisingly little is known of how basic cell expansion-related processes, such as cell wall biosynthesis, are affected during growth. The Arabidopsis (Arabidopsis thaliana) mutant root hair defective1 (rhd1) lacks a functional UDP-glucose 4-epimerase gene, UGE4, which is involved in channeling UDP-D-galactose (UDP-D-Gal) into cell wall polymers. Here, we use rhd1 as a genetic model to analyze the physiological and genetic controls of nucleotide sugar flux. We find that ethylene specifically suppresses all visible aspects of the rhd1 phenotype. The ethylene-triggered suppression of rhd1 is negatively regulated by CONSTITUTIVE TRIPLE RESPONSE1 and requires the function of the wild-type genes ETHYLENE INSENSITIVE2 (EIN2), EIN4, AUXIN-RESISTENT1, and ETHYLENE-INSENSITIVE ROOT1 but does not depend on the activity of wild-type ETHYLENE RECEPTOR1 or EIN3 genes, highlighting the nonlinearity of ethylene signal transduction. Ethylene does not induce the expression of alternative UGE genes but, instead, suppresses the expression of two isoforms, UGE1 and UGE3, in a tissue-specific manner. Ethylene restores the biosynthesis of galactose-containing xyloglucan and arabinosylated galactan cell wall polymers in rhd1 back to wild-type levels. However, the dependence on UGE4 of pectic (1-->4)-beta-D-galactan and glucuronosyl-modified AGP biosynthesis is exacerbated. Our data suggest that ethylene and auxin together participate in the flux control of UDP-D-Gal into cell wall polymers and that the genetic control of this process is qualitatively distinct from previously described responses to ethylene.

Arabidopsis↗
PubMed2004

The expression patterns of arabinogalactan-protein AtAGP30 and GLABRA2 reveal a role for abscisic acid in the early stages of root epidermal patterning.

In the Arabidopsis root, patterning of the epidermal cell types is position-dependent. The epidermal cell pattern arises early during root development, and can be visualized using reporter genes driven by the GLABRA (GL)2 promoter as markers. The GL2 gene is preferentially expressed in the differentiating hairless cells (atrichoblasts) during a period in which epidermal cell identity is believed to be established. We show that AtAGP30 is also expressed in atrichoblasts. This gene encodes an arabinogalactan-protein (AGP) that is known to play a role in root regeneration and increases abscisic acid (ABA)-response rates. Although the expression level of this gene is regulated by the plant growth factors ABA and ethylene, only ABA was found to affect the tissue-specific pattern of expression. ABA also disrupts the expression pattern of the GL2::GUS (beta-glucuronidase) reporter gene. Our results indicate that ABA regulates epidermal cell-type-specific gene expression in the meristematic zone of the Arabidopsis root, while ethylene is known to act at later stages of epidermal differentiation. Despite its effects on the early stages of root epidermal patterning, ABA does not affect root hair formation on mature wild-type epidermal cells, suggesting that other developmental cues, like positional information, can progressively over-ride the ABA-mediated disruption of early epidermal patterning.

Abscisic Acid↗
PubMed2003

"Big it up": endoreduplication and cell-size control in plants.

Cells undergoing endoreduplication replicate chromosomal DNA without intervening mitoses. The resulting larger, higher-ploidy nucleus is often associated with an increase in cell size, but the molecular basis for this correlation remains poorly understood. Recent advances in characterising various mutants and transgenic plants are beginning to unravel how this unique type of cell cycling is regulated and how it contributes to cell-size control. Both cell growth (i.e. increase in cytoplasmic macromolecular mass) and cell expansion (i.e. increase in cell volume through vacuolation) contribute independently to increases in cell size in plants. A total organ-size checkpoint may also help to coordinate cell size and cell number within an organ, and can contribute to final cell-size determination in plants.

Cell Cycle↗
PubMed2003

How do you size a nasopharyngeal airway.

OBJECTIVE: To measure an appropriately sized nasopharyngeal airway, it is taught that the size is related to the patients little finger or nostril (anterior nares). This study has been designed to identify whether these comparisons are valid. METHOD: Direct comparison of the dimensions of ten subjects' little fingers and anterior nares with the internal anatomy of their nose as visualised on coronal MRI scans. RESULTS: Neither method correlated statistically with the nasal anatomy of that subject. CONCLUSIONS: The methods used traditionally to size a nasopharyngeal airway do not correlate with the airway anatomy and are unreliable. It is more appropriate to size the airway dependent upon the patient's size, sex and race.

Adult↗
PubMed2003

A review of emergency equipment carried and procedures performed by UK front line paramedics.

OBJECTIVES: To assess which items of resuscitation equipment are carried on United Kingdom (UK) front line ambulances and what procedures paramedics are able to perform. To compare these findings with those of a previous survey in 1997. METHOD: Postal survey to the chief executives of all the UK ambulance services and direct comparison with the data from 1997. RESULTS: Nasopharyngeal airway usage (NPA) has increased (21-55%) and the laryngeal mask airway (LMA) (from 10 to 26%). No services employ the Combitube. 94% of services use a Hudson type trauma mask (increase of 17%). One service no longer allows its paramedics to deliver 12-15 l of oxygen. 68% of trust paramedics can perform needle thoracocentesis (increase of 35%). No paramedics perform chest drain insertion. All services have 14G intravenous cannulae available and 45% carry the intraosseous needle (increase of 30%). No services employ the MAST suit. There is an increase by 29% in the use of crystalloids and a decrease in the use of colloids of 22%. 23% of fluid resuscitation protocols are based upon the principles of hypotensive resuscitation. Spinal boards and extrication devices are used by 97% of services. The use of inflatable splints has decreased (38-23%). There has been a minor increase in the use of traction splints to 74% of services. The use of Entonox is universal. Nalbuphine (Nubain) is the most widely used opioid. The use of morphine/diamorphine has doubled to 10% with a further 26% to introduce their use. 29% of services have equipped some vehicles with automatic external defibrillators. CONCLUSIONS: The equipment available to UK paramedics and procedures that they may perform continues to expand. There are still variations in the basic management of airway, breathing and circulation care and only some services are keeping up to date with current medical thinking, for example the increasing use of crystalloids and hypotensive resuscitation. It remains to be seen whether the widespread use of Nalbuphine as a first line analgesic may decrease as the use of natural strong opiates becomes more widespread.

Ambulances↗
PubMed2003

AtAGP30, an arabinogalactan-protein in the cell walls of the primary root, plays a role in root regeneration and seed germination.

Arabinogalactan-proteins (AGPs) are extracellular proteoglycans that are implicated in many plant growth and developmental processes, but in no case has a biological function been assigned to a particular AGP. AtAGP30 is a non-classical AGP core protein from Arabidopsis that is expressed only in roots. Analysis of the corresponding mutant, agp30, has revealed that the wild-type gene product is required in vitro for root regeneration and in planta for the timing of seed germination. The mutant shows a suppression of the abscisic acid (ABA)-induced delay in germination and altered expression of some ABA-regulated genes. This suggests that AtAGP30 functions in the ABA response. By analogy to proteoglycan-mediated regulation of growth-factor-signalling pathways in animals, our data indicate that phytohormone activity in plants can be modulated by AGPs.

Amino Acid Sequence↗
PubMed2002

Galactose biosynthesis in Arabidopsis: genetic evidence for substrate channeling from UDP-D-galactose into cell wall polymers.

The biosynthesis of plant cell wall polysaccharides requires the concerted action of nucleotide sugar interconversion enzymes, nucleotide sugar transporters, and glycosyl transferases. How cell wall synthesis in planta is regulated, however, remains unclear. The root epidermal bulger 1 (reb1) mutant in Arabidopsis thaliana is partially deficient in cell wall arabinogalactan-protein (AGP), indicating a role for REB1 in AGP biosynthesis. We show that REB1 is allelic to ROOT HAIR DEFICIENT 1 (RHD1), one of five ubiquitously expressed genes that encode isoforms of UDP-D-glucose 4-epimerase (UGE), an enzyme that acts in the formation of UDP-D-galactose (UDP-D-Gal). The RHD1 isoform is specifically required for the galactosylation of xyloglucan (XG) and type II arabinogalactan (AGII) but is not involved either in D-galactose detoxification or in galactolipid biosynthesis. Epidermal cell walls in the root expansion zone lack arabinosylated (1-->6)-beta-D-galactan and galactosylated XG. In cortical cells of rhd1, galactosylated XG is absent, but an arabinosylated (1-->6)-beta-D-galactan is present. We conclude that the flux of galactose from UDP-D-Gal into different downstream products is compartmentalized at the level of cytosolic UGE isoforms. This suggests that substrate channeling plays a role in the regulation of plant cell wall biosynthesis.

Arabidopsis↗
PubMed2002

DNA topoisomerase VI is essential for endoreduplication in Arabidopsis.

Endoreduplication is a common process in eukaryotes that involves DNA amplification without corresponding cell divisions. Cell size in various organisms has been linked to endoreduplication, but the molecular mechanisms are poorly understood. We have used a genetic strategy to identify molecules involved in endocycles in Arabidopsis. We isolated two extreme dwarf mutants, hypocotyl6 (hyp6) and root hairless2 (rhl2) [3], and cells of these mutants successfully complete only the first two rounds of endoreduplication and stall at 8C. In both mutants, large cell types, such as trichomes and some epidermal cells, that normally endoreduplicate their DNA are much reduced in size. We show that HYP6 encodes AtTOP6B, a plant homolog of the archaeal DNA topoisomerase VI subunit B, and that RHL2 encodes AtSPO11-3, one of the three Arabidopsis subunit A homologs. We propose that this topoisomerase VI complex is essential for the decatenation of replicated chromosomes during endocycles and that successive rounds of endoreduplication are required for the full growth of specific cell types.

Arabidopsis↗
PubMed2002

The mechanics of cell fate determination in petals.

The epidermal cells of petals of many species are specialized, having a pronounced conical shape. A transcription factor, MIXTA, is required for the formation of conical cells in Antirrhinum majus; in shoot epidermal cells of several species, expression of this gene is necessary and sufficient to promote conical cell formation. Ectopic expression has also shown MIXTA to be able to promote the formation of multicellular trichomes, indicating that conical cells and multicellular trichomes share elements of a common developmental pathway. Formation of conical cells or trichomes is also mutually exclusive with stomatal formation. In Antirrhinum, MIXTA normally only promotes conical cell formation on the inner epidermal layer of the petals. Its restricted action in cell fate determination results from its specific expression pattern. Expression of MIXTA, in turn, requires the activity of B-function genes, and biochemical evidence suggests that the products of DEFICIENS, GLOBOSA and SEPALLATA-related genes directly activate MIXTA expression late in petal development, after the completion of cell division in the petal epidermis. A MIXTA-like gene, AmMYBML1, is also expressed in petals. AmMYBML1 expression is high early in petal development. This gene may direct the formation of trichomes in petals. In specifying the fates of different cell types in petals, regulatory genes like MIXTA may have been duplicated. Changes in the timing and spatial localization of expression then provides similar regulatory genes which specify different cell fates.

Antirrhinum↗
PubMed2002

Fucosylated arabinogalactan-proteins are required for full root cell elongation in arabidopsis.

The Arabidopsis thaliana mutant mur1 is affected in the biosynthesis of l-fucose and has less than 2% of the normal amounts of this sugar in the cell walls of its aerial parts. Although in roots the reduction of l-fucose is only 40%, this causes a decrease of about 50% in root cell elongation. Since arabinogalactan-proteins (AGPs) are known to play a role in plant cell expansion we studied the composition of mur1 root AGPs. Arabidopsis root AGPs were shown to contain l-fucose, which was reduced in level in mur1 AGPs. In wild-type plants, an l-fucose containing epitope is present in AGPs in the cell wall of differentiating root cells. Addition of eel lectin, which specifically recognizes this epitope, and not fucose in other wall polymers, can phenocopy mur1 roots. Several lines of evidence are presented to support the contention that l-fucose containing root AGPs are required for the full elongation of root cells.

Animals↗