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R S Quatrano

Publications and source records attributed to R S Quatrano.

At least 37 records · Page 2Linked to original sources
PubMed1996

Polar localization of a dihydropyridine receptor on living Fucus zygotes.

We have used a fluorescently-labeled dihydropyridine (FL-DHP) to vitally stain living Fucus zygotes during the establishment of cell polarity. Localization of FL-DHP is primarily at the plasma membrane and FL-DHP binding is competitively blocked by an unlabeled dihydropyridine. Distribution of FL-DHP is initially symmetrical before fixation of the polar axis, but becomes asymmetrical in response to a unilateral light gradient. The distribution of FL-DHP receptors can be relocalized when the direction of the photopolarizing stimulus is changed. Treatment of cells with cytochalasin B prior to axis fixation reversibly prevents localization of FL-DHP receptors. Observation of FL-DHP labeling by time-lapse fluorescence microscopy indicates that the existing receptors are redistributed during polar axis formation. The asymmetric distribution of FL-DHP receptors coincides temporally and spatially with increased local intracellular calcium ion concentrations, as measured by calcium green dextran. Based on the site, timing, photo-reversibility, and actin dependence of the asymmetric localization of FL-DHP receptors, we conclude that FL-DHP is a vital probe for the later stage of polar axis formation in Fucus zygotes. Furthermore, we propose that FL-DHP receptors correspond to ion channels that are transported to the future site of polar growth to create the changes in local calcium concentration required for polarity establishment.

Calcium↗
PubMed1995

Spatial redistribution of poly(A)+ RNA during polarization of the Fucus zygote is dependent upon microfilaments.

Asymmetrical distribution of mRNA has been associated with polarization and cell fate determination during early development of animal embryos. In this report we determine the distribution pattern of poly(A)+ RNA during early embryogenesis of the brown alga Fucus. Poly(A)+ RNA is symmetrically distributed in the egg and early zygote. Shortly after the polar axis is established, poly(A)+ RNA becomes segregated to the thallus pole of the zygote. Following cytokinesis, most of poly(A)+ RNA is partitioned into the thallus cell. We show that the spatial redistribution of poly(A)+ RNA requires intact microfilaments and the fixation of the polar axis, but is not dependent upon polarized growth of the rhizoid, intact microtubules, or orientation of the division plane.

Cell Division↗
PubMed1995

Structural features and phylogeny of the actin gene of Chondrus crispus (Gigartinales, Rhodophyta).

We have characterized the cDNA and genomic sequences that encode actin from the multicellular red alga Chondrus crispus. Southern-blot analysis indicates that the C. crispus actin gene (ChAc) is present as a single copy. Northern analysis shows that, like the GapA gene, the actin gene is well expressed in gametophytes but weakly in protoplasts. Compared to actin genes of animals, fungi, green plants and oomycetes, that of C. crispus displays a higher evolutionary rate and does not show any of the amino-acid signatures characteristic of the other lineages. As previously described for GapA, ChAc is interrupted by a single intron at the beginning of the coding region. The site of initiation of transcription was characterized by RNAse protection. The promoter region displays a CAAT box but lacks a canonical TATA motif. Other noticeable features, such as a high content of pyrimidines as well as a 14-nt motif found in both the 5'-untranslated region and the intron, were observed.

Actins↗
PubMed1995

ABA-regulated promoter activity in stomatal guard cells.

CDeT6-19 is an ABA-regulated gene which has been isolated from Craterostigma plantagineum. The CDeT6-19 gene promoter has been fused to the beta-glucuronidase reporter gene (GUS) and used to stably transform Arabidopsis thaliana and Nicotiana tabacum. This construct has been shown to be expressed in stomatal guard cells and often in the adjacent epidermal cells of both species in response to both exogenous ABA and drought stress. These results indicate that the stomatal guard cell is competent to relay an ABA signal to the nucleus. In contrast GUS expression directed by the promoter from a predominantly seed-specific, ABA-regulated gene, Em, or the promoter from the ABA-regulated CDeT27-45 gene is not detectable in the epidermal or guard cells of tobacco or Arabidopsis in response to ABA. The fact that not all ABA-regulated gene promoters are active in stomatal guard cells suggests that effective transduction of the signal is dependent upon particular regions within the gene promoter or that guard cells lack all or part of the specific transduction apparatus required to couple the ABA signal to these promoters. This suggests that there are multiple ABA stimulus response coupling pathways. The identification of a regulatory sequence from an ABA-induced gene which is expressed in stomatal guard cells creates the possibility of examining the role of Ca2+ and other second messengers in ABA-induced gene expression.

Abscisic Acid↗
PubMed1995

Overlap of Viviparous1 (VP1) and abscisic acid response elements in the Em promoter: G-box elements are sufficient but not necessary for VP1 transactivation.

The relationship between promoter sequences that mediate Viviparous1 (VP1) transactivation and regulation by abscisic acid (ABA) in the wheat Em promoter was investigated using deletion analysis and directed mutagenesis. The Em1a G-box is strongly coupled to VP1 transactivation as well as to ABA regulation; however, the Em promoter includes additional components that can support VP1 transactivation without ABA responsiveness or synergism. Oligonucleotide tetramers of several G-box sequences, including Em1a, Em1b, and the dyad G-box element from the UV light-regulated parsley chalcone synthase gene, were sufficient to confer VP1 transactivation and the synergistic interaction with ABA to the -45 cauliflower mosaic virus 35S core promoter. These data suggest that VP1 can activate transcription through at least two classes of cis-acting sequences, including the G-box elements and the Sph regulatory motif found in the C1 promoter. The contrasting roles of these motifs in the Em and C1 promoters suggest a basis for the differential regulation of the corresponding genes by VP1.

Abscisic Acid↗
PubMed1995

Digital photography for the light microscope: results with a gated, video-rate CCD camera and NIH-image software.

In this report, we describe a relatively inexpensive method for acquiring, storing and processing light microscope images that combines the advantages of video technology with the powerful medium now termed digital photography. Digital photography refers to the recording of images as digital files that are stored, manipulated and displayed using a computer. This report details the use of a gated video-rate charge-coupled device (CCD) camera and a frame grabber board for capturing 256 gray-level digital images from the light microscope. This camera gives high-resolution bright-field, phase contrast and differential interference contrast (DIC) images but, also, with gated on-chip integration, has the capability to record low-light level fluorescent images. The basic components of the digital photography system are described, and examples are presented of fluorescence and bright-field micrographs. Digital processing of images to remove noise, to enhance contrast and to prepare figures for printing is discussed.

Microscopy, Video↗
PubMed1994

Plant regulators. Insensitivity is in the genes.

The cloning of loci determining abscisic acid insensitivity in Arabidopsis has identified a phosphatase and a transcriptional activator that mediate responses to abscisic acid and so regulate plant growth and development.

Abscisic Acid↗
PubMed1993

The cis-regulatory element CCACGTGG is involved in ABA and water-stress responses of the maize gene rab28.

The maize gene rab28 has been identified as ABA-inducible in embryos and vegetative tissues. It is also induced by water stress in young leaves. The proximal promoter region contains the conserved cis-acting element CCACGTGG (ABRE) reported for ABA induction in other plant genes. Transient expression assays in rice protoplasts indicate that a 134 bp fragment (-194 to -60 containing the ABRE) fused to a truncated cauliflower mosaic virus promoter (35S) is sufficient to confer ABA-responsiveness upon the GUS reporter gene. Gel retardation experiments indicate that nuclear proteins from tissues in which the rab28 gene is expressed can interact specifically with this 134 bp DNA fragment. Nuclear protein extracts from embryo and water-stressed leaves generate specific complexes of different electrophoretic mobility which are stable in the presence of detergent and high salt. However, by DMS footprinting the same guanine-specific contacts with the ABRE in both the embryo and leaf binding activities were detected. These results indicate that the rab28 promoter sequence CCACGTGG is a functional ABA-responsive element, and suggest that distinct regulatory factors with apparent similar affinity for the ABRE sequence may be involved in the hormone action during embryo development and in vegetative tissues subjected to osmotic stress.

Abscisic Acid↗
PubMed1992

Role of a vitronectin-like molecule in embryo adhesion of the brown alga Fucus.

The rhizoid cell of the two-celled embryo of the brown alga Fucus is structurally and functionally differentiated from the thallus cell. The rhizoid cell is highly polar and transports directionally components of the cell wall to its elongating tip, which attaches the developing embryo to the substratum. Polyclonal antibodies to human vitronectin (Vn) recognize a vitronectin-like glycoprotein (Vn-F) in extracts of zygotes and two-celled embryos of Fucus, with a molecular mass (approximately 62 kDa) similar to that of human Vn. The specificity of the immunological cross-reactivity of Vn-F to rabbit polyclonal antibodies made to human Vn is demonstrated by competition experiments using pure human Vn and monospecific antibodies generated toward Vn-F. Vn-F possesses affinities for glass and heparin that are identical to those of human Vn. Immunolocalization and subcellular fractionation results demonstrate that Vn-F is localized first in the cytoplasm of the zygote, which is followed by the polar transport of Vn-F to its exclusive localization in the cell wall of the elongating rhizoid tip. Vn does not localize to the rhizoid tip under culture conditions that prevent two-celled embryos from attaching. Furthermore, an adhesion assay demonstrates that two-celled Fucus embryos do not adhere to the substratum in the presence of the Vn antibody, suggesting that the Vn-F in this brown alga not only possesses structural similarity to mammalian Vn but may also have a similar functional role in adhesion. The presence of Vn-F in brown algae suggests a high degree of evolutionary conservation of its structural and functional characteristics.

Cell Adhesion↗
PubMed1992

Abscisic Acid Elicits the Water-Stress Response in Root Hairs of Arabidopsis thaliana.

Water stress has been shown to cause root hairs to become short and bulbous. Because abscisic acid (ABA) mediates a variety of water-stress responses, we investigated the response of Arabidopsis thaliana root hairs to ABA. When wild-type root hairs were treated with ABA, they exhibited the water-stress response. The Arabidopsis mutants abi1 and abi2, which are insensitive to ABA at the seedling stage, did not display the root hair response. These data suggest that ABA may mediate the response of root hairs to water stress. The drought response of root hairs resulting in an inhibition of tip growth will provide an easy screen to select mutations that are insensitive to ABA and/or involved in tip growth.

Journal Article↗
PubMed1992

Regulation of Em Gene Expression in Rice : Interaction between Osmotic Stress and Abscisic Acid.

Expression of the Em gene was characterized in rice (Oryza sativa L.) suspension cultures following exposure of the cultures to various combinations of abscisic acid (ABA) and salt. Response-saturating concentrations of either ABA (50 micromolar) or NaCl (0.4 molar) rapidly induced (by 60 minutes) the accumulation of Em mRNA, with a maximum accumulation occurring 12 to 24 hours after treatment. NaCl-induced Em expression was accompanied by a doubling of endogenous ABA levels as determined by immunoassay. Inhibition of ABA biosynthesis by fluridone during NaCl treatment reduced the levels of endogenous ABA by fourfold and Em expression by 50%. Desiccation of the cultures to 12 to 15% of their initial fresh weight increased endogenous ABA more than twofold and was accompanied by an increase in Em mRNA levels. Exposure of the cultures to heat shock temperatures, chilling, or ultraviolet light neither increased endogenous ABA levels nor induced Em expression. When a subthreshold or saturating level of NaCl was added in combination with increasing levels of ABA, Em transcripts were detected at ABA concentrations that alone did not induce expression of Em. Treatment with saturating levels of both NaCl and ABA resulted in a doubling of Em transcript levels over the maximum signal for each treatment alone. Hence, our data suggested that salt interacted synergistically with ABA, in part because of the increased sensitivity of rice cells to ABA. The effect of salt stress on Em gene expression in rice suspension cells appeared to operate through two pathways: one is mediated through increases in the level of ABA; the other is via a unique salt response pathway that includes an intermediate that is common to both the salt and ABA response chains.

Journal Article↗
PubMed1991

Sequence analysis of a functional member of the Em gene family from wheat.

We report the complete sequence of one functional member of the Em gene family whose expression in wheat embryos is regulated by a complex set of environmental and developmental controls, including the phytohormone abscisic acid (ABA). The Em coding region contains one short intron, and there is an inverted repeat in the transcribed 3'-flanking region. A 646 bp fragment from the 5' promoter, which was previously shown to direct ABA-regulated expression in transformed tobacco tissue and rice cells, is characterized by: (1) three stretches of between 33 and 73 nucleotides of A/T rich (greater than 86%) boxes, (2) one copy of an eight bp palindrome (CATGCATG) which is identical to the RY repeat found in the 5' promoters of many legume genes expressed during embryo development, (3) 15 copies of a six bp repeat (PuCACGPy), found primarily in the 5' region, and (4) two sequences in the ABA-response region, CGAGCAG and a CACGT motif, both of which are conserved in 5' non-coding regions of other plant genes that are expressed in response to ABA and/or in embryos. These sequence comparisons are discussed in relation to the regulation of Em gene expression and other ABA-regulated genes.

Amino Acid Sequence↗
PubMed1991

Polar axis fixation in Fucus zygotes: components of the cytoskeleton and extracellular matrix.

Polar axis formation and polar axis stabilization (or fixation) can be separated and analyzed in synchronously developing zygotes of the brown alga Fucus. Extensive experimental evidence points to a role for both the cytoskeleton and the extracellular matrix (ECM) in the process of axis fixation in Fucus. A structural complex composed of the cytoskeleton and the ECM has been postulated to stabilize membrane asymmetries generated as a result of axis-forming vectors. This axis stabilizing complex (ASC) may take the form of transmembrane connections between the cytoskeleton on the cytoplasmic face and the ECM on the external side of the plasma membrane, similar to focal contacts in animal cells. At present we know of two components in the proposed ASC of Fucus: an adhesive sulfated glycoprotein which is localized in the ECM, and an actin network which is localized on the adjoining cytoplasmic face. This preliminary report describes evidence for the presence of molecules in two-celled Fucus embryos that are similar to those found in focal contacts in animal cells, i.e. vinculin, integrin and vitronectin. However, their localization and interaction with each other relative to the polar axis has yet to be determined. These initial observations will provide the basis to pursue further an analysis of these components in the process of polar axis fixation.

Cell Polarity↗
PubMed1990

A plant leucine zipper protein that recognizes an abscisic acid response element.

The mechanism by which phytohormones, like abscisic acid (ABA), regulate gene expression is unknown. An activity in nuclear extracts that interacts with the ABA response element (ABRE) from the 5' regulatory region of the wheat Em gene was identified. A complementary DNA clone was isolated whose product is a DNA binding protein (EmBP-1) that interacts specifically with an 8-base pair (bp) sequence (CACGTGGC) in the ABRE. A 2-bp mutation in this sequence prevented binding of EmBP-1. The same mutation reduced the ability of the ABRE to confer ABA responsiveness on a viral promoter in a transient assay. The 8-bp EmBP-1 target sequence was found to be conserved in several other ABA-responsive promoters and in promoters from plants that respond to signals other than ABA. Similar sequences are found in promoters from mammals, yeast, and in the major late promoter of adenovirus. The deduced amino acid sequence of EmBP-1 contains conserved basic and leucine zipper domains found in transcription factors in plants, yeast, and mammals. EmBP-1 may be a member of a highly conserved family of proteins that recognize a core sequence found in the regulatory regions of various genes that are integrated into a number of different response pathways.

Abscisic Acid↗
PubMed1989

Protein synthesis and morphogenesis are not tightly linked during embryogenesis in Fucus.

Fertilized eggs of the brown alga Fucus have long been used as model organisms for investigating the early events in the establishment of polarity and subsequent embryogenesis since large numbers of zygotes can easily be obtained. We have analyzed protein synthesis in eggs and embryos during the first day of development using two-dimensional gels and found that synthesis of 12 of the 60 most prominent proteins changed either qualitatively or quantitatively. Actin and beta-tubulin were identified by immunoblotting; synthesis of these cytoskeletal proteins was initiated at different times during the first 12 hr of development. Unique, reproducible patterns of protein synthesis observed during development in the light permitted accurate staging of developing embryos. Inhibitors such as cytochalasin and sucrose, however, blocked morphogenesis without affecting protein synthesis, and, conversely, growth in the dark delayed protein synthesis but had very little effect on the timing of morphogenesis. The data are consistent with morphogenesis and protein synthesis being relatively independent during early embryogenesis. Actinomycin D added soon after fertilization had no effect on protein synthesis 1 day later, indicating that the proteins analyzed were encoded by maternal mRNA stored in the egg.

Cell Division↗