Biomedical subjects

C Dumas

Publications and source records attributed to C Dumas.

At least 37 records · Page 2Linked to original sources

PubMed2001

Dynamic analyses of the expression of the HISTONE::YFP fusion protein in arabidopsis show that syncytial endosperm is divided in mitotic domains.

During early seed development, nuclear divisions in the endosperm are not followed by cell division, leading to the development of a syncytium. The simple organization of the Arabidopsis endosperm provides a model in which to study the regulation of the cell cycle in relation to development. To monitor nuclear divisions, we constructed a HISTONE 2B::YELLOW FLUORESCENT PROTEIN gene fusion (H2B::YFP). To validate its use as a vital marker for chromatin in plants, H2B::YFP was expressed constitutively in Arabidopsis. This enabled the observation of mitoses in living root meristems. H2B::YFP was expressed specifically in Arabidopsis syncytial endosperm by using GAL4 transactivation. Monitoring mitotic activity in living syncytial endosperm showed that the syncytium was organized into three domains in which nuclei divide simultaneously with a specific time course. Each mitotic domain has a distinct spatiotemporal pattern of mitotic CYCLIN B1;1 accumulation. The polar spatial organization of the three mitotic domains suggests interactions between developmental mechanisms and the regulation of the cell cycle.

Arabidopsis↗

PubMed2001

How to build patient loyalty.

When it comes to judging the quality of dental care they receive, patients often rely on more than the dental care itself. Patients judge quality by what they can see, touch, feel, and sense. A quality practice dedicated to nurturing loyal patients must create an environment that, in the eyes and perception of its patients, stands apart it from the competition. Many of today's service corporations can teach us how to market our practices and services and, in turn, keep our patients happy with their dental care and all aspects of the dental care experience.

Dentist-Patient Relations↗

PubMed2000

A calcium influx is triggered and propagates in the zygote as a wavefront during in vitro fertilization of flowering plants.

In this paper, we report direct measurement of an influx of extracellular Ca(2+) induced by gamete fusion in flowering plants. This result was obtained during maize in vitro fertilization with the use of an extracellular Ca(2+)-selective vibrating probe. Ca(2+) influx recorded at the surface of isolated egg cells, with or without adhesion of a male sperm cell, was close to zero and stable over time. Gamete fusion, however, triggered a Ca(2+) influx in the vicinity of the sperm entry site with a delay of 1.8 +/- 0.6 sec. The Ca(2+) influx spread subsequently through the whole egg cell plasma membrane as a wavefront, progressing at an estimated rate of 1.13 micrometer.(-1). Once established, Ca(2+) influx intensities were sustained, monotonic and homogeneous over the whole egg cell, with an average peak influx of 14.92 pmol .cm(-2).(-1) and an average duration of 24.4 min. The wavefront spread of channel activation correlates well with the cytological modifications induced by fertilization, such as egg cell contraction, and with the cytosolic Ca(2+) ((c)[Ca(2+)]) elevation previously reported. Calcium influx was inhibited effectively by gadolinium, possibly implicating mechanosensitive channels. Furthermore, artificial influxes created by incubation with Ca(2+) ionophores mimicked some aspects of egg activation. Taken together, these results suggest that, during fertilization in higher plants, gamete membrane fusion starts the first embryonic events by channel opening and Ca(2+) influx. In turn, (c)[Ca(2+)] may work as a trigger and possibly a space and time coordinator of many aspects of egg activation.

Calcium↗

PubMed2000

Multiple S gene family members including natural antisense transcripts are differentially expressed during development of maize flowers.

Within the large Brassica S gene family, SLG (S locus glycoprotein) and SRK (S locus receptor kinase) participate to the control of pollen-stigma self-incompatibility. In the self-compatible species maize, S gene family members are predominantly expressed in vegetative organs but are also expressed to a lesser extent in the stigma (silk). To determine if the expression of any S gene family members correlates with female receptivity, we analyzed their expression in developing maize silks. We show that a large family of maize S transcripts is expressed in developing silks. Surprisingly, we isolated a cDNA complementary to a large portion of the antisense strand of the maize receptor kinase S domain. Rapid amplification of cDNA ends (RACE)-polymerase chain reaction, RNase protection, and Northern hybridization with single-stranded riboprobes confirmed that natural antisense S transcripts exist in leaves and seedling shoots and in all sexual tissues tested except mature pollen. These natural antisense S transcripts co-exist with several less abundant sense S transcripts. The accumulation of sense and antisense S transcripts is differentially regulated during pollen and silk development. Thus, these results support a role for S gene family members in sexual tissue development and/or compatible pollination and reveal a new level of complexity in the regulation and function of the S gene family in maize.

Amino Acid Sequence↗

PubMed2000

Esr genes show different levels of expression in the same region of maize endosperm.

Esr genes share high homology among each other, code for small hydrophilic proteins, and are expressed in a restricted region of maize endosperm surrounding the embryo. We show here that not only Esr2 but also Esr1 and Esr3 are expressed in maize, and that the relative contribution of Esr1, Esr2 and Esr3 to total Esr mRNA is 17%, 55% and 28%, respectively. DNA sequence analysis of putative promoter fragments ranging from 0.53 kb to 3.54 kb revealed the presence of retrotransposons related to the Zeon and Cinful families in the distal parts of the promoters. The proximal parts show high homology that extended over 504bp between Esr2 and Esr3, and 265bp between Esr1 and the other two genes. The most conspicuous potential cis element is a fully conserved tandem repeat of the sequence CTACACCA close to the respective open reading frames (ORFs). By the analysis of transgenic maize plants carrying promoter-Gus fusions, it was shown that all three cloned upstream fragments contain functional promoters, that the spatial activity of all three Esr promoters is identical, and that the cis element(s) responsible for the expression in the embryo surrounding region reside in the 265 bp upstream of the respective ORFs.

Base Sequence↗

PubMed2000

The integral membrane S-locus receptor kinase of Brassica has serine/threonine kinase activity in a membranous environment and spontaneously forms oligomers in planta.

To gain further insight into the mode of action of S-locus receptor kinase (SRK), a receptor-like kinase involved in the self-incompatibility response in Brassica, different recombinant SRK proteins have been expressed in a membranous environment using the insect cell/baculovirus system. Recombinant SRK proteins exhibited properties close to those of the endogenous stigmatic SRK protein and were found to autophosphorylate on serine and threonine residues in insect cell microsomes. Autophosphorylation was constitutive because it did not require the presence of pollen or stigma extracts in the phosphorylation buffer. Phosphorylation was shown to occur in trans, suggesting the existence of constitutive homooligomers of membrane-anchored recombinant SRK. To investigate the physiological relevance of these results, we have examined the oligomeric status of SRK in planta in cross-linking experiments and by velocity sedimentation on sucrose gradients. Our data strongly suggest that SRK is associated both with other SRK molecules and other stigma proteins in nonpollinated flowers. These findings may have important implications for our understanding of self-pollen signaling.

Biopolymers↗

PubMed2000

HIV-2 and SIV nef proteins target different Src family SH3 domains than does HIV-1 Nef because of a triple amino acid substitution.

The nef gene is required for optimal viral spread of human and simian immunodeficiency viruses. However, the molecular mechanisms underlying the action of the Nef proteins may not be identical for all viral families. Here we investigate the interaction between the Nef protein of human and simian immunodeficiency viruses and SH3 domains from Src family kinases. Using the yeast two-hybrid system and immunoblotting we show that, in contrast to HIV-1 Nef, SIV and HIV-2 Nef poorly interact with Hck SH3 but bind to Src and Fyn SH3 domains. The molecular basis of these differences in SH3 targeting was revealed by sequence analysis and homology modeling of the putative SH3-Nef structures. Three amino acids (Trp-113, Thr-117, and Gln-118) that localize in a "hydrophobic pocket" implicated in SH3 binding of HIV-1 Nef, are systematically substituted in SIV/HIV-2 alleles (by Tyr, Glu, and Glu, respectively). We demonstrate that site-directed mutagenesis of these residues in SIV(mac239) Nef suffices to restore Hck SH3 binding and co-immunoprecipitation with full-length Hck from transfected cells. Our findings identify fundamental mechanistic differences in targeting of Src family kinases by HIV and SIV Nef. The herein described mechanism of SH3 selection by Nef via a "pocket" proximal to the canonical proline-rich motif may be a common feature for SH3 recognition by their natural ligands.

Amino Acid Sequence↗

PubMed2000

Late steps of egg cell differentiation are accelerated by pollination in Zea mays L.

Egg cells were analysed cytologically during the female receptivity period in maize (Zea mays L., line A 188). Three classes of egg cell were distinguished: type A--small, non-vacuolated cells with a central nucleus; type B--larger cells with small vacuoles surrounding the perinuclear cytoplasm located in the middle of the cell; type C--big cells with a large apical vacuole and the mid-basal perinuclear cytoplasm. The less-dense cytoplasm of the vacuolated egg cells usually contained numerous cup- or bell-shaped mitochondria. The three egg types appear to correspond to three late stages of egg cell differentiation. The frequencies of each of the three egg types were monitored in developing maize ears before and after pollination. In young ears, with the silks just extending out of the husks, small A-type cells were found in about 86% of ovules. Their frequency decreased to about 58% at the optimum silk length, remained unchanged in non-pollinated ears, and fell to 16% at the end of the female receptivity period. However, after pollination and before fertilisation the frequency of these cells decreased to about 33%, and the larger vacuolated egg cells (types B and C) prevailed. At various stages of the receptivity period, pollination accelerated changes in the egg population, increasing the number of ovules bearing larger, vacuolated egg cells. Experiments with silk removal demonstrated that putative pollination signals act immediately after pollen deposition and are not species-specific.

Cell Differentiation↗

PubMed2000

Episomal and stable expression of the luciferase reporter gene for quantifying Leishmania spp. infections in macrophages and in animal models.

We have expressed the reporter firefly luciferase gene (LUC) in Leishmania donovani and Leishmania major either as part of episomal vectors or integrated into the parasite genome under the control of their respective ribosomal promoter regions. An excellent linear correlation between parasite number and luciferase activity was observed with all the transfectants. LUC-expressing recombinant parasites were useful to monitor Leishmania spp. infections in macrophages or in animal models. For prolonged growth in absence of drug selection, such as within animal models, quantitation of parasites is more reliable when the reporter gene LUC is stably integrated in the parasite genome. These recombinant strains should be useful tools to monitor Leishmania growth under a number of conditions.

Animals↗

PubMed2000

Chromosome structure and sequence organization between pathogenic and non-pathogenic Leishmania spp.

We have used a chromosome fragmentation strategy based on systematic genomic insertions of the rare cutting yeast I-SceI endonuclease to assess structure and sequence organization of homologous chromosomes between evolutionary divergent pathogenic and non-pathogenic Leishmania species. This method was combined to physical mapping and hybridization studies using a number of specific chromosomal markers as probes. Our studies have concentrated on two different chromosomes of Leishmania major (L. major), L. donovani and L. infantum and of the non-pathogenic species L. tarentolae. Specific chromosome fragmentation events at the level of multiple I-SccI genomic integrations indicated that very similar distances separated internal genomic sequences between homologous chromosomes and that distances from chromosome ends were more variable. The order and orientation of genes along the homologous chromosomes were also conserved between species. With only few exceptions, genome organization between pathogenic and non-pathogenic Leishmania species was found to be highly conserved. Genomic comparison of pathogenic and non-pathogenic species may be useful for depicting regions involved in species-specific related pathologies.

Animals↗

PubMed2000

Mental and behavioural outcome of infantile epilepsy treated by vigabatrin in tuberous sclerosis patients.

Vigabatrin (VGB) has demonstrated high efficacy in infantile spasms (IS) due to tuberous sclerosis. Our first objective was to evaluate the cognitive long term effect outcome of children whose refractory spasms definitely disappeared when VGB was given as an add on drug. Our second objective was to determine the response of generalized epilepsy (infantile spasms) compared to partial epilepsy on cognitive impairment. A non selected series of 13 children underwent psychometric and behavioural evaluation before VGB initiation at a mean of 3 years on VGB treatment. Eight of them could perform detailed neuropsychological tests at follow-up. Seven had infantile spasms (Group I), they all were spasm free before 2 years of age and five remained with rare partial seizures (mean age, 5.5 years). Six others had partial epilepsy without spasms (Group II) and five remained with rare seizures (mean age, 7.5 years). Patients of Group I experienced dramatic changes. Developmental quotient (DQ) significantly rose in six out of seven by ten to more than 45 points (P = 0.03) and autistic behaviour disappeared in five out of the six who presented with. The four tested children had normal verbal level after 5 years and could integrate at school but they remained with marked visuospatial disabilities. By contrast, patients of Group II remained with an unchanged DQ of about 60 so that both groups had similar DQ levels on follow-up. The cessation of spasms with VGB is therefore associated with significant improvement of cognition and behaviour in children with tuberous sclerosis. Controlling secondary generalization induced by infantile spasms seems to be a key factor for mental development.

Anticonvulsants↗

PubMed2000

[Migraine, misunderstood pathology in children].

Although migraine is the main chronic headache in childhood and adolescence, it remains extensively misdiagnosed. Schematically, migraine is a severe headache evolving by stereotyped attacks frequently associated with marked digestive symptoms (nausea, vomiting, abdominal pain). Throbbing pain, sensitivity to sound, and light (and sometimes odors) are frequent additional symptoms. The attack is sometimes preceded by a visual or sensory aura. Rest brings relief, and sleep often ends the attack. Childhood migraine prevalence varies between 5 and 10%. Migraine episodes are frequently triggered by several factors: emotional stress (school pressure, vexation, excitement, upset), hypoglycemia, lack or excess of sleep (weekend migraine), sensory stimulation (loud noise, bright light, strong odor, heat or cold, etc.), sympathetic stimulation (sport, physical exercise). Attack treatments must be given at an early stage, oral ibuprofen (10 mg/kg) being particularly recommended. If the oral route is not available because of nausea or vomiting, rectal or nasal routes have then to be used. Non-pharmacological treatments (biofeedback and interventions combining progressive muscle relaxation) have demonstrated good efficacy as prophylactic measures. Daily prophylactic pharmacological treatments are prescribed as the second line after failure of non-pharmacological treatments.

Abdominal Pain↗

PubMed2000

Three-dimensional structure of nucleoside diphosphate kinase.

Three-dimensional structures are known from X-ray studies of the nucleoside diphosphate (NDP) kinase of many organisms from bacteria to human. All NDP kinases have subunits of about 150 residues with a very similar fold based on the alphabeta sandwich or ferredoxin fold. This fold is found in many nucleotide or polynucleotide-binding proteins with no sequence relationship to NDP kinase. This common fold is augmented here with specific features: a surface alpha-helix hairpin, the Kpn loop, and the C-terminal extension. The alpha-helix hairpin and Kpn loop make up the nucleotide binding site, which is unique to NDP kinase and different from that of other kinases or ATPases. The Kpn loop and the C-terminal extension are also involved in the quaternary structure. Whereas all known eukaryotic NDP kinases, including mitochondral enzymes, are hexamers, some bacterial enzymes are tetramers. However, hexameric and tetrameric NDP kinases are built from the same dimer. The structural environment of the active histidine is identical in all. The nucleotide binding site is also fully conserved, except for a feature implicating C-terminal residues in the hexamer, but not in the tetramer. Structural data on the native and phosphorylated enzyme, complexes with substrates, inhibitor, and a transition state analog, give a solid basis to a mechanism of phosphate transfer in which the largest contributors to catalysis are the 3'-OH of the sugar and the bound Mg2+ in the nucleotide substrate. In contrast, we still lack structural data relating to DNA binding and other functions of NDP kinases.

Animals↗

PubMed2000

Genes normally expressed in the endosperm are expressed at early stages of microspore embryogenesis in maize.

Reproduction in flowering plants is characterized by double fertilization and the resulting formation of both the zygotic embryo and the associated endosperm. In many species it is possible to experimentally deviate pollen development towards an embryogenic pathway. This developmental switch, referred to as microspore embryogenesis or androgenesis, leads to the formation of embryos similar to zygotic embryos. In a screen for genes specifically expressed during early androgenesis, two maize genes were isolated by mRNA differential display. Both genes represent new molecular markers expressed at a very young stage of androgenic embryogenesis. When their expression pattern was studied during normal reproductive development, both showed early endosperm-specific expression. Investigation of the cytological features of young androgenic embryos revealed that they present a partially coenocytic organization similar to that of early endosperm. These findings suggest that maize androgenesis may possibly involve both embryogenesis and the establishment of endosperm-like components.

Blotting, Southern↗

PubMed2000

Flexible search behavior in domestic cats (Felis catus): a case study of predator-prey interaction.

Domestic cats (Felis catus) were administered an object permanence task in a novel and a familiar situation to investigate flexibility (i.e., pause behavior and searching by following a path opposite of that taken by the object when it disappeared) in search behavior. Pause and opposite search were assumed to be independent, equiprobable, and randomly exhibited (i.e., random model). The random model predicted that cats would exhibit flexible behavior on 75% of the trials. The results revealed that flexible behavior occurred on 69% of the trials in the novel situation, but only on 52% of the trials in the familiar setting in which pauses were less frequent and shorter than in the novel situation. Thus, the random model provided a good fit of the data in the novel but not in the familiar situation. It is argued that pause and opposite search reflect decision processes when cats are dealing with the behavior of prey that has disappeared while being pursued.

Analysis of Variance↗

PubMed2000

Expression patterns of genes encoding HD-ZipIV homeo domain proteins define specific domains in maize embryos and meristems.

A family of homeo box genes with cell layer-specific expression patterns defining subdomains of the embryo and certain meristems has been isolated from maize. These genes encode proteins from the class of plant specific homeo domain-leucine zipper (HD-Zip) transcription factors containing the previously described ZmOCL1 protein, and have been designated ZmOCL2, ZmOCL3, ZmOCL4 and ZmOCL5. ZmOCL3, ZmOCL4 and ZmOCL5, like ZmOCL1, showed essentially L1 or epidermis-specific expression. However, each gene was expressed in a distinct region of the embryonic protoderm during early development, with ZmOCL3 showing suspensor-specific expression, ZmOCL4 transcripts being localized to the adaxial face of the embryo proper and ZmOCL5 showing a more abaxial expression pattern. All three genes were also expressed in vegetative, inflorescence and floral apices, although ZmOCL3 transcripts were excluded from meristems and very young organ primordia. In contrast, ZmOCL2 expression was entirely meristem-specific and was excluded from the L1 layer, appearing instead to be largely restricted to a cell layer directly beneath the L1, especially in floral meristems. This expression pattern is unprecedented and may indicate that cell-layer organization in maize meristems is more complex than that suggested by the classical L1/L2 (outer cell layer/inner cell mass) model. These differing expression patterns indicate that the members of the HD-ZipIV family of maize may not only play roles in defining different regions of the epidermis during embryonic development, but could also be responsible for maintaining cell-layer identity in meristematic regions.

Amino Acid Sequence↗

PubMed2000

Aquaporin PIP genes are not expressed in the stigma papillae in Brassica oleracea.

The pollen grains of angiosperms are usually desiccated at maturity. Following pollination, pollen hydrates on the stigma surface before germination takes place. Rehydration is an essential step for the success of pollination and depends on the movement of water from the stigmatic cells. This water flow has been shown to be biologically regulated, and components of both pollen and stigma surfaces have been demonstrated to play a role in the control of pollen hydration. Regulation of water transport between animal or plant cells involves membrane proteins, designated aquaporins, which possess water-channel activity. Such molecules may be candidates for controlling pollen hydration, and consequently we investigated whether aquaporins are present in the pollen and stigma cells in Brassica oleracea. Here, we report the identification of two new aquaporin genes, Bo-PIP1b1 and Bo-PIP1b2, which are highly homologous to PIP1b from Arabidopsis thaliana. Both Bo-PIP1b1 and Bo-PIP1b2 proteins are active water channels when expressed in Xenopus oocytes. Expression of Bo-PIP1b1 and Bo-PIP1b2 was observed in reproductive organs as well as in vegetative tissues. Interestingly, the use of a Bo-PIP1b2 cDNA probe revealed that PIP1-like transcripts were not present in the pollen grains or in the stigma papillae, but were present in the stigma cell layers underlying the papillar cells. This observation suggests that water flow between the pollen and stigma papillae may be dependent on aquaporins expressed in cells that are not directly in contact with the pollen grain.

Amino Acid Sequence↗

PubMed2000

Characterization and molecular basis of the oligomeric structure of HIV-1 nef protein.

The Nef protein of human immunodeficiency virus type I (HIV-1) is an important determinant for the onset of AIDS disease. The self-association properties of HIV-1 Nef are analyzed by chemical cross-linking, dynamic light scattering, equilibrium analytical ultracentrifugation, and NMR spectroscopy. The experimental data show that the HIV-1 Nef core domain forms stable homo-dimers and trimers in solution, but not higher oligomers. These Nef homomers are not covalently linked by disulfide bridges, and the equilibrium between these forms is dependent on the Nef concentration. We further provide the molecular basis for the Nef core dimers and trimers obtained by analysis of crystallographic models. Oligomerization of biological polypeptides is a common tool used to trigger events in cellular signaling and endocytosis, both of which are targeted by Nef. The quaternary structure of Nef may be of physiological importance and may help to connect its cellular targets or to increase affinity of the viral molecule for its ligands. The herein described models for Nef dimers and trimers will allow further mutational studies to elucidate their role in vivo. These results provide novel insight into the structural and functional relationships of this important viral protein. Moreover, the oligomer interface may represent a novel target for the design of antiviral agents.

Biopolymers↗