Search PubMed⌕ Search

Biomedical subjects

E Lam

Publications and source records attributed to E Lam.

At least 19 recordsLinked to original sources
PubMed1999

Nonspecific inhibition of myogenic tone by PD98059, a MEK1 inhibitor, in rat middle cerebral arteries.

Activation of MAP kinase kinase, also called ERK kinase (MEK), may lead to desinhibition of thin filament regulatory proteins and we therefore investigated the acute effects of the potent MEK inhibitor, PD98059 on the contractile properties of pressurized rat middle cerebral arteries. Cerebral arteries (diameter 100-150 microm) were mounted on a pressure myograph and PD98059 (10 microM, 40 microM) significantly inhibited (15% and 64%) myogenic tone (P < 0.001). At these concentrations, PD98059 also significantly reduced the vasopressin (0.1 microM)- and KCl (60 mM)-induced tone. Cumulative addition of exogenous Ca2+ (0.4-1.6 mM) increased myogenic tone to approximately 50% of constriction at 80 mmHg. This effect was inhibited by PD98059 (P < 0.001). These results demonstrate that pressure-induced myogenic tone is inhibited by PD98059 at the concentrations that have been reported to be selective for inhibition of MEK and the MAP kinase cascade. However, our results also demonstrate that PD98059 may have nonspecific effects on voltage-sensitive Ca2+ entry in vascular smooth muscle.

Animals↗
PubMed1998

Caspases and programmed cell death in the hypersensitive response of plants to pathogens.

The hypersensitive response (HR) is induced by certain plant pathogens and involves programmed cell death (PCD) to restrict the spread of pathogens from the infection site [1]. Concurrent with the induction of cell death, the host activates a defense response [2]. The cell death associated with the HR in several plant-pathogen systems has morphological similarities to animal apoptosis [3,4], which suggests that cell death mechanisms in plants and animals may share common components that lead to similar cellular events. Caspases are conserved cysteine proteases that regulate animal PCD [5]; caspase activity or an involvement of caspases in cell death has yet to be reported in plants. In this work, we investigated the participation of caspases in HR cell death. Caspase-specific peptide inhibitors, Ac-YVAD-CMK [6] and Ac-DEVD-CHO [7], could abolish bacteria-induced plant PCD but did not significantly affect the induction of other aspects of HR, such as the expression of defense genes. This result confirmed our previous model that cell death can be uncoupled from defense gene activation during HR [8]. Caspase-like proteolytic activity was detected in tobacco tissues that were developing HR following infection with tobacco mosaic virus (TMV). Our results provide evidence for the presence of caspase-like plant protease(s) that participate in HR cell death.

Amino Acid Chloromethyl Ketones↗
PubMed1998

Inhibition of myogenic tone by mibefradil in rat cerebral arteries.

The inhibitory effects of mibefradil (1.0 nM-1.0 microM), a putative selective inhibitor of T-type Ca2+ channels that has peripheral and coronary vasodilating properties with few negative inotropic effects, on pressure-induced vasoconstrictions were compared to depolarization- and [Arg8] vasopressin-induced tone in isolated middle cerebral arteries of the rat. The concentration-response relationships (IC50) for myogenic tone (70 +/- 20 nM), depolarization- (53 +/- 9 nM) and vasopressin-induced tone (70 +/- 10 nM) were equally inhibited by mibefradil. Pressure-induced responses were consistently inhibited by mibefradil throughout the myogenically active pressure range (20-100 mmHg). These results demonstrate that mibefradil is nonselective in inhibiting Ca2+ dependent cerebral artery tone due to myogenic activation, depolarization or receptor activation.

Animals↗
PubMed1998

Activation of hsr203, a plant gene expressed during incompatible plant-pathogen interactions, is correlated with programmed cell death.

hsr203J is a tobacco gene whose activation is rapid, highly localized, and specific for incompatible interactions between tobacco and the bacterial pathogen Ralstonia solanacearum. The effect of other hypersensitive response (HR)-inducing pathogens and elicitors has been tested with transgenic plants containing the hsr203J promoter-GUS reporter gene fusion, and confirms the generality of the preferential inducibility of the hsr203J gene promoter during incompatible interactions: bacterial and viral pathogens inducing an HR in tobacco were able to induce the promoter fusion, as were inducers of HR-like responses such as harpin, elicitins, and PopA1 proteins. A tomato hsr203 homologous cDNA was isolated (Lehsr203) and used to examine the effect of avr gene products on the expression of such genes. Lehsr203 was shown to be rapidly and transiently induced in leaves of the tomato Cf-9 line, following Avr9 product infiltration, but not in those of the Cf-0 line. Among potential effectors of HR or resistance such as H2O2, salicylic acid, methyl jasmonate, and 2,6-dichloro-isonicotinic acid (INA), none is able to induce a significant increase in promoter activation. In contrast, heavy metals that cause leaf necrosis can trigger such an activation. In addition, hsr203-GUS fusion expression is detected in transgenic tobacco lines expressing the bO gene and exhibiting spontaneous HR-like lesions. Taken together, these results demonstrate a strong correlation between hsr203 and genetically controlled cell death in tobacco and tomato. The expression of this gene should be a useful marker for programmed cell death occurring in response not only to diverse pathogens, but also to diverse death-triggering extracellular agents.

Amino Acid Sequence↗
PubMed1997

Pathogen-induced programmed cell death in plants, a possible defense mechanism.

As much as the definition of life may be controversial, the definition of death also may prove problematic. In recent years it became apparent that the death of a living cell may follow more than one possible scenario: it may result from an externally applied physical injury (an accidental death), or it may be the outcome of activating an internal pathway for cell suicide (a programmed death). That cells can participate in their own execution may indicate that certain types of cell deaths that were previously considered to be caused by foreign agents such as pathogens or drugs may actually result from the activation of a programmed cell death pathway that is normally latent in cells. Here, we describe the activation of such a cell suicide pathway in plant cells upon the recognition of an invading pathogen. We discuss the possible use of this pathway as a defense mechanism against infection and the possibility that in many ways the use of this type of cell death in plants is functionally analogous to that used by mammalian cells in response to infection by pathogens.

Apoptosis↗
PubMed1997

Switching of gene expression: analysis of the factors that spatially and temporally regulate plant gene expression.

In this chapter, we have reviewed the present research and understanding of several families of transcription factors in plants. From this information, it appears there is good conservation between the types of transcription factors in plants and animals. However, there are several types of factors which have been isolated in plants that remain to be documented in animals (e.g., HD-Zip and GT). These as well as the presence of two types of TATA-binding proteins (TBPs) in plants suggest that although transcription in eukaryotes is highly conserved, fundamental differences may exist. Despite the differences, the modes of regulating transcription are well conserved. Figure 3 summarizes these modes of regulation. In recent years, the role of chromatin structure as well as subcellular localization have been the focus of a vast amount of research in mammals, Drosophila and yeast. However, very little research in these areas has been done in plants. Isolation of genes such as Curly leaf suggest a conservation of genes that influence the formation of heterochromatin-like structures. Whether or not this gene influences chromatin/heterochromatin structure in plants, however, remains to be tested. The study of nuclear localization of factors such as COP1 and KN1 is now leading to models for regulating nuclear transport as well as intercellular transport of transcription factors. Further study of the inter- and intracellular movement of these and other transcription factors may provide information on new modes of regulating transcription. In addition to understanding the role chromatin structure and subcellular localization of transcription factors may have on transcription initiation, the biological role of many plant transcription factors remains to be identified. Several approaches may be taken to understand the mechanisms by which transcription factors influence biochemical and physiological processes in the plant. These steps include 1) identification of the DNA-binding sites of the factors as well as the promoter regions which contain these sites. Presently, this approach is limiting in that not many non-coding regions have been sequenced and characterized in detail. Furthermore, the presence of a putative binding site within a promoter does not necessarily indicate that the factor will bind to the site in vivo. 2) Analysis of the binding affinity for a particular factor to a binding site in comparison to other related factors, via in vitro competition assays and quantitative titrations. This will provide information on how strongly these factors are binding to the sites, but without knowledge of all the factors present in a single cell it is difficult to recreate the in vivo conditions. 3) Generation of transgenic plants or microinjection of DNA/RNA to express a particular factor ectopically, reduce expression of the factor via antisense expression, and creation of dominant negative mutants by overexpression of key dimerization domains may provide information concerning what biological pathways these factors influence. 4) Isolation of mutations in particular transcription factors has been extremely informative in floral development. However, this approach usually entails isolation of a mutant due to a phenotype and eventual mutated locus. The cloning of the locus may or may not involve a transcription factor. 5) Many plant transcription factors have been isolated via sequence similarity to other previously identified and/or characterized transcription factors. However, the biological role of may of these factors is not known. In addition to ectopic expression of these factors by creating transgenic plants, isolation of a loss-of-function mutation may provide valuable information concerning the role of this factor in vivo. Many loss-of-function mutations in MADS box genes have led to a better understanding of how the MADS domain proteins interact with one another as well as how they influence floral development. (ABSTRACT TRUNCATED)

Cell Compartmentation↗
PubMed1997

Characterization of nuclease activities and DNA fragmentation induced upon hypersensitive response cell death and mechanical stress.

Programmed cell death (PCD) is activated during the response of multicellular organisms to some invading pathogens. One of the key aspects of this process is the degradation of nuclear DNA which is thought to facilitate the recycling of DNA from dead cells. The PCD of tobacco plants (genotype NN) infected with tobacco mosaic virus (TMV) is accompanied by the induction of nuclease activities and the cleavage of nuclear DNA to fragments of about 50 kb. We examined the correlation between the increase in nuclease activities and the fragmentation of nuclear DNA during TMV- and bacteria-induced PCD in tobacco. We found that the increase in nuclease activities did not always correlate with fragmentation of nuclear DNA. Thus, in addition to pathogens that induce PCD, mechanical injury and infiltration of leaves with 1 M sucrose or bacteria that did not induce PCD also resulted in an increase in nuclease activities. Analysis of nuclease activities in total leaf extracts, nuclear extracts, and intercellular fluid (i.e., apoplast) revealed that at least four different nuclease activities are induced during PCD in tobacco; of these at least three appear to be secreted into the intercellular fluid. Although the latter were also induced in response to treatments that did not result in DNA fragmentation, they may function in the recycling of plant DNA during late stages of PCD when the integrity of the plasma membrane is compromised. This suggestion is supported by the finding that DNA degradation occurred late during TMV-induced PCD in tobacco. In addition, the finding of induced nuclease activities in the intercellular fluid raises the possibility that they may serve a protective function by degrading the DNA of invading pathogens.

Apoptosis↗
PubMed1997

DNA-binding properties, genomic organization and expression pattern of TGA6, a new member of the TGA family of bZIP transcription factors in Arabidopsis thaliana.

The TGA genes encode a family of basic domain-leucine zipper (bZIP) transcription factors that are conserved in higher plants. We have continued to unravel the complexity of this gene family by using a polymerase chain reaction (PCR)-based approach. Taking advantage of the conserved amino acid sequence in the bZIP domain found in all members of this gene family, two degenerate oligonucleotides were synthesized based on the sequence of this region in order to amplify by PCR the analogous genomic fragments from the various TGA loci in Arabidopsis. This approach has led us to the finding of a new member of the TGA gene family, and subsequently the isolation of a gene designated as TGA6. Further characterization of the TGA6 locus confirmed our prediction that the gene structure of this family is remarkably conserved. Genomic Southern blot analysis revealed that TGA6 is a single-copy gene in Arabidopsis. Based on the genomic sequence information, gene-specific primers were synthesized for isolating the cDNA that corresponds to the coding region. Subsequently, the cDNA for TGA6 was cloned and sequenced. Gel mobility shift assays with in vitro translated TGA6 protein showed that TGA6 is more like TGA5 in terms of its in vitro DNA-binding properties. The expression of TGA6 in different tissues was estimated by using reverse transcriptase (RT)-PCR and further analyzed in transgenic Arabidopsis lines expressing a TGA6 promoter-GUS fusion. TGA6 promoter activity is found primarily in roots of young seedlings. As seedlings develop, TGA6 is expressed in aging cotyledons, mesophyll cells of hydathodes on leaf margins, vascular tissue and trichomes of senescing rosette leaves, but is very low in primary roots of mature plants. High levels of expression persist in young lateral roots and in regions of the primary root where lateral roots are emerging. In flowers, the activity is localized predominantly to mature pollen grains. The expression pattern of TGA6 reported here is strikingly similar to that of an Arabidopsis acidic chitinase gene. Possible biological significance of TGA6 in cellular defense against pathogens and abiotic stress is discussed.

Amino Acid Sequence↗
PubMed1997

Characterization of acquired resistance in lesion-mimic transgenic potato expressing bacterio-opsin.

The lesion-mimic mutants of certain plants display necrotic lesions resembling those of the hypersensitive response and activate local and systemic defense responses in the absence of pathogens. We have engineered a lesion-mimic phenotype in transgenic Russet Burbank potato plants through constitutive expression of a bacterio-opsin (bO) proton pump derived from Halobacterium halobium. Transgenic potato plants exhibiting a lesion-mimic phenotype had increased levels of salicylic acid and overexpressed several pathogenesis-related messenger RNAs, all hallmarks of systemic acquired resistance (SAR). The lesion-mimic plants also displayed enhanced resistance to the US1 isolate (A1 mating type) of a fungal pathogen, Phytophthora infestans, a causal agent of late blight disease. In contrast, little resistance was observed against the US8 isolate (A2 mating type) of this pathogen. Furthermore, a majority of the transgenic plants displaying the lesion-mimic phenotype had increased susceptibility to potato virus X. The tubers of these plants were not resistant to the bacterial pathogen Erwinia carotovora. These results indicate that expression of bO can result in the activation of defense responses in transgenic potato plants and show for the first time that bO expression can confer resistance to a pathogenic fungus. However, our results also demonstrate that like SAR, this "engineered" resistance is likely to be limited to certain pathogens and particular cultivars.

Bacteriorhodopsins↗
PubMed1997

Pathogen-induced programmed cell death in tobacco.

Sacrificing an infected cell or cells in order to prevent systemic spread of a pathogen appears to be a conserved strategy in both plants and animals. We studied some of the morphological and biochemical events that accompany programmed cell death during the hypersensitive response of tobacco plants infected with tobacco mosaic virus. Certain aspects of this cell death process appeared to be similar to those that take place during apoptosis in animal cells. These included condensation and vacuolization of the cytoplasm and cleavage of nuclear DNA to 50 kb fragments. In contrast, internucleosomal fragmentation, condensation of chromatin at the nuclear periphery and apoptotic bodies were not observed in tobacco plants during tobacco mosaic virus-induced hypersensitive response. A unique aspect of programmed cell death during the hypersensitive response of tobacco to tobacco mosaic virus involved an increase in the amount of monomeric chloroplast DNA. Morphological changes to the chloroplast and cytosol of tobacco cells and increase in monomeric chloroplast DNA occurred prior to gross changes in nuclear morphology and significant chromatin cleavage. Our findings suggest that certain aspects of programmed cell death may have been conserved during the evolution of plants and animals.

Apoptosis↗
PubMed1996

lem7, a novel temperature-sensitive Arabidopsis mutation that reversibly inhibits vegetative development.

An important question in developmental biology concerns the mechanisms by which a few cells coordinate division and differentiation to yield the complex structures and organs found in multicellular organisms. During vegetative growth in plants, cells in the apical meristem must coordinate division and differentiation to yield the fully mature leaf organ. Alterations in these processes may result in an abnormal leaf. In this paper we present the isolation and characterization of an EMS-generated, cold-temperature-sensitive mutation in Arabidopsis thaliana, designated lem7 (leaf morphogenesis). lem7 is a semidominant mutation that maps to a novel locus on chromosome 2. When grown at 16 degrees C, lem7 reversibly arrests leaf development at the shoot apex. In contrast, lem7 grown at 30 degrees C appears phenotypically normal. Our data also suggest that the Lem7 locus may not be involved solely in leaf organogenesis, but may also play a role in floral development and the maintenance of patterns and structures after cellular differentiation. At an intermediate temperature of 23 degrees C, leaves on the lem7 plant emerged phenotypically normal but began to show drastic changes at about 13 days postgermination. These changes include a reduced bilateral symmetry, a rough leaf lamina, a reduced number of trichomes, and an altered vascular network. Leaves that developed at the permissive temperature (30 degrees C) and shifted to the nonpermissive temperature (16 degrees C) form tumor-like outgrowths. Histological analysis of these tumor-like outgrowths and leaves grown at the intermediate temperature reveal abnormally large mesophyll cells, a disorganized mesophyll layer, and collapsed epidermal cells. We propose that the reversible inhibition of leaf development in lem7 under nonpermissive temperatures may serve as a useful tool for identifying genes involved in Arabidopsis leaf organogenesis.

Arabidopsis↗
PubMed1996

The conserved ELK-homeodomain of KNOTTED-1 contains two regions that signal nuclear localization.

Nuclear localization serves as a regulatory mechanism in the activity of several transcription factors. KNOTTED-1 (Kn1) is a homeodomain protein likely to regulate vegetative development in maize. At least twelve genes related to Kn1 are known in maize and six in Arabidopsis. Ectopic expression of the maize, rice and Arabidopsis Kn1-related genes have been shown to alter cell fate determination. In this paper, we study the nuclear localization capabilities of the Kn1 homeodomain and the proximal amino acid residues (the ELK region) which is highly conserved among Kn1-related homeodomain proteins. The ELK homeodomain (ELK-HD) of Kn1 was fused to the reporter gene uidA encoding the bacterial enzyme beta-glucuronidase (GUS) and transformed into tobacco and onion cells. Quantitation of GUS activity in nuclear and total protein extracts from transgenic tobacco revealed a highly localized GUS activity in the nucleus for the ELK-HD/GUS fusion protein, as compared to the basal level of GUS activity in the nucleus for the GUS only protein. The ELK-HD/GUS transformants showed no unusual characteristics, thus indicating that expression of the putative Kn1 DNA-binding domain fused to GUS may be insufficient to create a dominant negative phenotype. Histochemical analysis of the onion epidermal cells transfected by particle bombardment demonstrated that greater than 50 % of the transformed onion epidermal cells showed higher levels of GUS staining in the nucleus relative to the cytoplasm. Deletion analysis of the ELK-HD revealed that the Kn1 homeodomain comprising the three predicted alpha-helices and the conserved ELK domain can each function independently as nuclear localization signals.

Amino Acid Sequence↗
PubMed1996

Coordinated activation of as-1-type elements and a tobacco glutathione S-transferase gene by auxins, salicylic acid, methyl-jasmonate and hydrogen peroxide.

The molecular mechanism of signal transduction pathways which mediate the action of phytohormones are poorly understood. Recently, we and others have shown that the as -1 type cis-acting elements can respond to auxin and salicylic acid, two well-characterized signaling molecules in plants. In the present work, we have examined a comprehensive set of physiological and abiotic agents and found that auxin, salicylic acid and methyl-jasmonate are three effective inducers of the as-1-type elements in transgenic tobacco. Using a cell suspension culture containing a synthetic promoter-GUS fusion, we demonstrated rapid and sensitive induction of the as-1-type element by these phytohormones. Furthermore, a tobacco glutathione S-transferase gene, GNT35, that contains an as-1-type binding site in its promoter is also inducible by auxin, salicylic acid and methyl-jasmonate with similar kinetics. As Ulmasov et al. have recently reported, we found that the as-1-type elements can also respond to weak/inactive analogues of auxin and salicylic acid. In addition, we show that hydrogen peroxide can also effectively activate the expression of GNT35 as well as the as-1-type element in a cell suspension culture, but not with whole seedlings. These results are discussed with respect to the possible mechanism(s) through which a single cis element may respond to a diverse array of molecules.

Acetates↗
PubMed1996

Isolation and analysis of the soybean SGA2 gene (cDNA), encoding a new member of the plant G-protein family of signal transducers.

We have isolated a cDNA clone from Glycine max, named SGA2, coding for a G alpha-subunit protein. The encoded polypeptide, SG alpha2, shows a molecular mass of 45 kDa and contains most of the conserved regions involved in guanine nucleotide binding and hydrolysis. Comparison at the nucleotide and amino acid sequence levels with the other plant G alpha's shows a high degree of conservation (>85% similarity). Phylogenetic analysis of these plant genes with the other G alpha's from different species clearly indicate that those proteins represent a new member of the heterotrimeric G-protein family, named Gp. Tissue localization of SGA2 transcripts in root, stem and leaf organs shows that this gene is widely expressed throughout the plant although it is most abundant in the vascular tissues of all these organs. Furthermore, the transcript is more abundant in young tissues and organ primordia than mature tissues. The high degree of sequence conservation among the plant G alpha's and the differences to other species of other kingdoms, suggest that plant G proteins may function in specialized signalling processes.

Amino Acid Sequence↗
PubMed1996

Sacrifice in the face of foes: pathogen-induced programmed cell death in plants.

Inhibition of pathogen growth during the defense of plants is thought to involve a rapid process of cell death localized to the site of invasion. Recent studies suggest that the death of plant cells during this 'hypersensitive response' process is likely to result from the activation of a programmed cell death pathway.

Amino Acid Sequence↗
PubMed1996

Induction of bimodal programmed cell death in malignant cells by the derivative Ukrain (NSC-631570).

Selective induction of malignant cell death is one of the major goals of effective and safe chemotherapy. Recent developments in the understanding of programmed cell death (PCD) or apoptosis are expected to provide new leads for a safer chemotherapy. The authors investigated whether the semisynthetic alkaloid thiophosphoric acid derivative Ukrain (NSC-631570) could induce PCD or apoptosis in human K562 leukaemia cells. Results showed that Ukrain induced two distinct modalities of cell death programmes. One modality corresponded morphologically to classical apoptosis or PCD characterized by blebbing and shedding of membrane vesicles with concomitant 51Cr release; however, the Ukrain-induced apoptosis was not associated with the characteristic nuclear DNA fragmentation. Higher concentrations of Ukrain induced a second cell death programme characterized by cell surface blister formation, high specific 51Cr release and extensive DNA polyploidy. These two cell death programmes are distinct from each other in that they are interphased by a silent period characterized by normal cell morphology and reduced specific 51Cr release.

Alkaloids↗