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1.
Opt Express ; 19(17): 15864-78, 2011 Aug 15.
Article in English | MEDLINE | ID: mdl-21934949

ABSTRACT

The principle, fabrication and characterization of a dielectric MEMS cantilever located a few 100 nm above a racetrack ring resonator are presented. After fabrication of the resonators on silicon-on-insulator (SOI) wafers in a foundry process, the cantilevers were integrated by surface micromachining techniques. Off-state deflections of the cantilevers have been optimized to appropriately position them near the evanescent field of the resonator. Using electrostatic actuation, moving the cantilevers into this evanescent field, the propagation properties of the ring waveguide are modulated. We demonstrate 122 pm tuning of the resonance wavelength of the optical ring resonator (in the optical C-band) without change of the optical quality factor, on application of 9 V to a 40 µm long cantilever. This compact integrated device can be used for tuning/switching a specific wavelength, with very little energy for operation and negligible cross talk with surrounding devices.

2.
Genes Dev ; 14(23): 2938-43, 2000 Dec 01.
Article in English | MEDLINE | ID: mdl-11114883

ABSTRACT

The developmental ontogeny of the vascular system (consisting of xylem, phloem and [pro]cambium) is poorly understood despite its central role in plant physiology. We show that in the Arabidopsis root meristem, xylem cell lineages are specified early, whereas phloem and procambium are established through a set of asymmetric cell divisions. These divisions require the WOODEN LEG (WOL) gene. The WOL gene encodes a novel two-component signal transducer with an unusual tandem arrangement of two receiver domains. It is expressed specifically in the vasculature from the early stages of embryogenesis on, consistent with a role as a sensor for vascular morphogenesis.


Subject(s)
Arabidopsis/enzymology , Arabidopsis/growth & development , Protein Kinases/metabolism , Amino Acid Sequence , Arabidopsis/genetics , Arabidopsis/metabolism , Base Sequence , Cell Division , DNA, Plant , Histidine Kinase , Molecular Sequence Data , Morphogenesis , Plant Roots/growth & development , Protein Kinases/genetics , Protein Kinases/physiology
3.
Mutat Res ; 434(1): 61-5, 1999 May 14.
Article in English | MEDLINE | ID: mdl-10377949

ABSTRACT

The relevance of p53 mutations to the neoplastic malignant transformation of rodent fibroblasts by genotoxic physical and chemical agents is not clear. In the present study, we investigated p53 mutations (in exons 5-8) in non-transformed and neoplastically transformed C3H 10T1/2 and severe combined immunodeficiency (SCID) cells. No p53 mutations were detected in 15 neoplastically transformed (two spontaneous, one 3-methylcholanthrene-induced, seven gamma-ray-induced and five 'hot particle'-induced) and two non-transformed 10T1/2 cells. Wild-type p53 gene was also detected in all non-transformed (immortalized) SCID cell lines analyzed (four lines) whereas all three neoplastically transformed (two spontaneous, one gamma-ray-induced) cell lines displayed missense mutations in the p53 gene. These mutations were all transitions: A > G in codon 123, G > A in codon 152, and C > T in codon 238. We conclude that mutation in the p53 gene appears to be an infrequent event in 10T1/2 cells regardless of the transforming agent, but a frequent event in the neoplastic transformation of immortalized SCID cells. Non-transformed SCID cells are deficient in repair of DNA double-strand breaks, and neoplastically transformed cells are assumed to be deficient as well.


Subject(s)
Cell Transformation, Neoplastic/genetics , Genes, p53/genetics , Animals , Cell Line , DNA/chemistry , DNA/genetics , DNA Mutational Analysis , Genes, Tumor Suppressor/genetics , Mice , Mice, Inbred C3H , Mice, SCID , Mutation , Mutation, Missense , Point Mutation
4.
Acta Derm Venereol ; 79(3): 217-20, 1999 May.
Article in English | MEDLINE | ID: mdl-10384921

ABSTRACT

Diabetic ulcers on the lower extremities present a difficult treatment problem, and some ulcers respond poorly to conventional topical and cast treatment. The purpose of this study was to assess the effect of cultured allogeneic keratinocyte epithelium and fibroblast-gelatin sponge on the healing of chronic, refractory diabetic leg and foot ulcers. Non-diabetic chronic leg ulcers were treated for comparison. This open study comprised 22 patients with type I or type II diabetes and 16 patients with leg or ankle ulcers of different aetiologies. A total of 26 diabetic and 25 non-diabetic ulcers were treated mainly with keratinocyte epithelium and/or fibroblast-gelatin sponge once weekly until complete healing or until no further healing could be observed despite several repeated treatments. The duration of diabetic ulcers was 10.3+/-15.8 (mean+/-SD) months and the size 3.1+/-6.6 cm2. The diabetic ulcers were located in the heel (7), toe (7), sole (5), leg (6) and Achilles (1). The mean duration of non-diabetic ulcers was 6.8+/-6.0 months and the size 10.5+/-11.8 cm2. A total of 12+/-11 skin cell transplantations were performed for the diabetic ulcers. All but 1 diabetic ulcer healed during the study. The time for 50% reduction in ulcer area was 32+/-32 days, but 99+/-110 days were needed for complete ulcer closure. The longer the ulcer had existed the longer was the healing time. Heel ulcers showed significantly slower healing response than leg, sole and toe ulcers. Preliminary results suggest that both keratinocytes and fibroblasts are equally effective in the healing process. The time required for healing of the diabetic ulcers did not differ markedly from that of the non-diabetic ulcers. The results suggest that cultured allogeneic skin cells used once weekly are effective in the treatment of recalcitrant diabetic ulcers.


Subject(s)
Cell Transplantation , Diabetic Foot/surgery , Foot Ulcer/surgery , Skin/cytology , Adult , Aged , Cells, Cultured , Chronic Disease , Diabetes Mellitus, Type 1/complications , Diabetes Mellitus, Type 2/complications , Diabetic Foot/complications , Female , Foot Ulcer/complications , Humans , Keratinocytes/cytology , Male , Middle Aged , Transplantation, Homologous , Treatment Outcome , Wound Healing
5.
DNA Cell Biol ; 14(10): 841-7, 1995 Oct.
Article in English | MEDLINE | ID: mdl-7546290

ABSTRACT

We have isolated and sequenced cDNA clones that encode human spermine synthase (EC 2.5.1.22). The total length of the sequenced cDNA was 1,612 nucleotides, containing an open reading frame encoding a polypeptide chain of 368 amino acids. All of the previously sequenced peptide fragments of human and bovine spermine synthase proteins could be located within the coding region derived from the cDNA. An unusual sequence of AATTAA apparently signaled the initiation of polyadenylation. Sequence comparisons between human spermine synthase and spermidine synthases from bacterial and mammalian sources revealed a nearly complete lack of similarity between the primary structures of these two enzymes catalyzing almost identical reactions. A modest similarity found was restricted to a relatively short peptide domain apparently involved in the binding of decarboxylated S-adenosylmethionine, the common substrate for both enzymes. The apparent lack of an overall similarity may indicate that spermine synthase, the enzyme found only in eukaryotes, and spermidine synthase with more universal distribution, although functionally closely related, have evolved separately.


Subject(s)
DNA, Complementary/genetics , Genes , Spermine Synthase/genetics , Amino Acid Sequence , Animals , Base Sequence , Brain/enzymology , Cattle , Cloning, Molecular , Humans , Molecular Sequence Data , Nerve Tissue Proteins/genetics , Polymerase Chain Reaction , Sequence Alignment , Sequence Homology, Amino Acid , Species Specificity , Spermidine Synthase/genetics , Spermine/biosynthesis , Substrate Specificity
6.
FEBS Lett ; 365(1): 61-5, 1995 May 22.
Article in English | MEDLINE | ID: mdl-7774716

ABSTRACT

An increased mRNA content of spermidine synthase was found in phytohemagglutinin stimulated human peripheral lymphocytes and in cultured human myeloma (Sultan) cells stimulated to grow by change of the culture medium. The many-fold increase in the amount of the message was accompanied by stimulation of the enzyme activity in activated lymphocytes, but not in stimulated myeloma cells. In the present study the effect of the 5'-untranslated region of spermidine synthase mRNA on the post-transcriptional control of its expression was studied both in vitro in rabbit reticulocyte system and in cultured mammalian cells. The results show that the GC-rich 5'-untranslated region of spermidine synthase mRNA has an inhibitory effect on its translation.


Subject(s)
Gene Expression Regulation, Enzymologic , Protein Biosynthesis , RNA, Messenger/genetics , Regulatory Sequences, Nucleic Acid/genetics , Spermidine Synthase/biosynthesis , Base Sequence , Cells, Cultured , Humans , Lymphocyte Activation , Lymphocytes/cytology , Lymphocytes/metabolism , Molecular Sequence Data , Multiple Myeloma/metabolism , Spermidine Synthase/genetics , Tissue Distribution
7.
Biochem J ; 293 ( Pt 2): 513-6, 1993 Jul 15.
Article in English | MEDLINE | ID: mdl-8343131

ABSTRACT

We have generated a transgenic mouse line harbouring the functional (chromosome-1-derived) human spermidine synthase (EC 2.5.1.16) gene in their genome. The transgenic animals expressed the human gene-derived mRNA, as revealed by reverse-transcriptase/PCR analysis, in all tissues studied and displayed tissue spermidine synthase activity that was 2-6 times that in their syngenic littermates. The elevated spermidine synthase activity, however, had virtually no effect on tissue putrescine, spermidine or spermine levels. The view that the accumulation of spermidine and spermine is possibly controlled by S-adenosylmethionine decarboxylase was further supported by the finding that tissue spermidine and spermine contents also remained practically normal in hybrid transgenic mice over-expressing both human ornithine decarboxylase and spermidine synthase genes.


Subject(s)
Spermidine Synthase/genetics , Animals , Base Sequence , Cloning, Molecular , DNA, Single-Stranded , Female , Humans , Male , Mice , Mice, Inbred BALB C , Mice, Inbred DBA , Mice, Transgenic , Molecular Sequence Data , Polymerase Chain Reaction , Putrescine/metabolism , Spermidine/metabolism , Spermidine Synthase/metabolism , Spermine/metabolism
8.
DNA Cell Biol ; 10(6): 467-74, 1991.
Article in English | MEDLINE | ID: mdl-2069720

ABSTRACT

The human spermidine synthase (EC 2.5.1.16) gene was isolated from a genomic library constructed with DNA obtained from a human immunoglobulin G (IgG) myeloma cell line. Subsequent sequence analyses revealed that the gene comprised of 5,818 nucleotides from the cap site to the last A of the putative polyadenylation signal with 8 exons and 7 intervening sequences. The 5'-flanking region of the gene was extremely GC rich, lacking any TATA box but containing CCAAT consensus sequences. No perfect consensus sequence for the cAMP-responsive element for the AP-1 binding site was found, yet the gene contained seven AP-2 binding site consensus sequences. The putative polyadenylation signal was an unusual AATACA instead of AATAAA. Polymerase chain reaction analysis with DNA obtained from human x hamster somatic cell hybrids indicated that human spermidine synthase genomic sequences segregate with human chromosome 1. Transfection of the genomic clone into Chinese hamster ovary cells displaying a low endogenous spermidine synthase activity revealed that the gene was transiently expressed and hence in all likelihood represents a functional gene.


Subject(s)
Chromosomes, Human, Pair 1 , Spermidine Synthase/genetics , Animals , Base Sequence , Cell Line , DNA , DNA Probes , Genomic Library , Humans , Hybrid Cells , Molecular Sequence Data , Polymerase Chain Reaction , Restriction Mapping , Transfection , Tumor Cells, Cultured
9.
Biochem J ; 267(3): 767-74, 1990 May 01.
Article in English | MEDLINE | ID: mdl-2339986

ABSTRACT

The effects of selenomethionine (SeMet) on the growth of 17 cultured cell lines were studied. SeMet in the culture medium of three hepatoma cell lines promoted cell growth at subcytotoxic levels (1-20 microM), but the growth of malignant lymphoid and myeloid cells was not stimulated. L-SeMet was cytotoxic to all 17 cell lines when assayed after culture for 3-10 days. A 50% growth inhibition was observed by 30-160 microM-SeMet in a culture medium containing 100 microM-methionine. SeMet cytotoxicity to normal (fibroblasts) and malignant cells was rather similar, excluding specific antineoplastic cytotoxicity. Cytotoxicity was increased by decreasing concentrations of methionine. The DL form of SeMet was less cytotoxic than the L form. L-SeMet was metabolized to a selenium analogue of S-adenosylmethionine approximately as effectively as the natural sulphur analogue methionine in malignant R1.1 lymphoblasts. Concomitantly, S-adenosylmethionine pools were decreased. This occurred early and at cytotoxic SeMet levels. Methionine adenosyltransferase activity was not altered by SeMet treatment. ATP pools were not affected early, and decreases in the synthesis of DNA and protein took place late and were apparently related to cell death. RNA synthesis was slightly stimulated at low cytotoxic SeMet levels by 24 h, but was markedly inhibited after 48 h. The SeMet analogue of S-adenosylmethionine could be effectively utilized in a specific enzymic transmethylation. Neither S-adenosylhomocysteine nor its selenium analogue accumulated in the treated cells. These findings together suggest a direct or indirect involvement of S-adenosylmethionine metabolism in SeMet cytotoxicity, but exclude a gross blockage of transmethylations.


Subject(s)
S-Adenosylmethionine/metabolism , Selenium/pharmacology , Selenomethionine/pharmacology , Cell Division/drug effects , Humans , Methionine/pharmacology , Methionine Adenosyltransferase/analysis , Methylation , Nucleotides/analysis , Tumor Cells, Cultured
10.
DNA Cell Biol ; 9(2): 103-10, 1990 Mar.
Article in English | MEDLINE | ID: mdl-2344393

ABSTRACT

Using a synthetic deoxyoligonucleotide mixture constructed for a tryptic peptide of the bovine enzyme as a probe, cDNA coding for the full-length subunit of spermidine synthase was isolated from a human decidual cDNA library constructed on phage lambda gt11. After subcloning into the Eco RI site of pBR322 and propagation, both strands of the insert were sequenced using a shotgun strategy. Starting from the first start codon, which was immediately preceded by a GC-rich region including four overlapping CCGCC consensus sequences, an open reading frame for a 302-amino-acid polypeptide was resolved. This peptide had an Mr of 33,827, started with methionine, and ended with serine. The identity of the isolated cDNA was confirmed by comparison of the deduced amino acid sequence with resolved sequences of the tryptic peptides of bovine spermidine synthase. The coding strand of the cDNA revealed no special regulatory or ribosome-binding signals within 82 nucleotides preceding the start codon and no polyadenylation signal within 247 nucleotides following the stop codon. The coding region, containing a 13-nucleotide repeat close to the 5' end, was longer than, and very different from, that of the bacterial counterpart. This region seems to be of retroviral origin and shows marked homology with sequences found in a variety of human, mammalian, avian, and viral genes and mRNAs. By computer analysis, the first 200 nucleotides of the 5' end of the coding strand appear able to form a very stable secondary structure with a free energy change of -157.6 kcal/mole.(ABSTRACT TRUNCATED AT 250 WORDS)


Subject(s)
Spermidine Synthase/genetics , Transferases/genetics , Amino Acid Sequence , Base Sequence , Blotting, Northern , Cloning, Molecular , Decidua/enzymology , Female , Humans , Molecular Sequence Data , Nucleic Acid Conformation , Pregnancy , RNA, Messenger/genetics
11.
Biochem J ; 259(3): 879-86, 1989 May 01.
Article in English | MEDLINE | ID: mdl-2730590

ABSTRACT

Spermidine synthase was purified to apparent homogeneity from human spleens (8700-fold) by affinity chromatography. The native enzyme was composed of two subunits of identical Mr (35,000) and showed an apparent Mr of 62,000 in pore-gradient gel electrophoresis. Its pI was 5.1, Spermine synthase was purified to apparent homogeneity from placenta (5300-fold) and from kidney (4600-fold). The native enzyme was composed of two subunits of identical Mr (45,000) and showed an apparent Mr of 78,000 in pore-gradient gel electrophoresis. In isoelectric focusing it revealed two bands, with pI values of 4.9 and 5.0. Both synthases were present in all human tissues studied, but revealed a clear tissue-specific pattern. Mouse antisera against spermidine synthase revealed only one band, of Mr 35,000, in all purified enzyme preparations and in crude human tissue extracts in immunoblotting. Antisera against spermine synthase showed an immunoreactive band corresponding to the Mr of the subunit of spermine synthase. These antisera did not indicate any cross-reactivity in immunoblotting. Thus spermine synthase and spermidine synthase do not share homologous antigenic sites and are totally different proteins.


Subject(s)
Spermidine Synthase/isolation & purification , Spermine Synthase/isolation & purification , Transferases/isolation & purification , Blotting, Western , Chromatography, Affinity , Electrophoresis, Polyacrylamide Gel , Humans , Isoelectric Focusing , Kinetics , Spermidine Synthase/metabolism , Spermine Synthase/metabolism
13.
Biochem J ; 231(1): 165-9, 1985 Oct 01.
Article in English | MEDLINE | ID: mdl-3933487

ABSTRACT

The putrescine-stimulated S-adenosyl-L-methionine decarboxylases from rat liver and yeast were strongly inhibited by Berenil and to a lesser extent by Pentamidine. Ten times greater drug concentrations were needed to achieve a similar level of inhibition of a Mg2+-stimulated bacterial enzyme. The inhibition was irreversible in that extensive dialyses or precipitation with (NH4)2SO4 did not restore enzyme activity. Putrescine did not protect the enzyme against Berenil, but adenosylmethionine either alone or with putrescine partially protected the irreversible action of Berenil. The compound 4,4'-diamidinodiphenylamine, which differs from Berenil only in lacking the azo group between benzene rings, was a weaker inhibitor than Berenil, and its inhibition was reversible. Berenil also inhibited the activity of adenosylmethionine decarboxylase in vivo, by depressing the activity of the enzyme in normal rat liver, for at least 24 h after a single injection (50 mg/kg body wt.) of the drug.


Subject(s)
Adenosylmethionine Decarboxylase/antagonists & inhibitors , Amidines/pharmacology , Carboxy-Lyases/antagonists & inhibitors , Diminazene/pharmacology , Pentamidine/pharmacology , Putrescine/pharmacology , Animals , Diminazene/analogs & derivatives , Diphenylamine/analogs & derivatives , Diphenylamine/pharmacology , Liver/enzymology , Pseudomonas aeruginosa/enzymology , Rats , S-Adenosylmethionine/pharmacology , Saccharomyces cerevisiae/enzymology
14.
Biochem Biophys Res Commun ; 130(2): 596-602, 1985 Jul 31.
Article in English | MEDLINE | ID: mdl-3861182

ABSTRACT

1-Aminooxy-3-aminopropane was shown to be a potent competitive inhibitor (Ki = 3.2 nM) of homogenous mouse kidney ornithine decarboxylase, a potent irreversible inhibitor (Ki = 50 microM) of homogeneous liver adenosylmethionine decarboxylase and a potent competitive (Ki = 2.3 microM) of homogeneous bovine brain spermidine synthase. It did not inhibit homogeneous bovine brain spermine synthase and it did not serve as a substrate for spermidine synthase. The compound did not inhibit tyrosine aminotransferase, alanine aminotransferase or aspartate aminotransferase, which are pyridoxal phosphate-containing enzymes like ornithine decarboxylase. The inactivation of adenosylmethionine decarboxylase was partially prevented by pyruvate, which is the coenzyme of adenosylmethionine decarboxylase, and by the substrate, adenosylmethionine. 1-Aminooxy-3-aminopropane at 0.5 mM concentration inhibited the growth of HL-60 promyelocytic leukemia cells and this inhibition was prevented by spermidine but not by putrescine.


Subject(s)
Polyamines/biosynthesis , Propylamines/pharmacology , Adenosylmethionine Decarboxylase/antagonists & inhibitors , Animals , Brain/enzymology , Cattle , Humans , Kidney/enzymology , Leukemia, Myeloid, Acute/enzymology , Mice , Ornithine Decarboxylase Inhibitors , Putrescine/pharmacology , Rats , Spermidine/pharmacology , Spermidine Synthase/antagonists & inhibitors
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