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1.
Neuron ; 32(5): 787-800, 2001 Dec 06.
Article in English | MEDLINE | ID: mdl-11738026

ABSTRACT

Transport of synaptic components is a regulated process. Loss-of-function mutations in the C. elegans unc-16 gene result in the mislocalization of synaptic vesicle and glutamate receptor markers. unc-16 encodes a homolog of mouse JSAP1/JIP3 and Drosophila Sunday Driver. Like JSAP1/JIP3, UNC-16 physically interacts with JNK and JNK kinases. Deletion mutations in Caenorhabditis elegans JNK and JNK kinases result in similar mislocalization of synaptic vesicle markers and enhance weak unc-16 mutant phenotypes. unc-116 kinesin heavy chain mutants also mislocalize synaptic vesicle markers, as well as a functional UNC-16::GFP. Intriguingly, unc-16 mutations partially suppress the vesicle retention defect in unc-104 KIF1A kinesin mutants. Our results suggest that UNC-16 may regulate the localization of vesicular cargo by integrating JNK signaling and kinesin-1 transport.


Subject(s)
Adaptor Proteins, Signal Transducing , Caenorhabditis elegans Proteins/metabolism , Caenorhabditis elegans Proteins/physiology , Caenorhabditis elegans/physiology , Mitogen-Activated Protein Kinases/metabolism , Proto-Oncogene Proteins c-jun/metabolism , Signal Transduction/physiology , Synaptic Vesicles/metabolism , Animals , Biological Transport, Active/genetics , Caenorhabditis elegans/genetics , Caenorhabditis elegans/metabolism , Caenorhabditis elegans Proteins/genetics , Carrier Proteins/genetics , Carrier Proteins/metabolism , Cell Line , Drosophila , Gene Deletion , Green Fluorescent Proteins , Humans , JNK Mitogen-Activated Protein Kinases , Luminescent Proteins/genetics , Luminescent Proteins/metabolism , Mice , Mitogen-Activated Protein Kinases/genetics , Molecular Sequence Data , Mutation/physiology , Nerve Tissue Proteins/genetics , Nerve Tissue Proteins/metabolism , Neurons/metabolism , Proto-Oncogene Proteins c-jun/genetics , Rabbits , Receptors, Glutamate/genetics , Receptors, Glutamate/metabolism , Signal Transduction/genetics , Synaptic Vesicles/genetics
2.
EMBO Rep ; 2(6): 530-5, 2001 Jun.
Article in English | MEDLINE | ID: mdl-11415987

ABSTRACT

The Caenorhabditis elegans excretory cell extends tubular processes, called canals, along the basolateral surface of the epidermis. Mutations in the exc-5 gene cause tubulocystic defects in this canal. Ultrastructural analysis suggests that exc-5 is required for the proper placement of cytoskeletal elements at the apical epithelial surface. exc-5 encodes a protein homologous to guanine nucleotide exchange factors and contains motif architecture similar to that of FGD1, which is responsible for faciogenital dysplasia. exc-5 interacts genetically with mig-2, which encodes Rho GTPase. These results suggest that EXC-5 controls the structural organization of the excretory canal by regulating Rho family GTPase activities.


Subject(s)
Caenorhabditis elegans Proteins , Caenorhabditis elegans/embryology , Guanine Nucleotide Exchange Factors/genetics , Guanine Nucleotide Exchange Factors/physiology , Helminth Proteins/genetics , Helminth Proteins/physiology , Amino Acid Sequence , Animals , Animals, Genetically Modified , Caenorhabditis elegans/chemistry , Caenorhabditis elegans/ultrastructure , Chromosome Mapping , DNA, Complementary/metabolism , Exons , Introns , Microscopy, Electron , Models, Genetic , Molecular Sequence Data , Mutation , Plasmids/metabolism , Protein Binding
3.
Cell ; 105(2): 221-32, 2001 Apr 20.
Article in English | MEDLINE | ID: mdl-11336672

ABSTRACT

A stochastic cell fate decision mediated by axon contact and calcium signaling causes one of the two bilaterally symmetric AWC neurons, either AWCL or AWCR, to express the candidate olfactory receptor str-2. nsy-1 mutants express str-2 in both neurons, disrupting AWC asymmetry. nsy-1 encodes a homolog of the human MAP kinase kinase kinase (MAPKKK) ASK1, an activator of JNK and p38 kinases. Based on genetic epistasis analysis, nsy-1 appears to act downstream of the CaMKII unc-43, and NSY-1 associates with UNC-43, suggesting that UNC-43/CaMKII activates the NSY-1 MAP kinase cassette. Mosaic analysis demonstrates that UNC-43 and NSY-1 act primarily in a cell-autonomous execution step that represses str-2 expression in one AWC cell, downstream of the initial lateral signaling pathway that coordinates the fates of the two cells.


Subject(s)
Calcium-Calmodulin-Dependent Protein Kinases/metabolism , Epistasis, Genetic , MAP Kinase Kinase Kinases/metabolism , MAP Kinase Signaling System/physiology , Mosaicism/genetics , Olfactory Receptor Neurons/physiology , Receptors, Odorant/metabolism , Transgenes/genetics , Amino Acid Sequence , Animals , Caenorhabditis elegans/cytology , Caenorhabditis elegans/embryology , Caenorhabditis elegans/genetics , Caenorhabditis elegans Proteins , Calcium-Calmodulin-Dependent Protein Kinase Type 2 , Cell Line , Gene Targeting , Genes, Reporter/genetics , Humans , Immunoblotting , MAP Kinase Kinase Kinases/genetics , MAP Kinase Signaling System/genetics , Microscopy, Confocal , Molecular Sequence Data , Phosphorylation , Receptors, Odorant/genetics , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/metabolism , Sequence Alignment
4.
J Biol Chem ; 276(26): 24396-400, 2001 Jun 29.
Article in English | MEDLINE | ID: mdl-11323434

ABSTRACT

TAK1, a member of the MAPKKK family, is involved in the intracellular signaling pathways mediated by transforming growth factor beta, interleukin 1, and Wnt. TAK1 kinase activity is specifically activated by the TAK1-binding protein TAB1. The C-terminal 68-amino acid sequence of TAB1 (TAB1-C68) is sufficient for TAK1 interaction and activation. Analysis of various truncated versions of TAB1-C68 defined a C-terminal 30-amino acid sequence (TAB1-C30) necessary for TAK1 binding and activation. NMR studies revealed that the TAB1-C30 region has a unique alpha-helical structure. We identified a conserved sequence motif, PYVDXA/TXF, in the C-terminal domain of mammalian TAB1, Xenopus TAB1, and its Caenorhabditis elegans homolog TAP-1, suggesting that this motif constitutes a specific TAK1 docking site. Alanine substitution mutagenesis showed that TAB1 Phe-484, located in the conserved motif, is crucial for TAK1 binding and activation. The C. elegans homolog of TAB1, TAP-1, was able to interact with and activate the C. elegans homolog of TAK1, MOM-4. However, the site in TAP-1 corresponding to Phe-484 of TAB1 is an alanine residue (Ala-364), and changing this residue to Phe abrogates the ability of TAP-1 to interact with and activate MOM-4. These results suggest that the Phe or Ala residue within the conserved motif of the TAB1-related proteins is important for interaction with and activation of specific TAK1 MAPKKK family members in vivo.


Subject(s)
HIV Envelope Protein gp120/chemistry , HIV Envelope Protein gp120/metabolism , MAP Kinase Kinase Kinases/metabolism , Recombinant Fusion Proteins/chemistry , Recombinant Fusion Proteins/metabolism , Amino Acid Motifs , Amino Acid Sequence , Animals , Binding Sites , Caenorhabditis elegans/genetics , Cell Line , Conserved Sequence , Enzyme Activation , Evolution, Molecular , HIV Envelope Protein gp120/genetics , Molecular Sequence Data , Mutation , Phenylalanine/genetics , Protein Structure, Tertiary , Recombinant Fusion Proteins/genetics , Sequence Homology, Amino Acid , Xenopus/genetics
5.
Mech Dev ; 99(1-2): 51-64, 2000 Dec.
Article in English | MEDLINE | ID: mdl-11091073

ABSTRACT

SR-protein kinases (SRPKs) and their substrates, serine/arginine-rich pre-mRNA splicing factors, are key components of splicing machinery and are well conserved across phyla. Despite extensive biochemical investigation, the physiological functions of SRPKs remain unclear. In the present study, cDNAs for SPK-1, a C. elegans SRPK homologue, and CeSF2, an SPK-1 substrate, were cloned. SPK-1 binds directly to and phosphorylates the RS domain of CeSF2 in vitro. Both spk-1 and CeSF2 are predominantly expressed in germlines. RNA interference (RNAi) experiments revealed that spk-1 and CeSF2 play an essential role at the embryonic stage of C. elegans. Furthermore, RNAi studies demonstrated that spk-1 is required for germline development in C. elegans. We provide evidence that RNAi, achieved by the soaking of L1 larvae, is beneficial in the study of gene function in post-embryonic germline development.


Subject(s)
Caenorhabditis elegans/enzymology , Embryo, Nonmammalian/physiology , src-Family Kinases/chemistry , src-Family Kinases/physiology , Amino Acid Sequence , Animals , Blotting, Northern , DNA, Complementary/metabolism , Evolution, Molecular , Female , Fluorescent Dyes/pharmacology , Glutathione Transferase/metabolism , In Situ Hybridization , Indoles/pharmacology , Infertility/genetics , Male , Molecular Sequence Data , Mothers , Phenotype , Phosphorylation , Protein Binding , Protein Structure, Tertiary , RNA/metabolism , Recombinant Proteins/metabolism , Sequence Homology, Amino Acid , Time Factors
6.
Science ; 288(5474): 2205-8, 2000 Jun 23.
Article in English | MEDLINE | ID: mdl-10864868

ABSTRACT

In Caenorhabditis elegans, the gonad acquires two U-shaped arms by the directed migration of its distal tip cells (DTCs) along the body wall basement membranes. Correct migration of DTCs requires the mig-17 gene, which encodes a member of the metalloprotease-disintegrin protein family. The MIG-17 protein is secreted from muscle cells of the body wall and localizes in the basement membranes of gonad. This localization is dependent on the disintegrin-like domain of MIG-17 and its catalytic activity. These results suggest that the MIG-17 metalloprotease directs migration of DTCs by remodeling the basement membrane.


Subject(s)
Caenorhabditis elegans Proteins , Caenorhabditis elegans/enzymology , Disintegrins/metabolism , Extracellular Matrix/metabolism , Metalloendopeptidases/metabolism , Amino Acid Sequence , Animals , Basement Membrane/enzymology , Caenorhabditis elegans/cytology , Caenorhabditis elegans/genetics , Caenorhabditis elegans/growth & development , Cell Movement , Cloning, Molecular , Disintegrins/chemistry , Disintegrins/genetics , Gene Expression Profiling , Genes, Helminth , Glycosylation , Gonads/cytology , Gonads/enzymology , Gonads/growth & development , Metalloendopeptidases/chemistry , Metalloendopeptidases/genetics , Molecular Sequence Data , Muscles/cytology , Muscles/enzymology , Mutation , Protein Structure, Tertiary , Recombinant Fusion Proteins/chemistry , Recombinant Fusion Proteins/metabolism , Sequence Alignment , Transgenes
7.
Nature ; 399(6738): 793-7, 1999 Jun 24.
Article in English | MEDLINE | ID: mdl-10391246

ABSTRACT

The signalling protein Wnt regulates transcription factors containing high-mobility-group (HMG) domains to direct decisions on cell fate during animal development. In Caenorhabditis elegans, the HMG-domain-containing repressor POP-1 distinguishes the fates of anterior daughter cells from their posterior sisters throughout development, and Wnt signalling downregulates POP-1 activity in one posterior daughter cell called E. Here we show that the genes mom-4 and lit-1 are also required to downregulate POP-1, not only in E but also in other posterior daughter cells. Consistent with action in a common pathway, mom-4 and lit-1 exhibit similar mutant phenotypes and encode components of the mitogen-activated protein kinase (MAPK) pathway that are homologous to vertebrate transforming-growth-factor-beta-activated kinase (TAK1) and NEMO-like kinase (NLK), respectively. Furthermore, MOM-4 and TAK1 bind related proteins that promote their kinase activities. We conclude that a MAPK-related pathway cooperates with Wnt signal transduction to downregulate POP-1 activity. These functions are likely to be conserved in vertebrates, as TAK1 and NLK can downregulate HMG-domain-containing proteins related to POP-1.


Subject(s)
Adaptor Proteins, Signal Transducing , Caenorhabditis elegans Proteins , Caenorhabditis elegans/genetics , Calcium-Calmodulin-Dependent Protein Kinases/metabolism , DNA-Binding Proteins/genetics , Down-Regulation , High Mobility Group Proteins/genetics , Intracellular Signaling Peptides and Proteins , JNK Mitogen-Activated Protein Kinases , MAP Kinase Kinase Kinases , Mitogen-Activated Protein Kinase Kinases , Mitogen-Activated Protein Kinases , Proto-Oncogene Proteins/metabolism , Zebrafish Proteins , Amino Acid Sequence , Animals , Caenorhabditis elegans/enzymology , Caenorhabditis elegans/metabolism , Carrier Proteins/genetics , Carrier Proteins/metabolism , Cell Line , Cloning, Molecular , Enzyme Activation , Gene Expression Regulation, Developmental , Genes, Helminth , Humans , MAP Kinase Kinase 4 , Membrane Proteins/genetics , Membrane Proteins/metabolism , Molecular Sequence Data , Potassium Channels, Voltage-Gated , Protein Kinases/metabolism , Protein Serine-Threonine Kinases/chemistry , Protein Serine-Threonine Kinases/genetics , Protein Serine-Threonine Kinases/metabolism , Sequence Homology, Amino Acid , Signal Transduction , Wnt Proteins
8.
EMBO J ; 18(13): 3604-15, 1999 Jul 01.
Article in English | MEDLINE | ID: mdl-10393177

ABSTRACT

The c-Jun N-terminal kinase (JNK) of the MAP kinase superfamily is activated in response to a variety of cellular stresses and is involved in apoptosis in neurons. However, the roles of the JNK signaling pathway in the nervous system are unknown. The genes for the Caenorhabditis elegans homolog of JNK, JNK-1, and its direct activator, JKK-1, were isolated based on their abilities to function in the Hog1 MAP kinase pathway in yeast. JKK-1 is a member of the MAP kinase kinase superfamily and functions as a specific activator of JNK. Both jnk-1 and jkk-1 are expressed in most neurons. jkk-1 null mutant animals exhibit defects in locomotion that can be rescued by the conditional expression of JKK-1 in mutant adults, suggesting that the defect is not due to a developmental error. Furthermore, ectopic expression of JKK-1 in type-D motor neurons is sufficient to rescue the movement defect. Thus, the C.elegans JNK pathway functions in type-D GABAergic motor neurons and thereby modulates coordinated locomotion.


Subject(s)
Caenorhabditis elegans Proteins , Caenorhabditis elegans/enzymology , Calcium-Calmodulin-Dependent Protein Kinases/metabolism , Mitogen-Activated Protein Kinase Kinases , Mitogen-Activated Protein Kinases , Motor Neurons/physiology , Protein Kinases/metabolism , Signal Transduction , gamma-Aminobutyric Acid/physiology , Amino Acid Sequence , Animals , Axons/enzymology , Caenorhabditis elegans/cytology , Caenorhabditis elegans/genetics , Caenorhabditis elegans/physiology , Calcium-Calmodulin-Dependent Protein Kinases/chemistry , Calcium-Calmodulin-Dependent Protein Kinases/genetics , Cell Line , Enzyme Activation , Gene Deletion , Gene Expression , Genes, Fungal , Genes, Helminth , Genetic Complementation Test , Humans , JNK Mitogen-Activated Protein Kinases , MAP Kinase Kinase 4 , Molecular Sequence Data , Motor Neurons/cytology , Motor Neurons/enzymology , Movement , Phylogeny , Protein Kinases/chemistry , Protein Kinases/genetics , Substrate Specificity
9.
J Biol Chem ; 274(15): 9976-83, 1999 Apr 09.
Article in English | MEDLINE | ID: mdl-10187773

ABSTRACT

To identify genes responsive to cold stress, we employed the differential display mRNA analysis technique to isolate a novel gene from Tetrahymena thermophila which encodes a protein kinase of 430 amino acids. A homolog of this kinase with 90% amino acid sequence identity was also found in T. pyriformis. Both kinases contain 11 subdomains typical of protein kinases. Sequence analysis revealed that the predicted amino acid sequences resemble those of mitogen-activated protein kinase (MAPK), especially p38 and stress-activated protein kinase which are known to be involved in various stress responses. However, it should be noted that the tyrosine residue in the normally conserved MAPK phosphorylation site (Thr-X-Tyr) is replaced by histidine (Thr226-Gly-His228) in this MAPK-related kinase (MRK). The recombinant MRK expressed in Escherichia coli phosphorylated myelin basic protein (MBP) and became autophosphorylated. However, the mutated recombinant protein in which Thr226 was replaced by Ala lost the ability to phosphorylate MBP, suggesting that Thr226 residue is essential for kinase activity. The MRK mRNA transcript in T. thermophila increased markedly upon temperature downshift from 35 to 15 degrees C (0.8 degrees C/min). Interestingly, osmotic shock either by sorbitol (100-200 mM) or NaCl (25-100 mM) also induced mRNA expression of the MRK in T. pyriformis. In addition, the activity of the kinase as determined by an immune complex kinase assay using MBP as a substrate was also induced by osmotic stress. This is the first demonstration of a MAPK-related kinase in the unicellular eukaryotic protozoan Tetrahymena that is induced by physical stresses such as cold temperature and osmolarity. The present results suggest that this MRK may function in the stress-signaling pathway in Tetrahymena cells.


Subject(s)
Calcium-Calmodulin-Dependent Protein Kinases/biosynthesis , Mitogen-Activated Protein Kinases , Tetrahymena thermophila/enzymology , Amino Acid Sequence , Animals , Calcium-Calmodulin-Dependent Protein Kinases/genetics , Cloning, Molecular , DNA, Complementary/isolation & purification , DNA, Protozoan/chemistry , Gene Expression Regulation, Enzymologic , JNK Mitogen-Activated Protein Kinases , Molecular Sequence Data , Osmolar Concentration , Phylogeny , Restriction Mapping , Temperature , Tetrahymena pyriformis/enzymology , Tetrahymena pyriformis/genetics , Tetrahymena thermophila/genetics
10.
Am J Med Sci ; 317(1): 59-62, 1999 Jan.
Article in English | MEDLINE | ID: mdl-9892274

ABSTRACT

Various extraintestinal manifestations may occur in juvenile polyposis, but hereditary hemorrhagic telangiectasia has rarely been reported in this type of polyposis. The authors treated a 14-year-old girl with rectal bleeding and anemia who had multiple polyps of the colorectum. Large polyps were removed and histologically diagnosed as juvenile polyps. She was later diagnosed as having telangiectasia of the skin, and arteriovenous malformations in the lung and in the liver, all of which were compatible with the diagnosis of hereditary hemorrhagic telangiectasia. At age 32, she had multiple ulcers in the ileum and in the colon. The coexistence of juvenile polyposis and hereditary hemorrhagic telangiectasia may be a clue for the understanding of the histopathogenesis of juvenile polyposis.


Subject(s)
Colonic Polyps/genetics , Telangiectasia, Hereditary Hemorrhagic/complications , Adolescent , Arteriovenous Malformations/genetics , Colonic Polyps/pathology , Female , Humans , Pulmonary Artery/abnormalities , Pulmonary Veins/abnormalities
11.
Mol Cell Biol ; 15(7): 3767-76, 1995 Jul.
Article in English | MEDLINE | ID: mdl-7791784

ABSTRACT

The Saccharomyces cerevisiae GLC7 gene encodes the catalytic subunit of type 1 protein phosphatase (PP1) and is required for cell growth. A cold-sensitive glc7 mutant (glc7Y170) arrests in G2/M but remains viable at the restrictive temperature. In an effort to identify additional gene products that function in concert with PP1 to regulate growth, we isolated a mutation (gpp1) that exacerbated the growth phenotype of the glc7Y170 mutation, resulting in rapid death of the double mutant at the nonpermissive temperature. We identified an additional gene, EGP1, as an extra-copy suppressor of the glc7Y170 gpp1-1 double mutant. The nucleotide sequence of EGP1 predicts a leucine-rich repeat protein that is similar to Sds22, a protein from the fission yeast Schizosaccharomyces pombe that positively modulates PP1. EGP1 is essential for cell growth but becomes dispensable upon overexpression of the GLC7 gene. Egp1 and PP1 directly interact, as assayed by coimmunoprecipitation. These results suggest that Egp1 functions as a positive modulator of PP1 in the growth control of S. cerevisiae.


Subject(s)
Genes, Fungal/genetics , Genes, Regulator/genetics , Nuclear Proteins , Phosphoprotein Phosphatases/metabolism , Repressor Proteins/genetics , Saccharomyces cerevisiae Proteins , Saccharomyces cerevisiae/growth & development , Schizosaccharomyces pombe Proteins , Amino Acid Sequence , Base Sequence , Cell Cycle/genetics , Cell Cycle Proteins , Cell Nucleus/pathology , Cloning, Molecular , Fluorescent Antibody Technique , Fungal Proteins/genetics , Fungal Proteins/isolation & purification , Gene Expression Regulation, Fungal , Genes, Lethal/genetics , Microtubules/pathology , Molecular Sequence Data , Mutagenesis , Phosphoprotein Phosphatases/isolation & purification , Precipitin Tests , Repressor Proteins/isolation & purification , Saccharomyces cerevisiae/cytology , Saccharomyces cerevisiae/genetics , Sequence Homology, Amino Acid
12.
Mol Cell Biol ; 14(5): 3158-65, 1994 May.
Article in English | MEDLINE | ID: mdl-8164671

ABSTRACT

We isolated a mutant carrying a conditional mutation in the GLC7 gene, encoding the catalytic subunit of a type 1 protein phosphatase, by selection of suppressors that restored the growth defect of cdc24 mutants at high temperature and simultaneously conferred cold-sensitive growth. This cold sensitivity for growth is caused by a single mutation (glc7Y-170) at position 170 of the Glc7 protein, resulting in replacement of cysteine with tyrosine. Genetic analysis suggested that the glc7Y-170 allele is associated with a recessive negative phenotype, reducing the activity of Glc7 in the cell. The glc7Y-170 mutant missegregated chromosome III at the permissive temperature, arrested growth as large-budded cells at the restrictive temperature, exhibited a significant increase in the number of nuclei at or in the neck, and had a short spindle. Furthermore, the glc7Y-170 mutant exhibited a high level of CDC28-dependent protein kinase activity when incubated at the restrictive temperature. These findings suggest that the glc7Y-170 mutation is defective in the G2/M phase of the cell cycle. Thus, type 1 protein phosphatase in Saccharomyces cerevisiae is essential for the G2/M transition.


Subject(s)
Cell Cycle , Chromosomes, Fungal , Fungal Proteins/metabolism , Genes, Fungal , Phosphoprotein Phosphatases/metabolism , Saccharomyces cerevisiae/enzymology , Saccharomyces cerevisiae/growth & development , Amino Acid Sequence , Base Sequence , CDC28 Protein Kinase, S cerevisiae/metabolism , Chromosome Mapping , Cloning, Molecular , Fungal Proteins/biosynthesis , Fungal Proteins/genetics , G2 Phase , Genetic Complementation Test , Genotype , Glycogen/metabolism , Mitosis , Molecular Sequence Data , Mutagenesis , Phenotype , Phosphoprotein Phosphatases/biosynthesis , Phosphoprotein Phosphatases/genetics , Plasmids , Protein Kinases/metabolism , Restriction Mapping , Saccharomyces cerevisiae/genetics , Sequence Homology, Amino Acid
14.
J Cell Sci ; 102 ( Pt 3): 393-400, 1992 Jul.
Article in English | MEDLINE | ID: mdl-1506422

ABSTRACT

The RCC1 protein that is required for coupling mitosis with the S phase has a DNA-binding domain in the N-terminal region outside the repeat. We found that RCC1 protein without any DNA-binding activity complemented the tsBN2 mutation with the same efficiency as that of intact RCC1 protein. In ts+ transformants of tsBN2 cells transfected with the RCC1 cDNA lacking the DNA-binding domain, an endogenous RCC1 disappeared at 39.5 degrees C, and the deleted RCC1 protein encoded by the transfected cDNA was found in the cytoplasm, but a significant amount of it was also found in the nuclei. This deleted RCC1 protein was eluted from the nuclei with the same concentration of NaCl and DNase I as was used for the intact RCC1 protein in BHK21 cells. Furthermore, the deleted RCC1 protein co-migrated with the nucleosome fraction on sucrose density gradient analysis. These results indicate that the RCC1 protein binds chromatin with the aid of other unknown protein(s). Thus, the DNA-binding domain of RCC1 protein is not essential for coupling between the S and M phases, but was shown instead to function as a nuclear translocation signal.


Subject(s)
Cell Cycle Proteins , DNA Replication/physiology , DNA-Binding Proteins/physiology , Guanine Nucleotide Exchange Factors , Mitosis/physiology , Amino Acid Sequence , Animals , Base Sequence , Cell Line , Cell Nucleus/metabolism , Chromatin/metabolism , Chromosome Deletion , DNA Mutational Analysis , Mice , Molecular Sequence Data , Nuclear Proteins/physiology , Protein Sorting Signals , Transfection
15.
Mol Cell Biol ; 11(6): 3317-25, 1991 Jun.
Article in English | MEDLINE | ID: mdl-2038334

ABSTRACT

The human CCG1 gene complements tsBN462, a temperature-sensitive G1 mutant of the BHK21 cell line. The previously cloned cDNA turned out to be a truncated form of the actual CCG1 cDNA. The newly cloned CCG1 cDNA was 6.0 kb and encoded a protein with a molecular mass of 210 kDa. Using an antibody to a predicted peptide from the CCG1 protein, a protein with a molecular mass of over 200 kDa was identified in human, monkey, and hamster cell lines. In the newly defined C-terminal region, an acidic domain was found. It contained four consensus target sequences for casein kinase II and was phosphorylated by this enzyme in vitro. However, this C-terminal region was not required to complement tsBN462 mutation since the region encoding the C-terminal part was frequently missing in complemented clones derived by DNA-mediated gene transfer. CCG1 contains a sequence similar to the putative DNA-binding domain of HMG1 in addition to the previously detected amino acid sequences common in nuclear proteins, such as a proline cluster and a nuclear translocation signal. Consistent with these predictions, CCG1 was present in nuclei, possessed DNA-binding activity, and was eluted with similar concentrations of salt, 0.3 to 0.4 M NaCl either from isolated nuclei or from a DNA-cellulose column.


Subject(s)
DNA-Binding Proteins/genetics , G1 Phase , Nuclear Proteins/genetics , TATA-Binding Protein Associated Factors , Transcription Factor TFIID , Amino Acid Sequence , Animals , Base Sequence , Cell Line , Cloning, Molecular , Codon/genetics , DNA/genetics , DNA/isolation & purification , DNA-Binding Proteins/isolation & purification , DNA-Binding Proteins/metabolism , Escherichia coli/genetics , Histone Acetyltransferases , Humans , Molecular Sequence Data , Molecular Weight , Nuclear Proteins/isolation & purification , Nuclear Proteins/metabolism , Oligonucleotide Probes , Polymerase Chain Reaction , Recombinant Proteins/isolation & purification , Sequence Homology, Nucleic Acid , Temperature
17.
Acta Endocrinol (Copenh) ; 120(6): 729-34, 1989 Jun.
Article in English | MEDLINE | ID: mdl-2786310

ABSTRACT

In a cross-sectional health screening 636 persons with negative urine glucose, a 75-g-oral glucose tolerance test was performed. We report the clinical features of the subjects with impaired glucose tolerance or diabetes mellitus. In 96 subjects with impaired glucose tolerance, the frequencies of alcohol dependency, fatty liver, and of increased levels of serum uric acid, cholesterol, triglycerides, total serum protein and gamma-glutamyl transpeptidase were significantly higher than in normal subjects. In 37 subjects with diabetes mellitus, the frequencies of fatty liver, hypertension and of increased erythrocyte sedimentation rate, triglycerides and gamma-glutamyl transpeptidase were significantly higher than in normal subjects. In addition, significant increases in serum gamma-glutamyl transpeptidase, triglycerides, serum total cholesterol and body mass index, and a significant decrease in high density lipoprotein cholesterol were also observed in subjects with impaired glucose tolerance and diabetes mellitus. These results suggest that alcohol dependency, fatty liver, obesity and hyperlipidemia are important concomitants of impaired glucose tolerance.


Subject(s)
Diabetes Mellitus/etiology , Glucose Tolerance Test , Adult , Aged , Alcoholism/complications , Blood Proteins/analysis , Cross-Sectional Studies , Diabetes Mellitus/blood , Humans , Lipids/blood , Middle Aged , Risk Factors
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