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1.
Tissue Antigens ; 82(1): 68-9, 2013 Jul.
Article in English | MEDLINE | ID: mdl-23745573

ABSTRACT

HLA-C*08:75 differs from C*08:02:01 by a non-synonymous mutation at codon 229 (GAG to AAG) in exon 4.


Subject(s)
Alleles , HLA-C Antigens/genetics , White People/genetics , Base Sequence , Exons/genetics , Humans , Molecular Sequence Data , Sequence Alignment
2.
Tissue Antigens ; 82(1): 69-70, 2013 Jul.
Article in English | MEDLINE | ID: mdl-23517135

ABSTRACT

HLA-C*08:76 differs from HLA-C*08:02:01 by one nonsynonymous nucleotide change at the codon 144 (CAG to AAG) in exon 3.


Subject(s)
Alleles , HLA-C Antigens/genetics , White People/genetics , Base Sequence , Exons/genetics , Humans , Molecular Sequence Data , Sequence Alignment
3.
J Biol Chem ; 276(27): 25030-6, 2001 Jul 06.
Article in English | MEDLINE | ID: mdl-11349131

ABSTRACT

In mammalian cells, CDK2 is part of a multiprotein complex that includes Cyclin A or E and cell cycle regulatory proteins such as p21(Cip1), PCNA, p27(Kip1), p45(SKP2), p19(SKP1), and CksHs1/CksHs2. While the role of some of these proteins has been well studied, the function of other proteins in the complex remains unclear. In this study, we showed that the carboxyl-terminal region of p45(SKP2) associates directly with CksHs1 and that CksHs1 negatively regulated the interaction between p45(SKP2) and CDK2. Moreover, we showed that overexpression of CksHs1 inhibits CDK2 kinase activity and that additional expression of p45(SKP2) overcame this inhibition and restored CDK2 kinase activity. We proposed that the association of CksHs1 and p45(SKP2) prevented CksHs1 from binding CDK2 and negatively regulating the CDK2 kinase activity.


Subject(s)
CDC2-CDC28 Kinases , Carrier Proteins/metabolism , Cell Cycle Proteins/metabolism , Cyclin-Dependent Kinases/metabolism , Protein Kinases , Protein Serine-Threonine Kinases/metabolism , Animals , COS Cells , Cyclin-Dependent Kinase 2 , Cyclin-Dependent Kinases/genetics , Electrophoresis, Polyacrylamide Gel , HeLa Cells , Humans , Macromolecular Substances , Multiprotein Complexes , Protein Binding , Protein Serine-Threonine Kinases/genetics , Rabbits , S-Phase Kinase-Associated Proteins , Transfection
4.
J Biol Chem ; 275(46): 36116-23, 2000 Nov 17.
Article in English | MEDLINE | ID: mdl-10948190

ABSTRACT

Trio is a complex protein containing two guanine nucleotide exchange factor domains each with associated pleckstrin homology domains, a serine/threonine kinase domain, two SH3 domains, an immunoglobulin-like domain, and spectrin-like repeats. Trio was originally identified as a LAR tyrosine phosphatase-binding protein and is involved in actin remodeling, cell migration, and cell growth. Herein we provide evidence that Trio not only activates RhoA but is also a RhoA target. The RhoA-binding site was mapped to the Trio immunoglobulin-like domain. RhoA isoprenylation is necessary for the RhoA-Trio interaction, because mutation of the RhoA carboxyl-terminal cysteine residue blocked binding. The existence of an intramolecular functional link between RhoA activation and RhoA binding is suggested by the finding that Trio exchange activity enhanced RhoA binding to Trio. Furthermore, immunofluorescence studies of HeLa cells showed that although ectopically expressed Trio was evenly distributed within the cell, co-expression of Trio with RhoA resulted in relocalization of Trio into punctate structures. Relocalization was not observed with Trio constructs lacking the immunoglobulin-like domain, indicating that RhoA acts to regulate Trio localization via binding to the immunoglobulin-like domain. We propose that Trio-mediated RhoA activation and subsequent RhoA-mediated relocalization of Trio functions to modulate and coordinate Trio signaling.


Subject(s)
Guanine Nucleotide Exchange Factors/chemistry , Guanine Nucleotide Exchange Factors/metabolism , Immunoglobulins/chemistry , Phosphoproteins/chemistry , Phosphoproteins/metabolism , Protein Serine-Threonine Kinases/chemistry , Protein Serine-Threonine Kinases/metabolism , rhoA GTP-Binding Protein/metabolism , Animals , Binding Sites , COS Cells , Cysteine/metabolism , Fluorescent Antibody Technique , Guanine Nucleotide Exchange Factors/genetics , HeLa Cells , Humans , Microscopy, Fluorescence , Mutation , Phosphoproteins/genetics , Protein Binding , Protein Prenylation , Protein Serine-Threonine Kinases/genetics , Protein Structure, Tertiary , Protein Transport , Recombinant Fusion Proteins , Transfection , rhoA GTP-Binding Protein/chemistry
5.
Hematol Cell Ther ; 41(4): 153-61, 1999 Aug.
Article in English | MEDLINE | ID: mdl-10543370

ABSTRACT

Several mutations prevent the expression of p53 in the human lymphoblastoid T cell line Jurkat. Restoration of p53 in Jurkat cells had no effect on the cell growth but markedly increased the amount of apoptosis induced by gamma-irradiation. Inhibition of RNA synthesis using 5,6-dichlorobenimidizole riboside had little effect on apoptosis induced by irradiation in the presence of p53 and did not affect the p53-independent apoptotic pathway. Expression of p53 also had no effect on the expression levels of proteins such as Fas, GADD45, Bax, Bcl-2, Bcl-x(L) or p53 induced proteins (PIGS) in resting cells or after irradiation. Activation of protein kinase C by phorbol 12-myristate 13-acetate produced an almost complete inhibition of p53-independent apoptosis following irradiation, whereas no significant effect was observed on the rate of p53-induced apoptosis. Although phorbol 12-myristate 13-acetate strongly induced p21 and stabilised p53 in the resting transfected Jurkat cells, neither apoptosis nor cell arrest was observed. In summary, this work shows that p53 enhances the radiosensitivity of Jurkat cells through an apoptotic process that is triggered by irradiation and is largely independent of RNA synthesis and protein kinase C activation. Apoptosis in p53- negative Jurkat cells is strongly inhibited by PMA indicating that the pathway triggered by p53 may be distinct from apoptotic pathways used in its absence.


Subject(s)
Apoptosis/drug effects , Apoptosis/radiation effects , Jurkat Cells/drug effects , Jurkat Cells/physiology , Tetradecanoylphorbol Acetate/pharmacology , Tumor Suppressor Protein p53/pharmacology , Apoptosis/genetics , Gamma Rays , Gene Expression Regulation/drug effects , Humans , Protein Kinase C , Time Factors , Transcription, Genetic , Transfection , Tumor Suppressor Protein p53/drug effects , Tumor Suppressor Protein p53/genetics
6.
J Cell Sci ; 112 ( Pt 12): 1825-34, 1999 Jun.
Article in English | MEDLINE | ID: mdl-10341202

ABSTRACT

Rho family GTPases regulate diverse cellular processes, including extracellular signal-mediated actin cytoskeleton reorganization and cell growth. The functions of GTPases are positively regulated by guanine nucleotide exchange factors, which promote the exchange of GDP for GTP. Trio is a complex protein possessing two guanine nucleotide exchange factor domains, each with adjacent pleckstrin homology and SH3 domains, a protein serine/threonine kinase domain with an adjacent immunoglobulin-like domain and multiple spectrin-like domains. To assess the functional role of the two Trio guanine nucleotide exchange factor domains, NIH 3T3 cell lines stably expressing the individual guanine nucleotide exchange factor domains were established and characterized. Expression of the amino-terminal guanine nucleotide exchange factor domain results in prominent membrane ruffling, whereas cells expressing the carboxy-terminal guanine nucleotide exchange factor domain have lamellae that terminate in miniruffles. Moreover, cells expressing the amino-terminal guanine nucleotide exchange factor domain display more rapid cell spreading, haptotactic cell migration and anchorage-independent growth, suggesting that Trio regulates both cell motility and cell growth. Expression of full-length Trio in COS cells also alters actin cytoskeleton organization, as well as the distribution of focal contact sites. These findings support a role for Trio as a multifunctional protein that integrates and amplifies signals involved in coordinating actin remodeling, which is necessary for cell migration and growth.


Subject(s)
Actins/ultrastructure , Cytoskeleton/ultrastructure , Protein Structure, Tertiary , Proteins/chemistry , 3T3 Cells , Animals , COS Cells , Cell Adhesion/physiology , Cell Division/physiology , Cell Movement/physiology , Guanine Nucleotide Exchange Factors , Mice
7.
Leuk Lymphoma ; 35(3-4): 237-43, 1999 Oct.
Article in English | MEDLINE | ID: mdl-10706446

ABSTRACT

Protein tyrosine phosphatases play an essential role in the control of leucocyte cell growth an differentiation. Recently a new receptor type membrane tyrosine phosphatase named CD148 has been identified. This molecule is present on the membrane of all the hematopoietic lineages as well as on several other cell types, mainly epithelial cells and its expression increases after cell activation. This molecule is able to act as a transducing molecule. Moreover, CD148 is able to modulate the signal transduction through the TCR/CD3 complex, in a manner similar to CD45. It has also been suggested that CD148 could be involved in mechanisms of differentiation and inhibition of cell growth. In addition, CD148 seems to be associated with a serine/threonine kinase in certain epithelial cell lines and leucocytes. Here, we review recent data on the expression and function of CD148 in both human, mouse and rat.


Subject(s)
Leukocytes/physiology , Protein Tyrosine Phosphatases/physiology , Signal Transduction , Animals , Cell Differentiation/physiology , Humans , Leukocytes/cytology , Mice , Rats , Receptor-Like Protein Tyrosine Phosphatases, Class 3
8.
J Biol Chem ; 273(25): 15611-20, 1998 Jun 19.
Article in English | MEDLINE | ID: mdl-9624153

ABSTRACT

LAR family transmembrane protein-tyrosine phosphatases function in axon guidance and mammary gland development. In cultured cells, LAR binds to the intracellular, coiled coil LAR-interacting protein at discrete ends of focal adhesions, implicating these proteins in the regulation of cell-matrix interactions. We describe seven LAR-interacting protein-like genes in humans and Caenorhabditis elegans that form the liprin gene family. Based on sequence similarities and binding characteristics, liprins are subdivided into alpha-type and beta-type liprins. The C-terminal, non-coiled coil regions of alpha-liprins bind to the membrane-distal phosphatase domains of LAR family members, as well as to the C-terminal, non-coiled coil region of beta-liprins. Both alpha- and beta-liprins homodimerize via their N-terminal, coiled coil regions. Liprins are thus multivalent proteins that potentially form complex structures. Some liprins have broad mRNA tissue distributions, whereas others are predominately expressed in the brain. Co-expression studies indicate that liprin-alpha2 alters LAR cellular localization and induces LAR clustering. We propose that liprins function to localize LAR family tyrosine phosphatases at specific sites on the plasma membrane, possibly regulating their interaction with the extracellular environment and their association with substrates.


Subject(s)
Caenorhabditis elegans Proteins , Cell Adhesion Molecules/genetics , Multigene Family , Phosphoproteins/genetics , Receptors, Cell Surface , Adaptor Proteins, Signal Transducing , Amino Acid Sequence , Animals , COS Cells , Caenorhabditis elegans , Cell Adhesion Molecules/chemistry , Evolution, Molecular , Humans , Intercellular Signaling Peptides and Proteins , Molecular Sequence Data , Phosphoproteins/chemistry , Protein Tyrosine Phosphatases/metabolism , RNA, Messenger/metabolism , Receptor-Like Protein Tyrosine Phosphatases, Class 4
9.
Proc Natl Acad Sci U S A ; 93(11): 5466-71, 1996 May 28.
Article in English | MEDLINE | ID: mdl-8643598

ABSTRACT

rho-like GTP binding proteins play an essential role in regulating cell growth and actin polymerization. These molecular switches are positively regulated by guanine nucleotide exchange factors (GEFs) that promote the exchange of GDP for GTP. Using the interaction-trap assay to identify candidate proteins that bind the cytoplasmic region of the LAR transmembrane protein tyrosine phosphatase (PT-Pase), we isolated a cDNA encoding a 2861-amino acid protein termed Trio that contains three enzyme domains: two functional GEF domains and a protein serine/threonine kinase (PSK) domain. One of the Trio GEF domains (Trio GEF-D1) has rac-specific GEF activity, while the other Trio GEF domain (Trio GEF-D2) has rho-specific activity. The C-terminal PSK domain is adjacent to an Ig-like domain and is most similar to calcium/calmodulin-dependent kinases, such as smooth muscle myosin light chain kinase which similarly contains associated Ig-like domains. Near the N terminus, Trio has four spectrin-like repeats that may play a role in intracellular targeting. Northern blot analysis indicates that Trio has a broad tissue distribution. Trio appears to be phosphorylated only on serine residues, suggesting that Trio is not a LAR substrate, but rather that it forms a complex with LAR. As the LAR PTPase localizes to the ends of focal adhesions, we propose that LAR and the Trio GEF/PSK may orchestrate cell-matrix and cytoskeletal rearrangements necessary for cell migration.


Subject(s)
GTP-Binding Proteins/chemistry , Guanine Nucleotide Exchange Factors , Phosphoproteins/chemistry , Phosphoproteins/metabolism , Protein Serine-Threonine Kinases/chemistry , Protein Serine-Threonine Kinases/metabolism , Protein Tyrosine Phosphatases/metabolism , Amino Acid Sequence , Animals , Binding Sites , Brain/metabolism , Cell Line , Cloning, Molecular , Consensus Sequence , DNA, Complementary , GTP-Binding Proteins/metabolism , Gene Expression , Humans , Molecular Sequence Data , Myocardium/metabolism , Phosphoproteins/biosynthesis , Protein Kinases/chemistry , Protein Serine-Threonine Kinases/biosynthesis , Receptor-Like Protein Tyrosine Phosphatases, Class 4 , Receptors, Cell Surface/metabolism , Recombinant Proteins/biosynthesis , Recombinant Proteins/chemistry , Recombinant Proteins/metabolism , Sequence Homology, Amino Acid , Transfection , Tumor Cells, Cultured
10.
Proc Natl Acad Sci U S A ; 92(25): 11686-90, 1995 Dec 05.
Article in English | MEDLINE | ID: mdl-8524829

ABSTRACT

The transmembrane protein-tyrosine-phosphatases (PTPases) LAR, PTP delta, and PTP sigma each contain two intracellular PTPase domains and an extracellular region consisting of Ig-like and fibronectin type III-like domains. We describe the cloning and characterization of human PTP sigma (HPTP sigma) and compare the structure, alternative splicing, tissue distribution, and PTPase activity of LAR, HPTP delta, and HPTP sigma, as well their ability to associate with the intracellular coiled-coil LAR-interacting protein LIP.1. Overall, these three PTPases are structurally very similar, sharing 64% amino acid identity. Multiple isoforms of LAR, HPTP delta, and HPTP sigma appear to be generated by tissue-specific alternative splicing of up to four mini-exon segments that encode peptides of 4-16 aa located in both the extracellular and intracellular regions. Alternative usage of these peptides varies depending on the tissue mRNA analyzed. Short isoforms of both HPTP sigma and HPTP delta were also detected that contain only four of the eight fibronectin type III-like domains. Northern blot analysis indicates that LAR and HPTP sigma are broadly distributed whereas HPTP delta expression is largely restricted to brain, as is the short HPTP sigma isoform containing only four fibronectin type III-like domains. LAR, HPTP delta, and HPTP sigma exhibit similar in vitro PTPase activities and all three interact with LIP.1, which has been postulated to recruit LAR to focal adhesions. Thus, these closely related PTPases may perform similar functions in various tissues.


Subject(s)
Isoenzymes/metabolism , Membrane Proteins/metabolism , Phosphoproteins/metabolism , Protein Tyrosine Phosphatases/metabolism , Receptors, Cell Surface , Adaptor Proteins, Signal Transducing , Amino Acid Sequence , Base Sequence , Blotting, Northern , Humans , Molecular Sequence Data , Protein Binding , Protein Conformation , RNA, Messenger/analysis , Receptor-Like Protein Tyrosine Phosphatases, Class 2 , Receptor-Like Protein Tyrosine Phosphatases, Class 4 , Sequence Homology, Amino Acid , Tissue Distribution
11.
EMBO J ; 14(12): 2827-38, 1995 Jun 15.
Article in English | MEDLINE | ID: mdl-7796809

ABSTRACT

Focal adhesions are sites of cell-extracellular matrix interactions that function in anchoring stress fibers to the plasma membrane and in adhesion-mediated signal transduction. Both focal adhesion structure and signaling ability involve protein tyrosine phosphorylation. LAR is a broadly expressed transmembrane protein tyrosine phosphatase comprised of a cell adhesion-like ectodomain and two intracellular protein tyrosine phosphatase domains. We have identified a novel cytoplasmic 160 kDa phosphoserine protein termed LAR-interacting protein 1 (LIP.1), which binds to the LAR membrane-distal D2 protein tyrosine phosphatase domain and appears to localize LAR to focal adhesions. Both LAR and LIP.1 decorate the ends of focal adhesions most proximal to the cell nucleus and are excluded from the distal ends of focal adhesions, thus localizing to regions of focal adhesions presumably undergoing disassembly. We propose that LAR and LIP.1 may regulate the disassembly of focal adhesions and thus help orchestrate cell-matrix interactions.


Subject(s)
Cell Adhesion/physiology , Phosphoproteins/metabolism , Protein Tyrosine Phosphatases/metabolism , Receptors, Cell Surface , Adaptor Proteins, Signal Transducing , Amino Acid Sequence , Cloning, Molecular , Cytoplasm/metabolism , Extracellular Matrix/metabolism , Humans , Molecular Sequence Data , Molecular Weight , Organ Specificity , Phosphoproteins/chemistry , Phosphoproteins/genetics , Protein Conformation , Protein Structure, Secondary , RNA, Messenger/analysis , Receptor-Like Protein Tyrosine Phosphatases, Class 4 , Recombinant Fusion Proteins/biosynthesis , Recombinant Fusion Proteins/metabolism , Sequence Analysis, DNA
12.
J Biol Chem ; 270(12): 6722-8, 1995 Mar 24.
Article in English | MEDLINE | ID: mdl-7896816

ABSTRACT

Protein-tyrosine phosphatases (PTPases) play an essential role in the regulation of cell activation, proliferation, and differentiation. A major subfamily of these enzymes is the transmembrane-type PTPases that contain extracellular regions comprised of Ig-like and fibronectin type III (FN-III)-like domains. Characterization of the human transmembrane PTPase delta (HPTP delta) revealed the existence of multiple HPTP delta isoforms that vary in their extracellular regions. The full-length HPTP delta isoform has an extracellular region containing three Ig-like and eight FN-III-like domains connected via a transmembrane peptide to an intracellular region with two PTPase domains, whereas another isoform lacks four of the eight FN-III like domains. Furthermore, other HPTP delta isoforms exist that lack 9 amino acids within the second Ig-like domain and 4 amino acids at the junction of the second and third Ig-like domains or 9 amino acids within the fifth FN-III-like domain. Reverse transcription polymerase chain reaction analysis demonstrated that HPTP delta isoforms lacking these short peptides are expressed in kidney, whereas isoforms containing these peptides are expressed in the brain. Analysis of HPTP delta biosynthesis demonstrated that HPTP delta is expressed as a complex of two noncovalently associated subunits derived from a proprotein and that the HPTP delta ectodomain is shed from the cell surface. Mutational analysis of the HPTP delta proprotein cleavage site revealed the existence of two or three functional and overlapping furin-like endoprotease cleavage sites.


Subject(s)
Isoenzymes/genetics , Protein Tyrosine Phosphatases/genetics , Alternative Splicing , Amino Acid Sequence , Base Sequence , DNA, Complementary/isolation & purification , Enzyme Precursors/metabolism , Humans , Molecular Sequence Data , Organ Specificity , Protein Tyrosine Phosphatases/biosynthesis , Protein Tyrosine Phosphatases/chemistry , RNA, Messenger/analysis
13.
J Biol Chem ; 269(38): 23632-41, 1994 Sep 23.
Article in English | MEDLINE | ID: mdl-8089133

ABSTRACT

The LAR transmembrane protein tyrosine phosphatase (PTPase) is expressed on the cell surface as a complex of two noncovalently associated subunits derived from a proprotein. The 150-kDa E-subunit contains most of the extracellular region, including the immunoglobulin-like and fibronectin type-III-like domains, whereas the 85-kDa P-subunit contains a short ectodomain, the transmembrane peptide, and the two intracellular PTPase domains. The LAR extracellular region is released from the cell surface, suggesting that shedding may be a mechanism to regulate LAR PTPase function. Functional regions necessary for LAR proprotein processing, subunit association, and shedding were determined by analyzing the effect of amino acid substitutions of residues surrounding the cleavage site and scanning the P-subunit ectodomain. Three amino acid residues were identified, two within a penta-arginine sequence and one C-terminal to the cleavage site, that are essential for efficient LAR proprotein cleavage. Several noncontiguous amino acid residues were also identified that play an essential role in LAR subunit association. LAR shedding is shown to be a consequence of proteolytic cleavage at a second site within the P-subunit ectodomain near the transmembrane peptide.


Subject(s)
Membrane Proteins/genetics , Protein Tyrosine Phosphatases/genetics , Amino Acid Sequence , Consensus Sequence , Extracellular Space/chemistry , Humans , Membrane Proteins/metabolism , Molecular Sequence Data , Mutagenesis, Site-Directed , Phorbol Esters/pharmacology , Protein Precursors/metabolism , Protein Processing, Post-Translational , Protein Tyrosine Phosphatases/metabolism , Sequence Alignment , Sequence Homology, Amino Acid , Structure-Activity Relationship
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