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M Jannatipour

Publications and source records attributed to M Jannatipour.

At least 19 recordsLinked to original sources
PubMed2001

Schwannomin isoform-1 interacts with syntenin via PDZ domains.

The neurofibromatosis type 2 gene (NF2) is involved in the pathogenesis of benign tumors of the human nervous system. The NF2 protein, called schwannomin or merlin, is inactivated in virtually all schwannomas and meningiomas. The molecular mechanisms by which schwannomin functions as a tumor suppressor is unknown but believed to involve plasma membrane-cytoskeletal interactions. Two major alternatively spliced isoforms of schwannomin differing in their C termini have been reported. Using the yeast two-hybrid system, we have identified syntenin as a binding partner for schwannomin isoform-1 (sch-1). Syntenin is an adapter protein that couples transmembrane proteoglycans to cytoskeletal components and is involved in intracellular vesicle transport. The C terminus 25 amino acids of sch-1 and the two PDZ domains of syntenin mediate their binding, and mutations introduced within the VAFFEEL region of sch-1 defined a sequence crucial for syntenin recognition. We have showed that the two proteins interacted in vitro and in vivo and localized underneath the plasma membrane. Fibroblast cells expressing heterologous antisense syntenin display alterations in the subcellular distribution of sch-1. Together, these results provide the first functional clue to the existence of schwannomin isoforms and could unravel novel pathways for the transport and subcellular localization of schwannomin in vivo.

3T3 Cells↗
PubMed2000

CAG tract of MJD-1 may be prone to frameshifts causing polyalanine accumulation.

Machado-Joseph disease (MJD) is one of several disorders caused by the expansion of a coding CAG repeat (exp-CAG). The presence of intranuclear inclusions (INIs) in patients and cellular models of exp-CAG-associated diseases has lead to a nuclear toxicity model. Similar INIs are found in oculopharyngeal muscular dystrophy, which is caused by a short expansion of an alanine-encoding GCG repeat. Here we propose that transcriptional or translational frameshifts occurring within expanded CAG tracts result in the production and accumulation of polyalanine-containing mutant proteins. We hypothesize that these alanine polymers deposit in cells forming INIs and may contribute to nuclear toxicity. We show evidence that supports our hypothesis in lymphoblast cells from MJD patients, as well as in pontine neurons of MJD brain and in in vitro cell culture models of the disease. We also provide evidence that alanine polymers alone are harmful to cells and predict that a similar pathogenic mechanism may occur in the other CAG repeat disorders.

Amino Acid Sequence↗
PubMed1999

Polyglutamine-containing proteins in schizophrenia.

Genetic anticipation, manifested by increased severity and earlier age-at-onset of the disease over successive generations, is reported in schizophrenia. The molecular basis of anticipation in several neurodegenerative diseases is unstable coding CAG repeat expansions. Anticipation was reported in schizophrenia. Recently, studies suggested that enlarged CAG/CTG repeats are over represented in schizophrenic patients compared to normal controls. Together, these observations suggest that unstable CAG repeats may play a role in the etiology of schizophrenia. The purpose of this study is to test for the presence of polyglutamine-expanded tracts, encoded by CAG repeats, in total protein extracts derived from lymphoblastoid cell lines of schizophrenic patients. Proteins from schizophrenic patients (n = 59) and normal controls (n = 73) were separated by means of SDS-polyacrylamide gel electrophoresis, wet blotted onto nitrocellulose membrane and probed with a monoclonal antibody (mab 1C2) recognizing expanded polyglutamine arrays. Three abnormal bands corresponding to protein(s) of molecular weight of approximately 50 kDa were identified in two unrelated schizophrenic patients and in a sibling of one of these patients. None of the normal controls tested positive for this abnormal band. These results suggest that expanded polyglutamine-containing proteins, though rare, may play a role in the pathogenesis of schizophrenia.

Age of Onset↗
PubMed1998

Calnexin and BiP interact with acid phosphatase independently of glucose trimming and reglucosylation in Schizosaccharomyces pombe.

The association of newly synthesized glycoproteins with the ER molecular chaperones calnexin and immunoglobulin binding protein (BiP) has been well documented in a variety of higher eukaryotes. Here we report that Cnx1p, the calnexin homologue in Schizosaccharomyces pombe, associates with newly synthesized molecules of the secreted glycoprotein acid phosphatase. Unlike ligand binding to mammalian calnexin, glucose trimming and reglucosylation of acid phosphatase by UDP-Glc:glycoprotein glucosyltransferase were shown to be dispensable for its binding to Cnx1p. Thus, despite the essentiality of Cnx1p for S. pombe viability, the glucose trimming and reglucosylation cycle does not appear to be required for protein folding in the fission yeast. The association of core-glycosylated acid phosphatase with Cnx1p after exposure of cells to heat shock or to DTT was shown to be reversible. However, Cnx1p stably associated with unglycosylated acid phosphatase after treatment with the core-glycosylation inhibitor tunicamycin. BiP was found to coprecipitate with Cnx1p, under normal and stress conditions, and following inhibition of protein synthesis by cycloheximide. We postulate that Cnx1p and BiP are part of a complex that is involved in the folding of both core-glycosylated trimmed ligands and unglycosylated proteins.

Acid Phosphatase↗
PubMed1998

A Schizosaccharomyces pombe gene encoding a novel polypeptide with a predicted alpha-helical rod structure found in the myosin and intermediate-filament families of proteins.

We have identified a Schizosaccharomyces pombe gene encoding a 461 amino acid polypeptide containing a predicted alpha-helical rod domain found in filamentous proteins. This gene, here designated noc1, is located immediately upstream from cnx1, the gene encoding the S. pombe homologue of mammalian calnexin, an endoplasmic reticulum chaperone [M. Jannatipour, L.A. Rokeach, J. Biol. Chem. 270 (1995) 4845-4853.]. Transcription of noc1 is divergent from that of cnx1. Northern blot analysis identified a single mRNA of approx. 2 kb whose expression was increased by heat shock and growth in the presence of beta-mercaptoethanol and 2-deoxyglucose.

Amino Acid Sequence↗
PubMed1996

A new stress protein: synthesis of Schizosaccharomyces pombe UDP--Glc:glycoprotein glucosyltransferase mRNA is induced by stress conditions but the enzyme is not essential for cell viability.

We have identified and begun the characterization of the gene encoding UDP-Glc:glycoprotein glucosyltransferase in Schizosaccharomyces pombe. This gene, here designated gpt1, codes for a polypeptide having a signal peptide of 18 amino acids followed by 1429 amino acids with no transmembrane domain, as expected for a soluble protein of the endoplasmic reticulum (ER). The C-terminal tetrapeptide PDEL most probably corresponds to a novel ER retention signal in this fission yeast. Synthesis of the corresponding mRNA was induced 2- to 9-fold by conditions known to affect glycoprotein folding in the ER (e.g. heat shock, culture in the presence of a Ca2+ionophore, 2-mercaptoethanol or inhibitors of protein N-glycosylation such as tunicamycin or 2-deoxyglucose). This is the first evidence obtained in vivo that supports the proposed involvement of the enzyme in the quality control of glycoprotein folding in the ER. Thus far, the said involvement was inferred solely from the ability of the enzyme to glucosylate misfolded but not native glycoproteins in cell-free assays. The gpt1 gene was disrupted and gpt1- cells were found to be viable. Moreover, no significant differences in the growth rate patterns at 18, 28 or 39 degrees C or in cell morphology between gpt1+ and gpt1- cells were observed, although they differed slightly in size.

Amino Acid Sequence↗
PubMed1996

Functional expression of human topoisomerase II alpha in yeast: mutations at amino acids 450 or 803 of topoisomerase II alpha result in enzymes that can confer resistance to anti-topoisomerase II agents.

DNA topoisomerase II is the target of a variety of important antitumor agents, including etoposide, adriamycin, and amsacrine. We have constructed a system for analyzing the action of anti-topoisomerase II agents using the yeast Saccharomyces cerevisiae and have constructed vectors for expressing human topoisomerase II functionally in yeast. We have demonstrated that temperature-conditional yeast TOP2 mutants can be complemented by expression of wild-type human topoisomerase II alpha. Furthermore, expression of human topoisomerase II in yeast results in a quantitatively unique pattern of sensitivity to amsacrine. We also have constructed mutations in human TOP2 based on previously identified mutations from a human cell line selected for resistance to teniposide. Our experiments demonstrate that mutation of either arginine 450 or proline 803 of human topoisomerase II can result in an enzyme that has altered sensitivity to anti-topoisomerase II agents, and that a human enzyme carrying both mutations confers a higher level of drug resistance than enzymes carrying either single mutation.

Alleles↗
PubMed1995

The Caenorhabditis elegans rop-1 gene encodes the homologue of the human 60-kDa Ro autoantigen.

As a first step toward establishing a genetic system for the elucidation of the cellular role(s) of the Ro ribonucleoproteins (RoRNP), we have cloned the gene encoding the homologue of the human 60-kDa Ro protein (Ro60) in Caenorhabditis elegans (Ce). This Ce gene is present as a single copy and contains a 643-codon open reading frame interrupted by three introns. The encoded protein, Rop1p, shares 40% identity and 63% overall similarity with both the human and amphibian Ro60. Recombinant protein has been produced in Escherichia coli and used to elicit anti-Rop1p antibodies. Immunological analysis indicated that the Ro60 epitopes have been poorly conserved. Gene-fusion expression studies in transgenic nematodes will provide a new avenue of research to shed light on the function of these particles.

Amino Acid Sequence↗
PubMed1995

The Schizosaccharomyces pombe homologue of the chaperone calnexin is essential for viability.

We have cloned a Schizosaccharomyces pombe gene, here designated cnx1, encoding the homologue of the endoplasmic reticulum molecular chaperone calnexin. Disruption of the cnx1 gene was lethal, demonstrating that it has an essential cellular function. Transcription of cnx1 mRNA is initiated at multiple sites, and it can be induced by various stress treatments that lead to the accumulation of unfolded and/or misfolded proteins in the endoplasmic reticulum. The encoded Cnx1p protein more closely resembles its plant and animal calnexin homologues than that of Saccharomyces cerevisiae. Cnx1p is acidic and migrates aberrantly on SDS-polyacrylamide gel electrophoresis, similar to its mammalian counterparts. Cnx1p contains the hallmark KPEDWD motifs that are found in all members of the calnexin/calreticulin family of proteins. Using an in vitro translation-processing system, we have shown that Cnx1p has the characteristic type I topology of calnexin proteins. Unlike its higher eukaryotic homologues, Cnx1p has a site for N-glycosylation that was modified in an in vitro translation-processing assay.

Amino Acid Sequence↗
PubMed1994

Yeast topoisomerase II mutants resistant to anti-topoisomerase agents: identification and characterization of new yeast topoisomerase II mutants selected for resistance to etoposide.

We describe a system that allows us to easily isolate and characterize mutants in yeast topoisomerase II that are resistant to antitumor agents that target this enzyme. The system uses yeast strains that are sensitive to those agents and that carry temperature-sensitive top2 mutations. The temperature-sensitive mutation allows the isolation of recessive drug-resistant mutations. The mutagenized TOP2 gene we have used is under the control of the yeast DED1 promoter; this overexpression of TOP2 is designed to avoid isolating mutants that are drug resistant solely because the mutated topoisomerase II has low enzymatic activity. We describe three mutants that we isolated using this system. Two of the three mutants show resistance to etoposide and amsacrine, while the third mutant is partially resistant to etoposide and fluoroquinolones but not to amsacrine. DNA sequence changes have been identified in all of these mutant TOP2 genes. The mutant with partial resistance to etoposide and fluoroquinolones has an amino acid change at position 738 of TOP2, which is three amino acids from the site homologous to Ser83 of E. coli gyrA, an amino acid which had previously been shown to be an important target for resistance to quinolones in bacteria. One of the alleles that confers resistance to both etoposide and amsacrine, top2-103, has changes in amino acid 824 and amino acid 1186 of TOP2. Reconstruction of the mutations by oligonucleotide-directed mutagenesis demonstrates that the change at amino acid 824 is responsible for the drug resistance of this allele.

Amino Acid Sequence↗
PubMed1993

Mouse cDNAs encoding a trifunctional protein of de novo purine synthesis and a related single-domain glycinamide ribonucleotide synthetase.

Three of the enzymatic activities of de novo purine synthesis, glycinamide ribonucleotide synthetase (GARS), aminoimidazole ribonucleotide synthetase (AIRS) and glycinamide ribonucleotide formyltransferase (GART), can be catalyzed by a single 110-kDa protein in mouse cells. Western blots using a polyclonal antibody (Ab) to this protein identified two species, the trifunctional 110-kDa protein and a 50-kDa cytosolic protein with GARS, but not GART activity. We used Ab and, subsequently, oligodeoxyribonucleotide screens to isolate cDNAs corresponding to these two proteins from mouse T-cell cDNA expression libraries. The sequence of one class of these cDNAs and the partial sequence of a corresponding genomic clone defined an open reading frame (ORF) encoding a 1010-amino-acid (aa) protein, individual domains of which showed high homology to each of the monofunctional bacterial GARS, AIRS and GART proteins, and to each domain of chicken and human trifunctional GARS-AIRS-GARTs. cDNAs corresponding to the smaller protein contained a 1.3-kb ORF with complete identity to the GARS domain of, but with a 3' untranslated region different from, the trifunctional cDNAs. Hence, both cDNAs appear to derive from the same gene due to either differential splicing or use of an intronic polyadenylation signal. The functional requirement for the expression of both trifunctional protein with GARS activity and monofunctional, catalytically active GARS is unknown.

Acyltransferases↗
PubMed1993

The top2-5 mutant of yeast topoisomerase II encodes an enzyme resistant to etoposide and amsacrine.

We have characterized a temperature-sensitive mutant in the yeast TOP2 gene that shows resistance to the anti-topoisomerase II agents amsacrine and etoposide. Cells carrying the top2-5 mutant have a minimum lethal concentration of amsacrine of greater than 100 micrograms/ml, compared to 10 micrograms/ml in isogenic wild-type cells, and a minimum lethal concentration of greater than 100 micrograms/ml etoposide, compared with 50 micrograms/ml for cells carrying wild-type topoisomerase II. We have cloned the top2-5 allele into a yeast vector that allows high level overexpression of the protein. As expected, the purified top2-5 activity is temperature-sensitive. The protein shows a 3-fold reduction of amsacrine-stabilized cleavage at the permissive temperature, confirming that the top2-5 protein is resistant to amsacrine in vitro. The protein also exhibits reduced cleavage in the presence of etoposide, with the largest effect at low concentrations of the drug. These results suggest that the top2-5 protein is altered in its sensitivity to anti-topoisomerase II agents. The relevant portion of the mutant allele has been sequenced, and several tightly clustered mutations have been identified. The location of the mutations identify a domain of the topoisomerase II protein that may be important in the interaction of the protein with anti-topoisomerase II anti-cancer drugs.

Alleles↗
PubMed1992

A dideazatetrahydrofolate analogue lacking a chiral center at C-6, N-[4-[2-(2-amino-3,4-dihydro-4-oxo-7H-pyrrolo[2,3-d]pyrimidin-5- yl)ethyl]benzoyl]-L-glutamic acid, is an inhibitor of thymidylate synthase.

N-[4-[2-(2-Amino-3,4-dihydro-4-oxo-7H-pyrrolo[2,3-d]pyrimidin-5- yl)ethyl]benzoyl]-L-glutamic acid (15), prepared in five steps from 2-pivaloyl-7-deazaguanine, has been found to be an antitumor agent with its primary site of action at thymidylate synthase rather than purine synthesis. This compound appears to be a promising candidate for clinical evaluation.

Animals↗
PubMed1992

Amsacrine and etoposide hypersensitivity of yeast cells overexpressing DNA topoisomerase II.

Increasing the cellular concentration of DNA topoisomerase II in yeast by expressing constitutively a plasmid-borne TOP2 gene encoding the enzyme greatly increases the sensitivity of the cells to amsacrine and etoposide (VP-16). This increased drug sensitivity at a higher intracellular DNA topoisomerase II level is observed in both RAD52+ repair-proficient strains and rad52 mutants that are defective in the repair of double-stranded breaks. These results provide strong support of the hypothesis that the cellular target of these drugs is DNA topoisomerase II, and that these drugs kill cells by converting DNA topoisomerase II into a DNA damaging agent.

Amsacrine↗
PubMed1992

Mapping of the immunoreactive domains of a small nuclear ribonucleoprotein-associated Sm-D autoantigen.

The Sm-D(D1) small nuclear ribonucleoprotein (snRNP) polypeptide is a major target of autoantibodies diagnostic for systemic lupus erythematosus. The cDNA encoding the protein from Raji cells was expressed in Escherichia coli as a fusion protein with anthranilate synthase (TrpE-Sm-D). When tested by protein blot, the recombinant polypeptide was strongly immunoreactive under defined blotting conditions, which appear to facilitate the refolding of the polypeptide into a native conformation. Multiple translational fusions between the trpE gene and fragments encompassing the length of the Sm-D coding sequence were constructed for epitope mapping. The results describe two general patterns of anti-Sm reactivity: (i) antibodies that recognize only the full-length antigen and are presumably directed against discontinuous epitopes, and (ii) antibodies that recognize the carboxy terminus of the antigen which embodies an extended/charged structure.

Amino Acid Sequence↗
PubMed1990

Heterologous expression and epitope mapping of a human small nuclear ribonucleoprotein-associated Sm-B'/B autoantigen.

The U snRNP associated B'/B polypeptides are primary targets of Sm autoantibodies in patients with systemic lupus erythematosus. We have bacterially expressed a Sm-B'/B autoantigen from Raji cells as a fusion with the anthranilate synthase protein from Escherichia coli. The recombinant Sm-B'/B fusion displays comparable immunologic reactivity to the native protein when tested with both monoclonal and polyclonal antibodies. To map Sm-B'/B epitopes, we constructed a series of 12 anthranilate synthase fusions spanning different regions of Sm-B'/B and tested such fusions on immunoblots against a panel of characterized sera. In this manner, we have identified six epitopes, five of which overlap the proline-rich carboxyl-terminus of the protein. Some of these epitopes appear to be conformational. The human sera tested can be divided, according to the epitopes they recognize, into six groups. Finally, we have shown that anti-Sm recognition of the (U1)RNP-specific A protein is attributable to cross-reactivity between the Sm-B'/B and A autoantigens.

Amino Acid Sequence↗
PubMed1989

Primary structure of a human small nuclear ribonucleoprotein polypeptide as deduced by cDNA analysis.

Anti-Sm is an antibody specificity often associated with the autoimmune disease systemic lupus erythematosus. The polypeptides Sm-B'/B (estimated molecular mass 27 and 26 kDa, respectively) are primary targets of Sm antibodies. Sm-B'/B are part of the core polypeptides of small ribonucleoprotein particles (snRNP) involved in pre-mRNA splicing. Sm-B'/B share the same amino-terminal sequence as we determined by microsequence analyses of the purified polypeptides. Oligonucleotide probes based on that sequence were used to isolate seven clones from a human lymphoblastoid cDNA library in lambda gt10. The clones contained a single coding region for a protein of approximately 25 kDa. The predicted amino-terminal sequence was identical to that of the isolated Sm-B'/B polypeptides. In vitro translation experiments produced a protein immunoreactive with human polyclonal anti-Sm antibodies. The isolation of only one unique cDNA sequence suggests that Sm-B'/B may be post-translational variants encoded by a single message. The specific structural features which distinguish Sm-B' from Sm-B have yet to be determined. Northern blot analysis confirmed the diverse tissue and species distribution expected for these immunologically conserved polypeptides. The Sm-B'/B primary sequence is rich in proline (20%) and glycine (15%) residues. The prolines are concentrated in the carboxyl-terminal half of the protein and display a repetitive unit that is shared with other snRNP and nucleic acid binding proteins. Analysis of these arrays suggests an eight residue proline-rich consensus sequence with potential as either an RNA binding domain, or as a site of protein/protein interaction.

Amino Acid Sequence↗