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E Durban

Publications and source records attributed to E Durban.

At least 19 recordsLinked to original sources
PubMed1998

Nucleolar protein p120 contains an arginine-rich domain that binds to ribosomal RNA.

Human proliferation-associated protein p120 has previously been shown to localize to the nucleolus, and several functional domains of p120 have been elucidated. By using a nitrocellulose filter binding assay and a Northwestern blotting procedure this study shows that recombinant p120 binds to an rRNA fragment in vitro with a dissociation constant of 4 nM. The specific RNA-binding region of p120 (residues 1-57) was identified with glutathione S-transferase-fused p120 deletion constructs and Northwestern blotting procedures. This RNA-binding region of p120, which includes the nucleolar localization signal of p120, is similar to the arginine-rich RNA-binding regions found in other RNA-binding proteins such as HIV Rev and Tat. Experiments in vivo with HeLa cell nucleolar extracts showed that p120 was associated with the 60-80S pre-ribosomal particles. This association is disrupted by treatment with either RNase A or buffer of high ionic strength. These results suggest that p120 might be involved in rRNA/ribosome maturation, consistent with the role of the yeast homologue Nop2p in rRNA biogenesis.

Amino Acid Sequence↗
PubMed1996

Increased prevalence of systemic sclerosis in a Native American tribe in Oklahoma. Association with an Amerindian HLA haplotype.

OBJECTIVE: To investigate a high prevalence of systemic sclerosis (SSc; scleroderma) in a well-defined population of 21,255 Choctaw Indians residing in 8 southeastern Oklahoma counties who were "users" of Indian Health Services. METHODS: A case-control study of 12 SSc cases and 48 matched non-SSc controls (4 per case) was conducted to investigate potential occupational, residential, and infectious exposures, as well as genetic factors which might predispose to SSc. HLA class II alleles were determined by DNA oligotyping, and class I and III alleles were defined serologically. RESULTS: The prevalence of SSc in full-blooded Choctaws was at least 8/1,704, or 469/100,000 (95% confidence interval [95% CI] 203-930) over the 4-year interval 1990-1994 and was significantly higher than that among non-full-blooded Choctaws (6/19,551, or 31/100,000) (P = 0.00001, odds ratio [OR] = 15.4, 95% CI 4.9-49.8). The overall prevalence of SSc in Oklahoma Choctaws (66/100,000) also was significantly higher than that in other Native Americans in Oklahoma (9.5/100,000) (P = 10(-6), OR = 6.95, 95% CI 3.3-13.7), who showed a prevalence similar to that reported for whites (2.1-25.3/100,000). Among the SSc cases, there was striking homogeneity of disease expression with the majority exhibiting diffuse scleroderma, pulmonary fibrosis, and autoantibodies to topoisomerase I. No environmental exposures were found to be in excess among cases versus controls. The strongest risk factor for SSc in cases (100%) versus controls (54%) was an HLA haplotype bearing the alleles B35, Cw4, DRB1*1602 (DR2), DQA1*0501, and DQB1*0301 (DQ7) (P = 0.002, Pcorr = 0.036, OR = 21, 95% CI 2.9-437). Survey of another group of Choctaws residing in another state revealed no cases of SSc despite a high frequency of the same HLA haplotype. CONCLUSION: Full-blooded Choctaw Native Americans living in southeastern Oklahoma have the highest prevalence of SSc yet found in any population. A major risk factor for disease is a uniquely Amerindian HLA haplotype; however, additional genes and/or an as-yet-unidentified environmental exposure seem likely.

Adult↗
PubMed1996

Overexpression of human nucleolar proteins in insect cells: characterization of nucleolar protein p120.

Nucleolar p120 is a proliferation-associated protein, which becomes detectable early in the G1 phase of the cell cycle and peaks early in the S phase. A variety of human malignant tumor cells contain much higher levels of p120 than normal resting cells. The cellular functions of p120 are unknown, and little information is available on the structural characteristics of the human p120 protein. For biochemical characterization, human p120 protein was expressed in a baculovirus system and purified to approximately 95% purity. By indirect immunofluorescence, most of the recombinant human p120 as well as recombinant human B23, C23, or fibrillarin were localized to insect cell nucleoli and to large globular nuclear inclusions. Like endogenous p120 in HeLa cells, recombinant p120 expressed in insect cells was phosphorylated. On sucrose density gradients, p120 from HeLa cells sedimented in the 60-80S region, in which preribosomal particles sedimented using similar extraction and centrifugation procedures. The sedimentation of p120 shifted to the 5-10S region by treatment with 1 M KCl or with RNAse which suggests that p120 is bound to RNA.

Amino Acid Sequence↗
PubMed1995

Emergence of salivary gland cell lineage diversity suggests a role for androgen-independent epidermal growth factor receptor signaling.

Diversity of cell lineages within glandular organs is generated postnatally by differentiation of committed progenitor cells. Fundamental regulatory aspects of this process are not understood. The mouse submandibular salivary gland (SSG) served as model to assess the role of epidermal growth factor (EGF) receptor signaling during emergence of cell lineage diversity. Temporal fluctuations in EGF receptor mRNA levels coincident with crucial differentiative cell lineage transitions were revealed by RNase protection analyses. Between days 2 and 5, when proacinar cells are maturing and striated duct cells emerge, EGF receptor mRNA levels were highest and all differentiating cells exhibited EGF receptor immunoreactivity. EGF receptor mRNA levels then declined sharply and immunoreactivity became confined to ductal cells. At day 11 in male mice, and days 11 and 16 in females, a second increase in EGF receptor mRNA was detected coincident with emergence of granular convoluted tubule (GCT) cells. With completion of androgen-dependent GCT cell differentiation at the onset of puberty, EGF receptor mRNA levels and intensity of immunoreactivity decreased. Androgen effects on EGF receptor mRNA or immunoreactivity could not be detected. These temporally distinct patterns of EGF receptor expression suggest that this signaling pathway is a mechanism of potential importance in emergence of cell lineage diversity in a glandular organ.

Androgens↗
PubMed1995

Functional domains of nucleolar phosphoprotein p120.

Nucleolar phosphoprotein p120 is a low abundance, proliferation-associated protein. Several functional domains have been characterized and are discussed here such as the antigenic domain recognized by a monoclonal antibody, the nuclear/nucleolar localization domain, phosphorylation domains of casein kinase II (CKII) and protein kinase C, a putative methylation domain and an RNA binding region. By sucrose gradient sedimentation analyses, protein p120 was shown to rapidly sediment with 60-80 S pre-rRNP particles but sedimented more slowly when treated with RNAse or salt suggesting binding to RNA. Nucleolar protein p120 differed from other nucleolar proteins such as C23 (nucleolin) and B23 (nucleophosmin) which sedimented more slowly near the top of the gradient.

Amino Acid Sequence↗
PubMed1994

Human topoisomerase I is phosphorylated in vitro on its amino terminal domain by protein kinase NII.

Topoisomerase I purified from HeLa cells was phosphorylated in vitro with protein kinase NII (pkNII) purified from calf thymus: this phosphorylation was inhibited by heparin. A peptide containing a sequence corresponding to a putative pkNII phosphorylation site in topoisomerase I was synthesized and phosphorylated with pkNII. HPLC and two-dimensional analysis show identity between the synthetic phosphorylated peptide and one topoisomerase I phosphopeptide indicating Ser10 as one of the in vitro pkNII phosphorylation sites in topoisomerase I.

Amino Acid Sequence↗
PubMed1993

Phosphorylation of human topoisomerase I by protein kinase C in vitro and in phorbol 12-myristate 13-acetate-activated HL-60 promyelocytic leukaemia cells.

Topoisomerase I was phosphorylated in vitro by protein kinase C (PKC) purified from rat brain with high affinity (Km about 0.1 microM). Tryptic phosphopeptide mapping indicated that two major topoisomerase I peptides phosphorylated in vivo were comigrating with minor peptides phosphorylated by PKC in vitro. Topoisomerase I phosphorylation was stimulated 3-fold in HL-60 cells exposed to the tumour promoter phorbol 12-myristate 13-acetate. The results suggest that topoisomerase I phosphorylation in HL-60 cells is indirectly controlled by PKC.

Animals↗
PubMed1992

Racial differences in the frequencies of scleroderma-related autoantibodies.

OBJECTIVE: To determine demographic differences in scleroderma-related autoantibodies. METHODS: One hundred fifty-six patients with systemic sclerosis were prospectively examined for anticentromere antibodies (ACA), anti-topoisomerase I (anti-topo I, or Scl-70), antinucleolar, and anti-U1 RNP autoantibodies. RESULTS: ACA was found in 36% of Caucasians and 4% of American blacks (P = 0.002, odds ratio [OR] 15). Anti-topo I was found in 37% of American blacks, compared with 17% of Caucasians (P = 0.04, OR 3). No significant differences in the frequencies of antinucleolar and anti-U1 RNP autoantibodies were found. CONCLUSION: These data suggest important demographic differences in scleroderma-associated autoantibodies.

Autoantibodies↗
PubMed1992

Association of amino acid sequences in the HLA-DQB1 first domain with antitopoisomerase I autoantibody response in scleroderma (progressive systemic sclerosis).

Previous studies in Caucasians with progressive systemic sclerosis (PSS) have suggested associations of antitopoisomerase I (antitopo I) autoantibodies with either serologically defined HLA-DR2 or DR5. To better define class II HLA associations with the antitopo I response, 161 PSS patients (132 Caucasians and 29 American blacks) were studied for antitopo I autoantibodies by immunodiffusion and immunoblotting, and their HLA-DRB1, DRB3, DQA1, and DQB1 alleles were determined by restriction fragment length polymorphic analysis and DNA oligotyping. Among Caucasians with antitopo I, HLA-DR5(DRB1*1101-*1104), DRB3*0202 and DQw3 (DQw7,8,9) were significantly increased in frequency. In American blacks, however, only HLA-DQB1*0301(DQw7) was significantly increased. The presence of HLA-DQB1*0301(DQw7) and other HLA-DQB1 alleles bearing the uncharged polar amino acid residue tyrosine at position 30 of the outermost domain was found in all antitopo I-positive Caucasian PSS patients compared with 66% of antitopo I-negative PSS patients (pc = 0.007) and 70% of normal controls (pc = 0.008), as well as all antitopo I-positive black patients. The association with HLA-DQB1 was independent of HLA-DR5(DRB1*1101-*1104) or any other HLA-DRB1, DRB3, or DQA1 alleles. Alternative or additional candidate epitopes for this autoimmune response include alanine at position 38 and threonine at position 77 of these same DQB1 alleles. These data suggest that genetic predisposition to the antitopo I response in PSS is associated most closely with the HLA-DQB1 locus.

Alleles↗
PubMed1991

Acidic pentapeptide phosphorylated in vitro by calf thymus protein kinase NII binds to DNA in the presence of Mg2+ cations.

The pentapeptide pyroGlu-Ala-Glu-Ser-Asn has been synthetized and phosphorylated in vitro at level of serine by protein kinase NII isolated from calf thymus chromatin. It is noteworthy that the calf thymus kinase NII shows a remarkable affinity for this peptide. The [32P]peptide is able to bind to several DNAs in the presence of Mg2+ (lambda phage, calf thymus, pBR540 plasmid). This binding appears not specific with regard to the type of DNA and its base sequence. These data support the hypothesis that phosphorylated acidic domains of nuclear nonhistone proteins could bind directly to DNA in the presence of Mg2+ cations.

Amino Acid Sequence↗
PubMed1990

The major phosphorylation site of nucleophosmin (B23) is phosphorylated by a nuclear kinase II.

Nucleophosmin (B23) was phosphorylated in vitro with [gamma-32P]ATP and a nuclear kinase (type II) purified from HeLa cells. The phosphorylation was inhibited by heparin and by 2,3-diphosphoglycerate. Peptide mapping analysis indicated that the phosphorylation site in vitro was identical to that in vivo. Purified nucleoli have a similar kinase that phosphorylated nucleophosmin at the same site. These results indicated that nucleophosmin is phosphorylated in vivo by a nucleolar kinase (type II).

2,3-Diphosphoglycerate↗
PubMed1989

Identification of consensus epitope structures expressed in recombinant DNA libraries.

In the course of screening cDNA expression libraries with a monospecific polyclonal antibody to topoisomerase I, we isolated three different immunopositive cDNA clones. By comparing their derived amino acid sequences, a consensus region of similarity in otherwise completely dissimilar sequences was identified as an epitope. The approach described here should identify both continuous and discontinuous topological epitopes. The probability of occurrence of "spurious" immunopositive clones is shown to depend on the number of codons of each critical amino acid within the antigenic determinant.

Amino Acid Sequence↗
PubMed1989

Synthetic model peptides phosphorylated in vitro by NII kinase and cation-mediated interactions with DNA.

Minimum substrate requirements for nuclear NII kinase (casein II kinase) were analyzed with synthetic peptides modeled according to amino acid composition of phosphopeptides isolated from chromatin. Uncharged blocked peptide termini decreased the requirement for acidic clusters neighboring the phosphate acceptor amino acid (serine) such that only one group immediately N-terminal to serine was sufficient for kinase activity. Studies on peptide interaction with DNA showed that the model phosphopeptides bound to DNA only in the phosphorylated form suggesting involvement of phosphorylation in protein-DNA interactions yet to be identified.

Amino Acid Sequence↗
PubMed1988

Characterization of a nuclear antigen (Mr 150,000) associated with cell proliferation.

In the course of development of a "library" of monoclonal antibodies to nucleolar proteins, a monoclonal antibody to a nuclear antigen with a molecular weight of 150,000 was obtained. Using this monoclonal antibody as an immunocytochemical probe, low immunofluorescence was demonstrated in human peripheral blood lymphocytes or HL-60 cells treated with retinoic acid. In contrast, a high degree of immunofluorescence was detected in rapidly proliferating human cells (HeLa, Hep-2, HL-60), in Novikoff hepatoma cells, and in phytohemagglutinin-activated human blood lymphocytes. Immunodot blot analysis indicated there were low levels of Mr 150,000 protein in quiescent lymphocytes and increasing amounts after stimulation with phytohemagglutinin. The increased level of Mr 150,000 protein thus is associated with cell proliferation.

Animals↗
PubMed1988

Partial amino acid sequence of rat topoisomerase I: comparison with the predicted sequences for the human and yeast enzymes.

The partial amino acid sequence of rat topoisomerase I was determined by gas-phase microsequencing. Seven tryptic peptides closely matched the sequences deduced from human topoisomerase I cDNA (94.5% homology). Similarity to sequences deduced from baker's yeast and fission yeast genomic DNA were restricted to conserved domains which may represent important sites of interaction with DNA or with other proteins.

Amino Acid Sequence↗
PubMed1987

Microinjection of anti-topoisomerase I immunoglobulin G into nuclei of Chironomus tentans salivary gland cells leads to blockage of transcription elongation.

Purified anti-topoisomerase I immunoglobulin G (IgG) was microinjected into nuclei of Chironomus tentans salivary gland cells, and the effect on DNA transcription was investigated. Synthesis of nucleolar preribosomal 38S RNA by RNA polymerase I and of chromosomal Balbiani ring RNA by RNA polymerase II was inhibited by about 80%. The inhibitory action of anti-topoisomerase I IgG could be reversed by the addition of exogenous topoisomerase I. Anti-topoisomerase I IgG had less effect on RNA polymerase II-promoted activity of other less efficiently transcribing heterogeneous nuclear RNA genes. The pattern of inhibition of growing nascent Balbiani ring chains indicated that the transcriptional process was interrupted at the level of chain elongation. The highly decondensed state of active Balbiani ring chromatin, however, remained unaffected after injection of topoisomerase I antibodies. These data are consistent with the interpretation that topoisomerase I is an essential component in the transcriptional process but not in the maintenance of the decondensed state of active chromatin.

Animals↗
PubMed1987

Novel nucleolar antigens in autoimmune disease.

Antinucleolar antibodies are of interest in 2 important areas, namely, autoimmune diseases and specific products involved in the "mitogenic cascade." The former have delineated a series of novel nuclear and nucleolar elements including the recently described proliferating cell nuclear antigens (PCNA), some of which are present in the nucleolus only at specific times in the G1-S phase of the cell cycle. Important new proteins such as "fibrillarin" (34 kD/pI 8.5) and a 125 kD nucleotide containing protein have been identified with antinucleolar antibodies. Protein p145 is a nucleolar PCNA that is present in growing and dividing cells but not in normal resting tissues. Antinucleolar antibodies offer powerful tools, not only for identification of specific nucleolar proteins important in the cell cycle, but also for purification of their genes and analysis of mechanisms of gene control that operate the "time windows" of the cell cycle.

Amino Acid Sequence↗