Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P01034 (cystatin C)
 3,397 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

10 derivations of rat tracheal epithelial (RTE) cells, including normal cells, normal primary cultures, 7 tumorigenic cell lines and 1 nontumorigenic cell line transformed in vitro by treatment with 7,12-dimethylbenz[a]anthracene (DMBA), benzo[a]pyrene (BP) and/or 12-O-tetradecanoylphorbol-13-acetate (TPA) were examined for oncogene alterations. No abnormalities of Ha-ras or Ki-ras were seen that were suggestive of amplification, rearrangement or the presence of RFLPs. Analysis of specific-point mutations in Ha-ras using Pst I digestion (codon 12, GGA to GCA) or Ha-ras and Ki-ras using Xba I (codon 61, CAA to CTA) were negative. In one cell line derived by DMBA treatment, changes in the c-myc restriction digest pattern were seen after incubation with Bam HI and Hind III. Northern analysis revealed consistent differences between normal and transformed cells when probed with Ha-ras; c-myc expression was of low intensity, and the expression of Ki-ras could not be detected. Transfection of RTE cell DNAs into NIH/3T3 cells did not result in the appearance of morphologic transformants. The studies suggest that Ha-ras or Ki-ras codon 61 A to T transversions (CAA to CTA) are not associated with the immortal/tumorigenic phenotype in RTE cells transformed by DMBA or TPA, and are in contrast to results reported in some other biological systems.
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PMID:Oncogene alterations in in vitro transformed rat tracheal epithelial cells. 169 45

Mouse skin tumors contain activated c-H-ras oncogenes, often caused by point mutations at codons 12 and 13 in exon 1 and codons 59 and 61 in exon 2. Mutagenesis by the noncoding apurinic sites can produce G-->T and A-->T transversions by DNA misreplication with more frequent insertion of deoxyadenosine opposite the apurinic site. Papillomas were induced in mouse skin by several aromatic hydrocarbons, and mutations in the c-H-ras gene were determined to elucidate the relationship among DNA adducts, apurinic sites, and ras oncogene mutations. Dibenzo[a,l]pyrene (DB[a,l]P), DB[a,l]P-11,12-dihydrodiol, anti-DB[a,l]P-11,12-diol-13,14-epoxide, DB[a,l]P-8,9-dihydrodiol, 7,12-dimethylbenz[a]anthracene (DMBA), and 1,2,3,4-tetrahydro-DMBA consistently induced a CAA-->CTA mutation in codon 61 of the c-H-ras oncogene. Benzo[a]pyrene induced a GGC-->GTC mutation in codon 13 in 54% of tumors and a CAA-->CTA mutation in codon 61 in 15%. The pattern of mutations induced by each hydrocarbon correlated with its profile of DNA adducts. For example, both DB[a,l]P and DMBA primarily form DNA adducts at the N-3 and/or N-7 of deoxyadenosine that are lost from the DNA by depurination, generating apurinic sites. Thus, these results support the hypothesis that misreplication of unrepaired apurinic sites generated by loss of hydrocarbon-DNA adducts is responsible for transforming mutations leading to papillomas in mouse skin.
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PMID:Relating aromatic hydrocarbon-induced DNA adducts and c-H-ras mutations in mouse skin papillomas: the role of apurinic sites. 747 97

The frequency and spectrum of Ha-ras mutations in benzo[a]pyrene (B[a]P)-initiated/12-O-tetradecanoylphorbol-13-acetate (TPA)-promoted CD-1 mouse skin papillomas were characterized by amplifying high molecular weight papilloma DNA using the polymerase chain reaction (PCR) followed by direct DNA sequencing. Analysis of 10 individual B[a]P-initiated early emergence papillomas indicated that 90% contained a Ha-ras mutation. Twenty percent of these papillomas contained a GGA-->GTA transversion in the 12th codon, 50% contained a GGC-->GTC transversion in the 13th codon and 20% contained a CAA-->CTA transversion in the 61st codon. A characteristic of 7,12-dimethylbenz[a]anthracene (DMBA)-initiated papillomas, which contain an A-->T mutation in the 61st codon of Ha-ras, is that they exhibit a constitutive decrease in both protein kinase C (PKC) activity and PKC alpha and beta 2 isozyme levels when compared to epidermis. In the present study we found that total PKC activity, as well as PKC alpha and beta 2 isoforms, were markedly decreased in B[a]P-initiated early emergence papillomas and that this decrease was also accompanied by an altered subcellular distribution of PKC activity. The particulate/cytosolic (P/C) ratio of PKC activity in the epidermis was 0.39, whereas the P/C ratio in the papillomas was 0.77. These results demonstrate that B[a]P-initiated/TPA-promoted papillomas exhibit a high incidence of specific ras mutations and that PKC levels are constitutively decreased in these papillomas, indicating that an activated ras gene is associated with and may contribute to the observed decrease in PKC levels.
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PMID:Characterization of benzo[a]pyrene-initiated mouse skin papillomas for Ha-ras mutations and protein kinase C levels. 824 57

One of the key events in tumor initiation in mouse skin is mutational activation of the H-ras gene. Papillomas induced by the most carcinogenic environmental polycyclic aromatic hydrocarbon (PAH), dibenzo[a,l]pyrene (DB[a,l]P), in SENCAR mouse skin contain a specific H-ras codon 61 (CAA-->CTA) mutation. We describe here detection of these mutations in preneoplastic skin by measuring the frequency of an induced XbaI RFLP, created by the mutation. Development of the PCR-XbaI RFLP method, sensitive enough to detect 1 codon 61 mutant allele among 10,000 wild-type genes, is described. The results indicate that codon 61 mutations are induced 1 day (0.1%) after DB[a,l]P treatment on mouse skin, reach a high value (5%) by day 3, rapidly decline between days 7-9 and increase again during the clonal expansion of pre-papillomas into tumors. The detection of codon 61 mutations 1 day after DB[a,l]P exposure suggests that mutations occurred by pre-replication misrepair.
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PMID:Detection of dibenzo[a,l]pyrene-induced H-ras codon 61 mutant genes in preneoplastic SENCAR mouse skin using a new PCR-RFLP method. 967

Initiation of skin tumors in mice is associated with the formation of oncogenic mutations in the H-ras gene. Mice treated on the dorsal skin with the potent polycyclic aromatic hydrocarbon (PAH) carcinogen dibenzo[a,l]pyrene (DB[a,l]P) form papillomas carrying the H-ras codon 61 (CAA to CTA) mutations. These mutations are induced in early preneoplastic skin within 1 day after DB[a,l]P treatment (Oncogene 16 (1998) 3203-3210) and appear to be related to DB[a,l]P-Ade-depurinating adducts (Proc. Natl. Acad. Sci. U. S. A. 92 (1995) 10422-10426). The rapid kinetics of mutation induction suggests that abasic sites generated from base depurination may undergo error-prone excision repair in pre-S-phase cells to induce these mutations. Analysis of mutations in the H-ras exon 1 and 2 region in DB[a,l]P-treated early preneoplastic skin indicated great changes in mutation spectra in the preneoplastic period. The initial spectra contained abundant A-->G mutations, which frequently occurred 3' to a putative conserved sequence (TGN-doublet). These mutations appeared to be induced initially as mismatched (G.T) heteroduplexes and then converted into double-stranded mutations by one round of replication. Unlike the A-->G mutations found in DB[a, l]P-treated skin (which forms 99% depurinating adducts), A-->G mutations found in anti-DB[a,l]P-diol epoxide-treated skin (forms 97% stable adducts) did not appear to be G.T heteroduplexes. These results, therefore, suggest that under these conditions, the repair errors occurred only from abasic sites but not from stable adducts. Initiated cells carrying specific oncogenic mutations, formed presumably by misrepair, underwent rapid clonal expansion and regression (transient clonoplasia). The multiplication of initiated stem cells during transient clonoplasia may be a factor determining the tumor-initiating potential of some PAH carcinogens.
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PMID:Evidence that error-prone DNA repair converts dibenzo[a,l]pyrene-induced depurinating lesions into mutations: formation, clonal proliferation and regression of initiated cells carrying H-ras oncogene mutations in early preneoplasia. 1108 92

Benzo[a]pyrene (BP), a prototype polycyclic aromatic hydrocarbon (PAH), can be metabolically activated to the enantiomeric benzo[a]pyrene diol epoxides (BPDEs), (+)-(7R,8S,9S,10R)-7,8-dihydroxy-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene and the (-)-(7S,8R,9R,10S) enantiomer. These can react with adenine residues in DNA, to produce the stereoisomeric 10S (+)- and 10R (-)-trans-anti-[BP]-N(6)-dA adducts. High-resolution NMR solution studies indicate that in DNA duplexes the 10R (-) adduct is intercalated on the 5'-side of the modified adenine, while the 10S (+) adduct is disordered, exhibits multiple adduct conformations, and is positioned on the 3'-side of the modified adenine. Duplexes containing the 10S (+) adduct positioned at A within codon 61 of the human N-ras sequence CAA are thermodynamically less stable and more easily excised by human DNA repair enzymes than those containing the 10R (-) adduct. However, the molecular origins of these differences are not understood and represent a fascinating opportunity for elucidating structure-function relationships. We have carried out a computational investigation to uncover the structural and thermodynamic origins of these effects in the 11-mer duplex sequence d(CGGACAAGAAG).d(CTTCTTGTCCG) by performing a 2-ns molecular dynamics simulation using NMR solution structures as the basis for the starting models. Then, we applied the MM-PBSA (molecular mechanics Poisson-Boltzmann surface area) method to compute free energy differences between the stereoisomeric adducts. The 10R (-) isomer is more stable by approximately 13 kcal/mol, of which approximately 10 kcal/mol is enthalpic, which agrees quite well with their observed differences in thermodynamic stability. The lower stability of the 10S (+) adduct is due to diminished stacking by the BP moiety in the intercalation pocket, more helix unwinding, and a diminished quality of Watson-Crick base pairing. The latter stems from conformational heterogeneity involving a syn-anti equilibrium of the glycosidic bond in the modified adenine residue. The lower stability and conformational heterogeneity of the 10S (+) adduct may play a role in its enhanced susceptibility to nucleotide excision repair.
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PMID:Stereochemical, structural, and thermodynamic origins of stability differences between stereoisomeric benzo[a]pyrene diol epoxide deoxyadenosine adducts in a DNA mutational hot spot sequence. 1145 84

Topical application of dibenzo[a,l]pyrene (DB[a,l]P) to the dorsal skin of SENCAR mice induces codon 61 (CAA Gln to CTA Leu) mutations in the Harvey (H)-ras gene within 12 h after treatment. Between days 1 and 3, the frequency of these mutations increases rapidly, suggesting that skin cells carrying the codon 61 mutations proliferate in this period. We have investigated DB[a,l]P-treated mouse skin (12 h-7 d) for further evidence of H-ras expression and epidermal cell proliferation. Two waves of cell proliferation were observed: the first wave (1-2 d) correlated with the clonal proliferation of codon 61-mutated cells, and the second wave (3-7 d) correlated with DB[a,l]P-induced hyperplasia. DB[a,l]P-induced early preneoplastic cell proliferation correlated with H-ras and specific G1 cyclin expression. Total H-ras protein and cyclin D1 were found to increase during DB[a,l]P-induced hyperplasia, but the levels of guanosine triphosphate-bound (active) H-ras protein and cyclin E were increased during the putative clonal proliferation of codon 61-mutated cells. These results suggest that DB[a,l]P-induced oncogenically mutated cells proliferate in early preneoplastic skin. As this proliferation occurs in the absence of any promoting treatment, we propose that this phenomenon is a tumor initiation event.
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PMID:Harvey-ras gene expression and epidermal cell proliferation in dibenzo[a,l]pyrene-treated early preneoplastic SENCAR mouse skin. 1611

Polycyclic aromatic hydrocarbons (PAH) form stable and depurinating DNA adducts in mouse skin to induce preneoplastic mutations. Some mutations transform cells, which then clonally expand to establish tumors. Strong clues about the mutagenic mechanism can be obtained if the PAH-DNA adducts can be correlated with both preneoplastic and tumor mutations. To this end, we studied mutagenesis in PAH-treated early preneoplastic skin (1 day after exposure) and in the induced papillomas in SENCAR mice. Papillomas were studied by PCR amplification of the H-ras gene and sequencing. For benzo[a]pyrene (BP), BP-7,8-dihydrodiol (BPDHD), 7,12-dimethylbenz[a]anthracene (DMBA) and dibenzo[a,l]pyrene (DB[a,l]P), the codon 13 (GGC to GTC) and codon 61 (CAA to CTA) mutations in papillomas corresponded to the relative levels of Gua and Ade-depurinating adducts, despite BP and BPDHD forming significant amounts of stable DNA adducts. Such a relationship was expected for DMBA and DB[a,l]P, as they formed primarily depurinating adducts. These results suggest that depurinating adducts play a major role in forming the tumorigenic mutations. To validate this correlation, preneoplastic skin mutations were studied by cloning H-ras PCR products and sequencing individual clones. DMBA- and DB[a,l]P-treated skin showed primarily A.T to G.C mutations, which correlated with the high ratio of the Ade/Gua-depurinating adducts. Incubation of skin DNA with T.G-DNA glycosylase eliminated most of these A.T to G.C mutations, indicating that they existed as G.T heteroduplexes, as would be expected if they were formed by errors in the repair of abasic sites generated by the depurinating adducts. BP and its metabolites induced mainly G.C to T.A mutations in preneoplastic skin. However, PCR over unrepaired anti-BPDE-N(2)dG adducts can generate similar mutations as artifacts of the study protocol, making it difficult to establish an adduct-mutation correlation for determining which BP-DNA adducts induce the early preneoplastic mutations. In conclusion, this study suggests that depurinating adducts play a major role in PAH mutagenesis.
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PMID:The role of polycyclic aromatic hydrocarbon-DNA adducts in inducing mutations in mouse skin. 1793 59

DNA structural perturbations that are induced by site specifically and stereospecifically defined benzo[a]pyrene-7,8-dihydrodiol-9,10-epoxide (BPDE) adducts are directly correlated with mutagenesis, leading to cellular transformation. Although previous investigations had established that replication of DNAs containing N(6) -BPDE dA adducts at the second position in the N-ras codon 61(CAA) (61(2) ) resulted exclusively in A to G transitions, NMR analyses not only established the structural basis for this transition mutation but also predicted that if the adduct were positioned at the third position in the same codon, an expanded spectra of mutations was possible. To test this prediction, replication of DNAs containing C10 S-BPDE and C10 R-BPDE lesions linked through the N(6) position of adenine in the sequence context N-ras codon 61, position 3 (C10 S-BPDE and C10 R-BPDE at 61(3) ) was carried out in Escherichia coli, and these data revealed a wide mutation spectrum. In addition to A to G transitions produced by replication of both lesions, replication of the C10 S-BPDE and C10 R-BPDE adducts also yielded A to C and A to T transversions, respectively. Analyses of single nucleotide incorporation using Sequenase 2.0 and exonuclease-deficient E. coli Klenow fragment and pol II not only revealed high fidelity synthesis but also demonstrated the same hierarchy of preference opposite a particular lesion, independent of the sequence context. Primer extension assays with the two lesions at N-ras 61(3) resulted in truncated products, with the C10 S-BPDE adducts being more blocking than C10 R-BPDE lesions, and termination of synthesis was more pronounced at position 61(3) than at 61(2) for each of the lesions.
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PMID:Sequence context modulation of polycyclic aromatic hydrocarbon-induced mutagenesis. 2391 16