Impaired Translesion Synthesis in Xeroderma Pigmentosum Variant Extracts

Xeroderma pigmentosum variant (XPV) cells are characterized by a cellular defect in the ability to synthesize intact daughter DNA strands on damaged templates. Molecular mechanisms that facilitate replication fork progression on damaged DNA in normal cells are not well defined. In this study, we used single-stranded plasmid molecules containing a single N-2-acetylaminofluorene (AAF) adduct to analyze translesion synthesis (TLS) catalyzed by extracts of either normal or XPV primary skin fibroblasts. In one of the substrates, the single AAF adduct was located at the 3' end of a run of three guanines that was previously shown to induce deletion of one G by a slippage mechanism. Primer extension reactions performed by normal cellular extracts from four different individuals produced the same distinct pattern of TLS, with over 80% of the products resulting from the elongation of a slipped intermediate and the remaining 20% resulting from a nonslipped intermediate. In contrast, with cellular extracts from five different XPV patients, the TLS reaction was strongly reduced, yielding only low amounts of TLS via the nonslipped intermediate. With our second substrate, in which the AAF adduct was located at the first G in the run, thus preventing slippage from occurring, we confirmed that normal extracts were able to perform TLS 10-fold more efficiently than XPV extracts. These data demonstrate unequivocally that the defect in XPV cells resides in translesion synthesis independently of the slippage proces

Mutagenicity of Acridines in a Reversion Assay Based on Tetracycline Resistance in Plasmid pBR322 in Escherichia Coli

The mutagenicity of a series of acridine compounds was studied in an assay based on the reversion of mutations in the tetracycline-resistance gene (tet) of plasmid pBR322 in Escherichia coli. Mutations that restore the tetracycline-resistant phenotype were detected in tetracycline-sensitive strains carrying mutant plasmids. Mutations that revert by +2, +1, −1, and −2 frameshift mutations and by base-pair substitutions were used to analyze the mutagenicity of two simple acridines, two acridine mustards, and a nitroacridine. The simple acridines (9-aminoacridine and quinacrine) effectively induced −1 frameshifts and weakly induced +1 frameshifts. The acridine mustards (quinacrine mustard and ICR-191) were more potent inducers of −1 and +1 frameshifts than the simple acridines. Reactive acridines, including both the mustards and the nitroacridine Entozon, were effective inducers of −2 frameshifts but the simple acridines were not. The two classes of reactive acridines differed from one another, in that the mustards were better inducers of +1 frameshifts than Entozon, whereas Entozon was a particularly potent inducer of −2 frameshifts. None of the compounds induced +2 frameshifts, and the induction of base-pair substitutions was negligible. These results confirm and extend studies showing that adduct-forming acridines are stronger frameshift mutagens than simple intercalating acridines and that the acridines differ from one another not only in overall mutagenic potency but also in the prevalence of different classes of frameshift mutations

Optimal Eavesdropping in Quantum Cryptography. II. Quantum Circuit

It is shown that the optimum strategy of the eavesdropper, as described in the preceding paper, can be expressed in terms of a quantum circuit in a way which makes it obvious why certain parameters take on particular values, and why obtaining information in one basis gives rise to noise in the conjugate basis.Comment: 7 pages, 1 figure, Latex, the second part of quant-ph/970103

Gaps and forks in DNA replication: Rediscovering old models

Most current models for replication past damaged lesions envisage that translesion synthesis occurs at the replication fork. However older models suggested that gaps were left opposite lesions to allow the replication fork to proceed, and these gaps were subsequently sealed behind the replication fork. Two recent articles lend support to the idea that bypass of the damage occurs behind the fork. In the first paper, electron micrographs of DNA replicated in UV-irradiated yeast cells show regions of single-stranded DNA both at the replication forks and behind the fork, the latter being consistent with the presence of gaps in the daughter-strands opposite lesions. The second paper describes an in vitro DNA replication system reconstituted from purified bacterial proteins. Repriming of synthesis downstream from a blocked fork occurred not only on the lagging strand as expected, but also on the leading strand, demonstrating that contrary to widely accepted beliefs, leading strand synthesis does not need to be continuous

Involvement of DnaE, the second replicative DNA polymerase from Bacillus subtilis, in DNA mutagenesis

In a large group of organisms including low G + C bacteria and eukaryotic cells, DNA synthesis at the replication fork strictly requires two distinct replicative DNA polymerases. These are designated pol C and DnaE in Bacillus subtilis. We recently proposed that DnaE might be preferentially involved in lagging strand synthesis, whereas pol C would mainly carry out leading strand synthesis. The biochemical analysis of DnaE reported here is consistent with its postulated function, as it is a highly potent enzyme, replicating as fast as 240 nucleotides/s, and stalling for more than 30 s when encountering annealed 5'-DNA end. DnaE is devoid of 3' --> 5'-proofreading exonuclease activity and has a low processivity (1-75 nucleotides), suggesting that it requires additional factors to fulfill its role in replication. Interestingly, we found that (i) DnaE is SOS-inducible; (ii) variation in DnaE or pol C concentration has no effect on spontaneous mutagenesis; (iii) depletion of pol C or DnaE prevents UV-induced mutagenesis; and (iv) purified DnaE has a rather relaxed active site as it can bypass lesions that generally block other replicative polymerases. These results suggest that DnaE and possibly pol C have a function in DNA repair/mutagenesis, in addition to their role in DNA replication

Kinematics of Metal-Poor Stars in the Galaxy. II. Proper Motions for a Large Non-Kinematically Selected Sample

We present a revised catalog of 2106 Galactic stars, selected without kinematic bias, and with available radial velocities, distance estimates, and metal abundances in the range 0.0 <= [Fe/H] <= -4.0. This update of the Beers and Sommer-Larsen (1995) catalog includes newly-derived homogeneous photometric distance estimates, revised radial velocities for a number of stars with recently obtained high-resolution spectra, and refined metallicities for stars originally identified in the HK objective-prism survey (which account for nearly half of the catalog) based on a recent re-calibration. A subset of 1258 stars in this catalog have available proper motions, based on measurements obtained with the Hipparcos astrometry satellite, or taken from the updated Astrographic Catalogue (AC 2000; second epoch positions from either the Hubble Space Telescope Guide Star Catalog or the Tycho Catalogue), the Yale/San Juan Southern Proper Motion (SPM) Catalog 2.0, and the Lick Northern Proper Motion (NPM1) Catalog. Our present catalog includes 388 RR Lyrae variables (182 of which are newly added), 38 variables of other types, and 1680 non-variables, with distances in the range 0.1 to 40 kpc.Comment: 31 pages, including 8 figures, to appear in AJ (June 2000), full paper with all figures embedded available at http://pluto.mtk.nao.ac.jp/people/chiba/preprint/halo4