Expression of the E. coli uvrA gene is inducible

UvrA+-dependent excision repair is one of the most important systems in Escherichia coli for repairing UV-induced pyrimidine dimers and a variety of other forms of DNA damage. The uvrA protein acts in conjunction with the uvrB and uvrC gene products to introduce a nick at the of a DNA lesion and thus initiate the repair process. We have recently used the Mud(Ap, lac) operon fusion vector to identify a set of genes whose expression is induced by DNA damage. One Mud(Ap, lac) insertion mapped at the uvrA locus and made the cells sensitive to UV light. In this fusion strain, beta-galactosidase expression was induced by DNA-damaging agents in a recA+lexA+-dependent fashion. We were surprised by this result because uvrA+-dependent excision repair is observed both in cells in which protein synthesis has been inhibited and in recA- and lexA- cells, findings which have led to the conclusion that the uvrA gene product is constitutively expressed and not under the control of the complex recA+lexA+ regulatory circuitry (see below). We have investigated this possibility further and describe here the generation and characterization of a set of fusions of the lac genes to the promoter of the uvrA gene. We confirm that the uvrA gene product is induced by DNA damage in a recA+lexA+-dependent fashion.     

A model for SOS-lesion-targeted mutations in Escherichia coli

The UmuD'2C protein complex (Escherichia coli pol V) is a low-fidelity DNA polymerase (pol) that copies damaged DNA in the presence of RecA, single-stranded-DNA binding protein (SSB) and the beta,gamma-processivity complex of E. coli pol III (ref. 4). Here we propose a model to explain SOS-lesion-targeted mutagenesis, assigning specific biochemical functions for each protein during translesion synthesis. (SOS lesion-targeted mutagenesis occurs when pol V is induced as part of the SOS response to DNA damage and incorrectly incorporates nucleotides opposite template lesions.) Pol V plus SSB catalyses RecA filament disassembly in the 3' to 5' direction on the template, ahead of the polymerase, in a reaction that does not involve ATP hydrolysis. Concurrent ATP-hydrolysis-driven filament disassembly in the 5' to 3' direction results in a bidirectional stripping of RecA from the template strand. The bidirectional collapse of the RecA filament restricts DNA synthesis by pol V to template sites that are proximal to the lesion, thereby minimizing the occurrence of untargeted mutations at undamaged template sites.     

Sequence of the human insulin gene

The human insulin gene contains two intervening sequences, one is within the region transcribed into the 5'-untranslated segment of the mRNA and the other interrupts the C-peptide encoding region. A comparison of the human with the rat insulin genes indicates potential regulatory regions in the DNA segment preceding the gene and suggests that the ancestral form of the insulin gene had two intervening sequences.     

用计算机对人类TSPYl基因P53结合位点的鉴定

根据p53下游基因在其调节区域（启动子或内含子）含有与P53蛋白特异性结合的一致性序列5’-RRRCWWGYYYN（0-13）RRRCWWGYYY-3’，R—G或A，W—T或A，Y—C或T，N—A，C，T，G。用计算机对人类基因组中P53结合位点进行了研究，发现Y染色体上的TSPY1基因内含子中含有这样的一致性序列5’-GGGCTAGTTTtgGAGCTAGCCT-3’，意味着TSPY1基因有可能是一个p53下游基因。     

Requirement for the replication protein SSB in human DNA excision repair

Replication and repair are essential processes that maintain the continuity of the genetic material. Dissection of simian virus 40 (SV40) DNA replication has resulted in the identification of many eukaryotic replication proteins, but the biochemistry of the multienzyme process of DNA excision repair is less well defined. One protein that is absolutely required for semiconservative replication of SV40 DNA in vitro is human single-stranded DNA-binding protein (SSB, also called RF-A and RP-A). SSB consists of three polypeptides of relative molecular mass 70,000, 34,000 and 13,000, and acts with T antigen and topoisomerases to unwind DNA, allowing the access of other replication proteins. Human SSB can also stimulate the activity of polymerases alpha and delta, suggesting a further role in elongation during DNA replication. We have now found a role for human SSB in DNA excision repair using a cell-free system that can carry out nucleotide excision repair in vitro. Monoclonal antibodies against human SSB caused extensive inhibition of DNA repair in plasmid molecules damaged by ultraviolet light or acetylaminofluorene. Addition of purified SSB reversed this inhibition and further stimulated repair synthesis by increasing the number of repair events. These results show that a mammalian DNA replication protein is also essential for repair.     

Stimulation of protein-directed strand exchange by a DNA helicase

The protein-mediated exchange of strands between a DNA double helix and a homologous DNA single strand involves both synapsis and branch migration, which are two important aspects of any general recombination reaction. Purified DNA-dependent ATPases from Escherichia coli (recA protein), Ustilago (rec 1 protein) and phage T4 (uvsX protein) have been shown to drive both synapsis and branch migration in vitro. The T4 gene 32 protein is a helix-destabilizing protein that greatly stimulates uvsX-protein-catalysed synapsis, and the E. coli SSB (single-strand binding) protein stimulates the analogous recA-protein-mediated reaction to a lesser degree. One suspects that several other proteins also play a role in the strand exchange process. For example, a DNA helicase could in principle accelerate branch migration rates by helping to melt the helix at the branch point. The T4 dda protein is a DNA helicase that is required to move the T4 replication fork past DNA template-bound proteins in vitro. Previously, we have shown that the dda protein binds to a column that contains immobilized T4 uvsX protein. We show here that this helicase specifically stimulates the branch migration reaction that the uvsX protein catalyses as a central part of the genetic recombination process in a T4 bacteriophage-infected cell.     

yigP——大肠杆菌生长所必需的基因

yigP基因在大肠杆菌基因组上位于辅酶Q生物合成相关基因ubiE和ubiB之间,3者构成一个操纵子。利用温敏质粒pMAK705系统敲除大肠杆菌基因组上的yigP基因后,发现二次重组子内的温敏质粒无法通过高温培养丢失,即染色体上的yigP基因被敲除后,必须依靠质粒上完整的yigP基因才能存活;通过功能回补yigP基因的表...     

N-Acetoxy-acetylaminofluorene reacts preferentially with a control region of intracellular SV40 chromosome

Many chemical carcinogens or their metabolites react with DNA; thus it is of interest to determine what effect chromosomal structure has on these reactions. The chromosome of simian virus 40 (SV40) is well suited for such studies; like chromatin of eukaryotic cells, it is organized into nucleosomes. The nucleotide sequence of SV40 is known, together with much about the pattern of viral gene expression and DNA replication, and the structure of the viral chromosome. We have investigated the binding of the ultimate carcinogen, N-acetoxy-acetylaminofluorene (AAAF), to specific regions of the SV40 chromosome in situ in the intact infected cell. The results, reported here, indicate that a region containing regulatory functions on the intracellular SV40 chromosome has unique structural properties which render it more susceptible to attack by AAAF than the rest of the SV40 genome. The preferential binding of AAAF to regulatory regions of chromatin may have implications for the mechanism of action of this and similar carcinogens.     

Sequence of the lacI gene.

The structural gene for the lac repressor of Escherichia coli, the lacI gene has been sequenced. This 1,080 base pair region of the E. coli chromosome codes for the lac repressor protein of 360 amino acids. The DNA sequence largely confirms but extends the previously reported protein sequence and allows a structural analysis of genetic phenomena.