Biomedical subjects

Y L Lyu

Publications and source records attributed to Y L Lyu.

5 recordsLinked to original sources

PubMed2001

Inverted repeats as genetic elements for promoting DNA inverted duplication: implications in gene amplification.

Inverted repeats are important genetic elements for genome instability. In the current study we have investigated the role of inverted repeats in a DNA rearrangement reaction using a linear DNA substrate. We show that linear DNA substrates with terminal inverted repeats can efficiently transform Escherichia coli. The transformation products contain circular inverted dimers in which the DNA sequences between terminal inverted repeats are duplicated. In contrast to the recombination/rearrangement product of circular DNA substrates, which is exclusively one particular form of the inverted dimer, the rearrangement products of the linear DNA substrate consist of two isomeric forms of the inverted dimer. Escherichia coli mutants defective in RecBCD exhibit much reduced transformation efficiency, suggesting a role for RecBCD in the protection rather than destruction of these linear DNA substrates. These results suggest a model in which inverted repeats near the ends of a double-strand break can be processed by a helicase/exonuclease to form hairpin caps. Processing of hairpin capped DNA intermediates can then yield inverted duplications. Linear DNA substrates containing terminal inverted repeats can also be converted into inverted dimers in COS cells, suggesting conservation of this type of genome instability from bacteria to mammalian cells.

Animals↗

PubMed2001

Suppression of gene amplification and chromosomal DNA integration by the DNA mismatch repair system.

Mismatch repair (MMR)-deficient cells are shown to produce >15-fold more methotrexate-resistant colonies than MMR normal cells. The increased resistance to methotrexate is primarily due to gene amplification since all the resistant clones contain double-minute chromosomes and increased copy numbers of the DHFR gene. In addition, integration of linearized or retroviral DNAs into chromosomes is also significantly elevated in MMR-deficient cells. These results suggest that in addition to microsatellite instability and homeologous recombination, MMR is also involved in suppression of other genome instabilities such as gene amplification and chromosomal DNA integration.

Adaptor Proteins, Signal Transducing↗

PubMed1999

Inversion/dimerization of plasmids mediated by inverted repeats.

In contrast with earlier studies on the lambda and Escherichia coli genomes, recombination between inverted repeats on plasmids is highly efficient and shown to be recA-independent. In addition, the recombination product is exclusively a head-to-head inverted dimer. Here, we show that this recombination/rearrangement event can occur on different plasmid replicons and is not specific to the particular sequence within the inverted repeats. Transcription readthrough into the inverted repeats has little effect on this event. Genetic analysis has also indicated that most known recombination enzymes are not involved in this process. Specifically, single or double mutants defective in Holliday junction resolution systems (RuvABC and/or RecG/RusA) do not abolish this recombination/rearrangement event. This result does not support the previous models (i.e. the reciprocal-strand-switching and the cruciform-dumbbell models) in which intermediates containing Holliday junctions are proposed. Further analysis has demonstrated that the recombination/rearrangement frequency is dramatically (over 1000-fold) reduced if mismatches (2.8 %) are present within the inverted repeats. Mutations in dam, mutH and mutL genes partially or completely restored the recombination/rearrangement frequency to the level exhibited by the perfect inverted repeats, suggesting the formation of heteroduplexes during recombination/rearrangement. Sequencing analysis of the recombination/rearrangement products have indicated that the majority of the products do not involve crossing-over. We discuss a possible mechanism in which blockage of the lagging strand polymerase by a hairpin triggers recombination/rearrangement mediated by inverted repeats.

Base Sequence↗

PubMed1997

A cruciform-dumbbell model for inverted dimer formation mediated by inverted repeats.

Small inverted repeats (small palindromes) on plasmids have been shown to mediate a recombinational rearrangement event in Escherichia coli leading to the formation of inverted dimers (giant palindromes). This recombinational rearrangement event is efficient and independent of RecA and RecBCD. In this report, we propose a cruciform-dumbbell model to explain the inverted dimer formation mediated by inverted repeats. In this model, the inverted repeats promote the formation of a DNA cruciform which is processed by an endonuclease into a linear DNA with two hairpin loops at its ends. Upon DNA replication, this linear dumbbell-like DNA is then converted to the inverted dimer. In support of this model, linear dumbbell DNA molecules with unidirectional origin of DNA replication (ColE1 ori ) have been constructed and shown to transform E.coli efficiently resulting in the formation of the inverted dimer. The ability of linear dumbbell DNA to transform E.coli suggests that the terminal loops may be important in bypassing the requirement of DNA supercoiling for initiation of replication of the ColE1 ori.

DNA↗

PubMed1995

Specific stimulation of recA-independent plasmid recombination by a DNA sequence at a distance.

Recombination between directly repeated DNA sequences can occur via both recA-dependent and recA-independent mechanisms. They are differentially affected by the length of the repeat and the distance separating the repeats, respectively. Interestingly, a 623 base-pairs long DNA sequence of the plasmid pBR322 was found to stimulate specifically recA-independent recombination between tandem repeats. Analysis of this stimulating sequence has revealed the following features. (1). It is cis-acting. (2). No specific region of it appears to be essential for the effect. Moreover, a neutral sequence of comparable size was able to substitute for the sequence in influencing recombination. (3). The sequence affects recombination between tandem repeats within the tetA but not the bla gene of pBR322. (4). The sequence exerts its effect in a position-dependent manner. (5). It changes not only the frequency but also the products of recombination. Our results provide an example that recA-independent recombination can be influenced by DNA sequences at a distance.

Antiporters↗