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Biomedical subjects

M E Woodworth

Publications and source records attributed to M E Woodworth.

5 recordsLinked to original sources
PubMed2001

DNA replication efficiency depends on transcription factor-binding sites.

Naturally arising variants of simian virus 40 (SV40), generated by serial passage of the virus at high multiplicities of infection, provide important insight into the role of transcription factor-binding sites in enhancing DNA replication. Although the variants that arise from numerous recombination events are the result of selective pressure to replicate more efficiently than the other variants in the infection, there is no transcription pressure. Therefore, it is interesting that a minimum of two viral Sp1 transcription factor-binding sites are retained and that host AP-1 and NF-1 transcription factor-binding sites are incorporated into the 100-bp regulatory region that maximizes DNA replication in these variants. We cotransfected COS-1 cells (that provide viral large T antigen for DNA replication) to examine the effect of transcription factor-binding sites on the replication of plasmid constructs that contain the SV40 origin of replication (ori). The level of relative replication efficiency (RRE) depends on the number and type of transcription factor-binding sites. Replication increases as the number of transcription factor-binding sites increases within the regulatory region of the variants; AP-1 sites are more effective than NF-1 transcription factor-binding sites. Competition between constructs in transfections magnifies the difference in their RREs. The results indicate that transcription factor-binding sites play an important role in enhancing DNA replication.

Animals↗
PubMed1999

Conformational changes in simian virus 40 rearranged regulatory regions: effects of the 21-base-pair promoters and their location.

Simian virus 40 (SV40) is an excellent model system for investigating the cis- and trans-acting factors involved in eukaryotic DNA replication because it uses host enzymes, with the exception of the virus-encoded T-antigen (T-ag), to replicate its genome. Although its origin of replication (ori) is essential for DNA replication, there are transcriptional promoters and enhancers that affect DNA replication efficiency. T-ag binds to sites I to III within and around ori with different affinities and induces structural changes. We were interested in determining if the position of the promoters relative to ori influences the binding of T-ag to these regions. Furthermore, we characterized the DNA structural changes that occur as a result of protein binding when the promoters are absent and also when the promoters are moved from their wild-type position upstream of ori to a position downstream of ori. Using sequence- and conformation-specific chemical probes, our data indicate that (i) the conformation of site I is influenced by T-ag binding and by flanking sequences, (ii) the conformation of the promoters after T-ag binding is dependent on their location, and (iii) unwinding of ori is influenced by the location of the promoters and their presence or absence. These differences in DNA conformation may help explain decreases in relative DNA replication efficiency that occur when the promoters are absent or located downstream of ori.

Animals↗
PubMed1993

Characterization of murine middle repetitive DNA.

We have characterized four sequences from a small library containing a subset of the repetitive families of the mouse. Each clone has a repetition frequency of less than 1,100 copies per genome and each clone represents a unique family of middle repetitive DNA. One clone (pMR111) shares homology with mouse intracisternal A-particle (IAP) elements, a second clone (pMR89) has partial homology with a sequence in the 3' untranslated region of the human fibulin gene, while two clones (pMR6, pMR66) are new that have no homology to any reported DNA sequence. Each clone hybridizes to one or two discrete RNA transcripts from one or more tissues of the mouse. Clone pMR66 detected restriction fragment length polymorphisms (RFLPs) at three loci in genomic mouse DNA, defining loci at the distal end of chromosome 5 and the proximal end of chromosome 7. The third locus is unmapped. This study demonstrates that cloned repeats from a repetitive DNA library are a potential source of genetic markers.

Amino Acid Sequence↗