Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.5.1.3 (dihydrofolate reductase)
 5,819 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A procedure was developed for mapping origins of DNA replication in mammalian cell chromosomes based on determining the relative abundance of nascent DNA strands throughout a specific genomic region. The method entails purification of short strands of nascent DNA derived from recently activated origins and the quantification, within this sample, of the relative abundances of different adjacent DNA segments by a competitive polymerase chain reaction technique. It is expected that the abundance of defined markers within the origin region is greatest at the site where DNA replication begins. This origin mapping procedure (i) allows analysis of single-copy genomic regions, (ii) can be performed on cultured and primary cells in the absence of any chemical treatment, (iii) does not require cell synchronization, and (iv) allows mapping origins to within a few hundred base pairs. This high degree of resolution permits a study of the cis- and trans-acting elements required for origin function. Application of this method to single-copy sequences in mammalian cells has identified replication origins within an approximately 500-bp segment in the human lamin B2 gene domain and within an approximately 800-bp segment in the hamster dihydrofolate reductase gene locus.
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PMID:Mapping replication origins by quantifying relative abundance of nascent DNA strands using competitive polymerase chain reaction. 944 56

Replication of eukaryotic cell genomes is a tightly controlled process occurring once and only once per cell cycle. Replication initiates at several thousand origins, whose cis-acting sequences and trans-acting proteins have been partially characterized in the yeast S. cerevisiae in the last few years. In contrast, identification of origins of DNA replication in mammalian cells have proven much more difficult. Currently, less then 20 bona fide mammalian origins have been identified, of which only few characterized in detail. Here we discuss the available methods for origin identification in mammalian DNA and the main results, sometimes controversial, so far generated by their application. In particular, we review the currently available information concerning the three best characterized origins, namely those in the lamin B2 and b-globin gene domains in human cells and the one located downstream of the dihydrofolate reductase gene in hamster cells.
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PMID:Replication origins of mammalian chromosomes: the happy few. 1057 95

A small DNA fragment containing the high-frequency initiation region (IR) ori-beta from the hamster dihydrofolate reductase locus functions as an independent replicator in ectopic locations in both hamster and human cells. Conversely, a fragment of the human lamin B2 locus containing the previously mapped IR serves as an independent replicator at ectopic chromosomal sites in hamster cells. At least four defined sequence elements are specifically required for full activity of ectopic ori-beta in hamster cells. These include an AT-rich element, a 4-bp sequence located within the mapped IR, a region of intrinsically bent DNA located between these two elements, and a RIP60 protein binding site adjacent to the bent region. The ori-beta AT-rich element is critical for initiation activity in human, as well as hamster, cells and can be functionally substituted for by an AT-rich region from the human lamin B2 IR that differs in nucleotide sequence and length. Taken together, the results demonstrate that two mammalian replicators can be activated at ectopic sites in chromosomes of another mammal and lead us to speculate that they may share functionally related elements.
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PMID:Defined sequence modules and an architectural element cooperate to promote initiation at an ectopic mammalian chromosomal replication origin. 1512 36