Cell
Volume 52, Issue 3, 12 February 1988, Pages 375-383
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Article
Enhancer-origin interaction in plasmid R6K involves a DNA loop mediated by initiator protein

https://doi.org/10.1016/S0092-8674(88)80030-8Get rights and content

Summary

Initiation of DNA replication from on β of plasmid R6K requires the presence of the on γ sequence in cis. We demonstrate that binding of initiator protein to the seven strong, tandem binding sites in γ increases binding of the protein at the very weak binding site present in on β by cooperativity at a distance. The γ-β interaction via the initiator results in a DNA loop, as revealed by the novel technique of cyclization enhancement and as confirmed by exonuclease III protection, electron microscopy, and chemical footprinting. The protein-mediated γ-β interaction in vitro suggests that the cooperative interaction of γ-bound protein with the β sequence by DNA looping is an early step in the initiation of DNA replication at the β origin of R6K.

References (32)

Cited by (97)

  • Plasmid R6K replication control

    2013, Plasmid
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    π protein facilitates interaction between the iterons in γ ori and iteron-like sequences in oris α and β (Mukherjee et al., 1988b; Miron et al., 1992; Saxena et al., 2010a, 2010b). This results in a looping of the plasmid DNA, which is believed to transmit the replication signal from the internal iteron cluster (7 DRs) to the outlying R6K oris (Mukherjee et al., 1988a, 1988b). Studies of the interaction between γ and α oris suggest that both monomers and dimers of π protein participate in the looping process (Saxena et al., 2010a, 2010b).

  • Regulation of Replication Termination by Reb1 Protein-Mediated Action at a Distance

    2010, Cell
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    We wished to confirm the aforementioned ligation enhancement results by an independent method. The method is schematically shown in Figure 3A (Mukherjee et al., 1988; Zzaman and Bastia, 2005). The plasmid DNA substrate pSW1 contained a M7 and a WT Ter site cloned on either sides of the unique EcoR1 site.

  • Investigations of π initiator protein-mediated interaction between replication origins α and γ of the plasmid R6K

    2010, Journal of Biological Chemistry
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    The replication origins α and β are located ∼3800 and ∼1200 bp away, respectively, from ori γ and at its opposite side (11, 15). We have previously reported that π-mediated DNA looping between the single iteron at α or the half-iteron at β with the γ iteron array appears to activate the two distantly located origins, respectively (9, 13, 16). Models of activation of each of the three origins, that are mutually exclusive, and the postulated looped structures involved in the process are shown in Fig. 1.

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