Affiliation:
1. Department of Biological Sciences, University of Illinois at Chicago Circle, Chicago, Illinois 60680
Abstract
A hypermodified base (Y-Thy) replaces 20% of the thymine (Thy) in mature DNA of
Bacillus subtilis
phage SP10. Two noncomplementing hypermodification-defective (
hmd
) mutants are described. At 30°C,
hmd
phage carried out a normal program, but at temperatures of ≥37°C, the infection process was nonproductive. When cells were infected at 37°C with
hmd
phage, DNA synthesis started at its usual time (12 min), proceeded at about half the normal rate for 6 to 8 min, and then stopped or declined manyfold. All, or nearly all, of the DNA made under
hmd
conditions consisted of fully hypermodified parental DNA strands H-bonded to unhypermodified nascent strands. The reduced levels of DNA synthesis observed under
hmd
conditions were accompanied by weak expression of late genes. A sucrose gradient analysis of SP10
hmd
+
replicating DNA intermediates was made. Two intermediates, called VG and F, were identified. VF consisted of condensed DNA complexed to protein; VF also contained negatively supercoiled domains covalently joined to relaxed regions. F was composed of linear concatenates from which mature DNA was cleaved. None of those intermediates was evident in cells infected at 37°C with
hmd
phage. Shiftup experiments were performed wherein cells infected with
hmd
phage at 30°C were shifted to 37°C at a time when replication was well under way. DNA synthesis stopped or declined manyfold 10 min after shiftup. The
hmd
DNA made after shiftup was conserved as a form sedimentationally equivalent to the F intermediate, but little mature DNA was evident. It is proposed that Y-Thy is required for replication and DNA maturation because certain key proteins involved with these processes interact preferentially with hypermodified DNA.
Publisher
American Society for Microbiology
Subject
Virology,Insect Science,Immunology,Microbiology
Cited by
3 articles.
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