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1.
Early in infection by bacteriophage T4, before replication has commenced, one can detect the presence of newly synthesized DNA which cosediments with parental phage DNA on sucrose gradients. As shown earlier (R. E. Murray and C. K. Mathews, 1969), some of this represents covalent attachment of new material to parental phage DNA molecules. However, as shown herein, most of it is bacterial DNA, which is synthesized after infection and presumably degraded to T4 DNA-sized pieces. The small amount of phage-specific DNA synthesis which occurs is apparently a repair process, for its extent is greatly increased if the phage are irradiated with ultraviolet light prior to infection. Analysis by means of pulse labeling with [(3)H]thymidine and DNA-DNA hybridization shows that host DNA synthesis continues at a significant rate (40 to 80% of the preinfection rate) as late as 10 min after infection at 37 C. Very early in infection this is primarily replicative synthesis, but later a repair process predominates. Presumably this represents attempted repair of damage being inflicted on host DNA by phage-coded nucleases.  相似文献   

2.
The synthesis of membrane protein after infection with bacteriophage T4 was examined. Protein constituents of both the cytoplasmic and outer membrane are made during the infective cycle. In addition, newly synthesized membrane protein is found in material which has a buoyant density greater than that of either of the two host membrane fractions. Polyacrylamide gel analyses and solubilization studies using the detergent Sarkosyl indicate that synthesis of most of the membrane proteins made during the first 5 min of infection is directed by bacterial genes. New membrane proteins synthesized at times greater than 6 min after infection appear to be distinct from those of the host, and new proteins of the outer membrane are different from those of the inner. Proteins in the new dense membrane fraction are similar to those of the outer membrane.  相似文献   

3.
4.
Cryptic Mutants of Bacteriophage T4   总被引:1,自引:0,他引:1       下载免费PDF全文
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5.
Proteins that associate with cellular membrane during the first 5 min after infection with bacteriophage T4 were examined. Several procedures, including electrophoretic separations in three sodium dodecyl sulfate polyacrylamide gel systems and inhibition of host protein synthesis by UV irradiation, were employed to distinguish host-specified proteins from those induced by T4. Residual host protein synthesis was found to account for much of the new protein in preparations of the total membrane and for almost all of the newly synthesized protein in the outer membrane. Preliminary evidence indicates that the synthesis of some host membrane proteins is shut off less rapidly than is host synthesis of soluble protein. One host-directed polypeptide of the outer membrane was unique in that its synthesis or incorporation into the membrane was preferentially inhibited by infection. Also, it was found that the detergent Sarkosyl solubilizes all early T4 membrane proteins; this observation provides the basis for a simple procedure for distinguishing phage proteins from host outer membrane proteins.  相似文献   

6.
The DNA-Delay Mutants of Bacteriophage T4   总被引:10,自引:6,他引:10  
Mutants of phage T4 defective in genes 39, 52, 58-61, and 60 (the DNA delay or DD genes) are characterized by a delay in phage DNA synthesis during infection of a nonpermissive Escherichia coli host. Amber (am) mutants defective in these genes yield burst sizes varying from 30 to 110 at 37 C in E. coli lacking an am suppressor. It was found that when DD am mutants are grown on a non-permissive host at 25 C, rather than at 37 C, phage yield is reduced on the average 61-fold. At 25 C incorporation of labeled thymidine into phage DNA is also reduced to 3 to 10% of wild-type levels. Mutants defective in the DD genes were found to promote increased recombination as well as increased base substitution and addition-deletion mutation. These observations indicate that the products of the DD genes are necessary for normal DNA synthesis. The multiplication of the DD am mutants on an Su host at 37 C is about 50-fold inhibited if prior to infection the host cells were grown at 25 C. This suggests that a compensating host function allows multiplication of DD am mutants at 37 C in the Su host, and that this function is active in cells grown at 37 C prior to infection, but is inactive when the prior growth is at 25 C. Further results are described which suggest that the products of genes 52, 60, and 39 as well as a host product interact with each other.  相似文献   

7.
The folded genome of Escherichia coli is converted to a slower-sedimenting form within 5 min after infection with bacteriophage T4 or T4nd28(den A)-amN82(44). Chloramphenicol sensitivity and response to UV-irradiation of the phage suggest participation of viral-induced functions.  相似文献   

8.
The degradation of bacterial deoxyribonucleic acid (DNA) was studied after infection of Escherichia coli B with DNA-negative amber mutants of bacteriophage T7. Degradation occurred in three stages. (i) Release of the DNA from a rapidly sedimenting cellular structure occurred between 5 and 6 min after infection. (ii) The DNA was cleaved endonucleolytically to fragments having a molecular weight of about 2 x 10(6) between 6 and 10 min after infection. (iii) These fragments of DNA were reduced to acid-soluble products between 7.5 and 15 min after infection. Stage 1 did not occur in the absence of the gene 1 product (ribonucleic acid polymerase sigma factor), stage 2 did not occur in the absence of the gene 3 product (phage T7-induced endonuclease), and stage 3 did not occur in the absence of the gene 6 product.  相似文献   

9.
Mapping Experiments with R Mutants of Bacteriophage T4d   总被引:6,自引:8,他引:6       下载免费PDF全文
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10.
Simultaneous addition of T4 phage and ghosts to host cells prevents infective center formation. Cells which have been infected with phage for less than 2 min are also inhibited by superinfecting ghosts. After this time, a chloramphenicol-inhibitable reaction occurs which causes the phage-infected cells to become increasingly tolerant of added ghosts.  相似文献   

11.
12.
Endonuclease II-deficient, ligase-deficient double mutants of phage T4 induce considerably more deoxyribonucleic acid (DNA) synthesis after infection of Escherichia coli B than does the ligase-deficient single mutant. Furthermore, the double mutant can replicate 10 to 15% as well as wild-type T4, whereas the single mutant fails to replicate. When the E. coli host is also deficient in ligase, the double mutant resembles the single mutant. The results indicate that host ligase can substitute for phage ligase when the host DNA is not attacked by the phage-induced endonuclease II.  相似文献   

13.
Superinfection by phage T4 of cells infected by the ribonucleic acid (RNA) phage f2 results in inhibition of further f2 production. Experiments using rifampin show that the exclusion of f2 requires T4 gene function soon after T4 infection. By using a sensitive new peptide-mapping procedure to identify f2 coat protein in infected cells, we show that synthesis of the f2 coat occurs at a reduced level until 4 min after T4 superinfection and then ceases abruptly. Within 4 min after T4 superinfection, there are also several changes in f2 RNA metabolism, all of which require T4 gene function: preexisting f2 replicative intermediate RNA and f2 single-stranded RNA are degraded to small but still acid-precipitable fragments, and most f2-specific RNA is released from polyribosomes. We favor the hypothesis that T4 induces the synthesis of a specific endoribonuclease which degrades f2 RNA and that the inhibition of f2 protein synthesis may be a consequence of this degradation, rather than a direct effect of T4 upon translation.  相似文献   

14.
Five temperature-sensitive sporulation mutants of Bacillus cereus T have been isolated. These mutants are blocked at stage 0 of sporulation at the restrictive temperature (37 C) but are able to sporulate at nearly normal frequencies at the permissive temperature (26 C). A bacteriophage that forms a stable lysogen in the parent strain is induced at increased frequencies in the mutants. This induction is accompanied, in some of the mutants, by a reduction in immunity to the phage. Revertants, selected for their ability to sporulate normally at both temperatures, lose their ability to produce high titers of the phage. In addition to this lytic phage, an apparently defective phage has been found in lysates of the mutants. Strains cured of the plaque-forming phage still carry the defective phage. Comparisons of physical and biological properties of the plaque-forming phage with those of the two Bacillus cereus phages most similar to it have shown that this phage is not identical to either of them. The maximal titer of phage produced in cultures of the parent strain is about 10(3) plaque-forming units (PFU) per ml at both temperatures. The maximal titers of phage produced by the mutant are 4 x 10(9) PFU/ml at 37 C and 7 x 10(8) PFU/ml at 26 C. Both mutant and parent strains release over 90% of the phage they produce after the onset of stationary phase.  相似文献   

15.
J. D. Karam  M. Leach    L. J. Heere 《Genetics》1979,91(2):177-189
T4 phage completely defective in both gene 30 (DNA ligase) and the rII gene (function unknown) require at least normal levels of host-derived DNA ligase (E. coli lig gene) for growth. Viable E. coli mutant strains that harbor less than 5% of the wild-type level of bacterial ligase do not support growth of T4 doubly defective in genes 30 and rII (T4 30- rII- mutants). We describe here two classes of secondary phage mutations that permit the growth of T4 30- rII- phage on ligase-defective hosts. One class mapped in T4 gene su30 (Krylov 1972) and improved T4 30- rII- phage growth on all E. coli strains, but to varying degrees that depended on levels of residual host ligase. Another class mapped in T4 gene 32 (helix-destabilizing protein) and improved growth specifically on a host carrying the lig2 mutation, but not on a host carrying another lig- lesion (lig4). Two conclusions are drawn from the work: (1) the role of DNA ligase in essential DNA metabolic processes in T4-infected E. coli is catalytic rather than stoichiometric, and (2) the E. coli DNA ligase is capable of specific functional interactions with components of the T4 DNA replication and/or repair apparatus.  相似文献   

16.
Deoxyribonucleic acid replication and genetic recombination were investigated after infection of Escherichia coli with ligase-deficient rII bacteriophage T4D. The major observations are: (i) deoxyribonucleic acid synthesis is discontinuous, (ii) the discontinuities are more slowly repaired than in wild-type infection, (iii) host ligase is required for viability, and (iv) genetic recombination is increased.  相似文献   

17.
Synthesis of host-specific and phage-specific messenger ribonucleic acid (mRNA) was studied in bacteria infected by unmodified (T1 . B) or modified [T1 . B(P1)] bacteriophage T1. In a "standard" infection of Escherichia coli B by T1 . B (no host-controlled modification involved), the rate and amount of T1 mRNA synthesis was intermediate between those values reported for infections by a virulent phage such as T4 or a temperate phage such as lambda. The initial rate of mRNA synthesis was slightly increased after T1 . B(P1) infection of E. coli B in comparison with T1 . B infection of the same host. Little or no phage mRNA synthesis could be detected in T1 . B infection of E. coli B(P1). Phage mRNA synthesis in T1 . B(P1)-infected E. coli B(P1) cells was approximately the same in amount as that seen in T1 . B(P1) infection of E. coli B. Synthesis of host-specific mRNA continued throughout the latent period in all infections studied. However, the enzyme beta-galactosidase could not be induced, except after T1 . B infection of E. coli B(P1). In an attempt to understand the apparent differences in mRNA synthesis after infection of E. coli B by phages T1 . B or T1 . B(P1), the effect of altered T1 deoxyribonucleic acid (DNA) methylation on mRNA synthesis was studied. Methyl-deficient T1 DNA, made in cells infected with ultraviolet-irradiated phage T3, inhibited (14)C-uridine incorporation more strongly than normal T1. One passage of methyl-deficient T1 through E. coli B restored uracil incorporation rates to those seen with ordinary T1. This suggests that methylation of T1 DNA can influence the rate of phage mRNA synthesis. However, attempts to relate the difference in mRNA synthesis seen between T1 . B and T1 . B(P1) in E. coli B to the activity of the P1 modification gene were not conclusive.  相似文献   

18.
Presumptive D2a Point Mutants of Bacteriophage T4   总被引:1,自引:3,他引:1       下载免费PDF全文
Mutants of bacteriophage T4 have been isolated that show phenotypes seen previously with those rII deletions that extend into the nearby D2a region. Analysis indicates that rII mutations are not necessary in order to get the D2a phenotypes.  相似文献   

19.
The Gp59 protein of bacteriophage T4 promotes DNA replication by loading the replicative helicase, Gp41, onto replication forks and recombination intermediates. Gp59 also blocks DNA synthesis by Gp43 polymerase until Gp41 is loaded, ensuring that synthesis is tightly coupled to unwinding. The distinct polymerase blocking and helicase loading activities of Gp59 likely involve different binding interactions with DNA and protein partners. Here, we investigate how interactions of Gp59 with DNA and Gp32, the T4 single-stranded DNA (ssDNA)-binding protein, are related to these activities. A previously characterized mutant, Gp59-I87A, exhibits markedly reduced affinity for ssDNA and pseudo-fork DNA substrates. We demonstrate that on Gp32-covered ssDNA, the DNA binding defect of Gp59-I87A is not detrimental to helicase loading and translocation. In contrast, on pseudo-fork DNA the I87A mutation is detrimental to helicase loading and unwinding in the presence or absence of Gp32. Other results indicate that Gp32 binding to lagging strand ssDNA relieves the blockage of Gp43 polymerase activity by Gp59, whereas the inhibition of Gp43 exonuclease activity is maintained. Our findings suggest that Gp59-Gp32 and Gp59-DNA interactions perform separate but complementary roles in T4 DNA metabolism; Gp59-Gp32 interactions are needed to load Gp41 onto D-loops, and other nucleoprotein structures containing clusters of Gp32. Gp59-DNA interactions are needed to load Gp41 onto nascent or collapsed replication forks lacking clusters of Gp32 and to coordinate bidirectional replication from T4 origins. The dual functionalities of Gp59 allow it to promote the initiation or re-start of DNA replication from a wide variety of recombination and replication intermediates.  相似文献   

20.
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