Specialized Transduction

Transduction is the transfer of genes from one host cell to another by a virus. There are two types: generalized (which results from errors in packaging) and specialized (which results from errors in excision from the chromosome of a lysogenic bacterium). Generalized transduction is not directly related to lysogeny. It is most easily observed with temperate phages, but only because the potential transduc-tants formed by a virulent phage are generally destroyed by lysis. This lysis is not caused by the transducing particles, which usually contain only host DNA, but rather by the phage particles in the lysate.

Specialized transduction occurs only with temperate phages, and is observed only in lysates produced by inducing lytic development in a lysogen, not in lysates produced by

Figure 7.3 Formation of specialized transducing phages by bacteriophage l. The central column shows normal excision of the phage genome, mediated by integrase and excisionase. For every such excision, there are about 10~5 abnormal excisions, where either gal or bio is effectively cloned into l. These are rare because they require breakage and joining of heterologous DNA. The reciprocal product (a deleted chromosome), which may or may not be formed in the same event, is not shown. The presence of the cos site allows the excised DNA to be packaged into virions and injected into other cells. Since l is cut at cos during packaging, part of cos occurs at each end of the packaged DNA.

infection. Specialized transduction results from rare faulty excision from the lysogenic chromosome. Instead of the precise excision mediated by integrase and excisionase (figure 7-1), an unknown enzymology causes breakage and joining in heterologous DNA to produce packageable chimeric genomes, part phage and part host. This can be viewed as a natural form of gene cloning. Whereas any part of the host genome can be transferred in generalized transduction, specialized transduction is possible only for a limited portion of the host chromosome flanking the phage insertion site.

Figure 7-3 shows specialized transduction by l of some nearby genes—gal from one side and bio from the other. l gal and l bio arise in separate events. Several constraints on the process may be noted: (i) The genome of a specialized transducing phage is a connected segment of the lysogenic chromosome. Therefore, any l carrying gal will also carry all the genes between gal and attl, and any l carrying the distal galE gene must also carry galK. (ii) The amount of DNA is restricted by the packaging limits of the phage. l can package up to 110% of its 48.7 kb genome, so no gene can be packaged that is farther than that from l cos. By extension, if a l has accidentally inserted somewhere other than its normal site, it will now transduce only genes close to the new site. (iii) Specialized transducing phages are frequently missing some phage genes. As implied by figure 7-3, acquisition of host DNA to the left of the prophage is accompanied by loss of genes from the right end of the prophage. The phage genes that are eliminated may be vital for the viral life cycle, so propagation of the specialized transducing variant can take place only in cells coinfected with a complete l. (iv) The packaging site, cos, cannot be lost because it is needed in cis. Therefore, all gal-transducers will include genes int-cos, and all bio transducers will include cos-J.

The last line of figure 7-3 compares the structures of l gal and l bio DNA with that of normal l, as packaged in the virion. The termini (produced by cutting at cos) are the same. l gals have a segment of host DNA whose left margin may vary widely but whose right end is always at the phage insertion site; in l bios, the host DNA runs rightward from the insertion site.

Figure 7-4 shows insertion by the l gal specialized transducing phage. A gal may insert into the chromosome either by general, homology-dependent recombination or by site-specific recombination at att. The insertion is sometimes accompanied by insertion of a coinfecting normal l (not shown in figure 7-4). The requirements for the integrase reaction using the hybrid site attL on the l gal are different from those for a phage attP site; this generates a helping effect by the coinfecting l. The product of lysogen-ization contains two copies of gal: the indigenous copy already present before infection (shown as gal~ in figure 7-4) and the added copy from the l gal (gal+ in figure 7-4). Homology-dependent recombination within the chromosome can "loop out'' one copy or the other, so that lysogens of l gal are inherently unstable.

Insertion and excision by l gal have served as a model for replacement of genes by cloned, mutated copies. Whereas l insertion requires integrases, l gal insertion can also take place through homologous recombination with bacterial DNA. The haploid segregants can be either GaP (reversal) or Gal+ (replacement) depending on which side of gal the crossover occurs to loop out the l gal.

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