The future of bacteriophage biology

Abstract

After an illustrious history as one of the primary tools that established the foundations of molecular biology, bacteriophage research is now undergoing a renaissance in which the primary focus is on the phages themselves rather than the molecular mechanisms that they explain. Studies of the evolution of phages and their role in natural ecosystems are flourishing. Practical questions, such as how to use phages to combat human diseases that are caused by bacteria, how to eradicate phage pests in the food industry and what role they have in the causation of human diseases, are receiving increased attention. Phages are also useful in the deeper exploration of basic molecular and biophysical questions.

Figure 1. Life cycle of the typical temperate phage coliphage-λ.

The phage particle encounters and attaches to the cell surface by the tip of its tail and phage DNA enters, which leaves an empty protein shell attached to the outside of the cell. Next, the ends of the linear DNA molecule join to form a circle. The point of closure is called the cohesive site (cos). In some infected cells, the DNA is transcribed, translated and replicated. There are two pathways of replication: θ-form and rolling circle. Rolling-circle replication generates multigenomic tails of linear double-stranded DNA (dsDNA) from which DNA is drawn into preformed protein shells; tails are added and the cell lyses to liberate a crop of phage progeny. In other infected cells, phage development is repressed and phage DNA integrates into the bacterial chromosome. The resulting lysogenic cell can replicate indefinitely, but can be induced to return to the lytic cycle with the excision of phage DNA from the chromosome.

Figure 2. The φ29 DNA-packaging mechanism.

An illustration of one cycle in the mechanism that rotates the connector and translates the DNA into the head. The view down the connector axis (top row) is towards the head, whereas the bottom row shows side views that correspond to panel b. Of the twelve subunits (A–L) of the connector, eleven are shown in green, and the one 'active' monomer is shown in red. The connector is represented as a set of small spheres at the narrow end and a set of larger spheres at the wide end, which are connected by a line that represents the central helical region. The plasmid RNA (pRNA)–ATPase complexes (I–V), which surround the narrow end, are shown by a set of four blue spheres and one red sphere. The DNA base that is aligned with the active connector monomer is also shown in red. a | The active pRNA–ATPase I interacts with the adjacent connector monomer (A), which in turn contacts the aligned DNA base. b | The narrow end of the connector has moved anti-clockwise by 12 °, to place the narrow end of monomer C opposite ATPase II, the next ATPase to be fired; this causes the connector to expand lengthwise by slightly altering the angle of the helices in the central domain (white arrow with asterisk). c | The wide end of the connector has followed the narrow end, as the connector relaxes and contracts (white arrow with two asterisks), thereby causing the DNA to be translated into the phage head. For the next cycle, ATPase II is activated, which causes the connector to be rotated by another 12 °, and so on. Figure reproduced with permission from Ref. .

Figure 3. The phage-λ lysis–lysogeny decision circuit.

a | In the bacteriophage life cycle (Fig. 1), the phage can either enter the lytic cyle — in which it reproduces and then lyses its host — or the lysogenic cycle — in which the phage DNA is incorporated into the host DNA and is replicated indefinitely. Dashed boxes enclose operator sites that comprise a promoter-control complex. The three operator sites O_R1–3 of the 'λ-switch' control the promoters P_RM and P_R. Cro and CI dimers bind to the three sites with different affinities and in the opposite order to control the activation of the P_RM and P_R promoters. If protein N is available, transcribing RNA polymerases (RNAPs) can be ANTI-TERMINATED at the NUT_R and NUT_L sites; the termination sites T_R1 and T_L1 are inoperative for anti-terminated RNAPs. The CI dimer acts as either a repressor or activator of the promoter P_RM, depending on its concentration. P1 and P2 are proteases that degrade the CII phage protein. CIII is a phage gene product that is also a substrate for P1. By binding to P1, CIII inhibits the degradation of CII. b | λ-decision-circuit DNA organization. Phage-encoded genetic elements of the decision circuit are located in a 5,000-nucleotide region of the phage DNA. Genes are separated into leftward and rightward transcribed strands as indicated by the arrows. Rightward extensions of the anti-terminated P_R transcript transcribe the O and P genes that are essential for phage genome replication, and the Q gene that controls the transcription of later genes on the lytic pathway. Leftward extension of the anti-terminated P_L transcript transcribes xis and int genes, which are essential for phage-chromosome integration and excision both into and out of the host chromosome. The locations of four termination sites are indicated by T_R1–2 and T_L1–2. Figure modified with permission from Ref. .