Bacteriophages & CRISPR

1. T4: Lytic Phage Anatomy

Bacteriophage T4 of E. coli is the textbook lytic phage. ~169 kb dsDNA in an elongated head; a baseplate with six tail fibres for receptor recognition (LPS for T4); a contractile tail sheath. On binding, the sheath contracts (an ATP-independent spring-loaded mechanism), drives a hollow tube through the cell envelope, and injects the genome. The cycle: 5-min eclipse, ~25-min latent period, burst size ~200 progeny per cell. T4 makes its own DNA polymerase, primase, helicase, ligase — encoded in just ~170 kb. The economy is striking.

2. λ: The Lytic/Lysogenic Switch

Bacteriophage λ introduced biology to the concept of a stable epigenetic switch. After infection, λ chooses between two fates:

• Lytic — replicate, lyse the cell, release ~100 progeny.
• Lysogenic — integrate into the bacterial chromosome at attB (between gal and bio), become a quiescent prophage, replicate passively with the host, and reactivate later (typically on SOS-response activation).

The choice is governed by competing repressor and antirepressor circuits (CI vs Cro) at the \(P_R/P_L\) operators. Mark Ptashne’s A Genetic Switch (1986) made the λ circuit a paradigm of gene regulation; modern systems biology rederives bistability from those equations as the canonical bistable switch.

Lysogeny is not a curiosity. Cholera toxin is encoded by the CTXφ prophage; diphtheria toxin by β-corynebacteriophage; Shiga toxin by Stx phages of E. coli O157:H7. Many of the most virulent bacterial pathogens are virulent because they carry phage-encoded toxins.

3. Filamentous Phages (M13, fd)

Long thin rods (~6 nm wide, ~900 nm long), ssDNA genome ~6.4 kb, no lysis. Released continuously through the bacterial envelope by extrusion. Famous for phage display (George Smith, Greg Winter; Nobel 2018) — expressing peptide libraries on the M13 surface to select binders. The technology underlies modern therapeutic-antibody discovery (adalimumab/Humira was developed via phage display).

4. Phage Therapy

Felix d’Herelle co-discovered phages in 1917 and immediately began using them therapeutically. Phage therapy was largely abandoned in the West with the advent of antibiotics but persisted in the Soviet Union (Eliava Institute, Tbilisi). The antibiotic-resistance crisis has reawakened interest. Notable cases: Tom Patterson’s 2016 cure of multidrug-resistant Acinetobacter baumanniiby personalised phage cocktails; cystic-fibrosis lung infections by Mycobacterium abscessus treated with engineered mycobacteriophages (Hatfull lab). Regulatory frameworks for phage therapeutics are now developing in the EU and US.

5. CRISPR-Cas: Bacteria’s Adaptive Immune System

Bacteria face constant phage attack. Their adaptive defence: CRISPR (clustered regularly interspaced short palindromic repeats) and the Cas (CRISPR-associated) proteins. Three stages:

1. Adaptation — on phage infection, Cas1–Cas2 captures a short fragment (~30 bp) of phage DNA and integrates it into the CRISPR array as a new spacer.
2. Expression — the array is transcribed and processed into individual crRNAs (CRISPR RNAs), each carrying one spacer + a flanking handle.
3. Interference — the crRNA loads onto an effector protein (Cas9 in Type II; Cas12 in Type V; Cas13 targets RNA in Type VI; Cas3 + Cascade in Type I). When the loaded effector encounters DNA matching the spacer (and a PAM motif), it cleaves — destroying the phage genome.

The bacterial CRISPR array is a heritable record of past infections. Because the array preserves spacer order, ancient infections sit at the cassette’s end and recent ones at the leader — a generational immune memory.

6. CRISPR as a Tool: Doudna, Charpentier, Zhang

Jennifer Doudna and Emmanuelle Charpentier’s 2012 paper (Science, with Martin Jinek) showed Cas9 + a single guide RNA (sgRNA fusing crRNA + tracrRNA) could programme any DNA cleavage in vitro. Within months, Feng Zhang and others demonstrated CRISPR-Cas9 editing in mammalian cells. The 2020 Nobel Prize in Chemistry went to Doudna and Charpentier.

CRISPR has transformed every branch of biology. The first CRISPR-edited therapy — Casgevy (exa-cel) for sickle-cell disease — was approved by the FDA in 2023. Base editors (Liu lab, single-base C→T or A→G without double-strand break), prime editors, and CRISPR-mediated diagnostics (SHERLOCK, DETECTR for Cas13/Cas12-based detection) are extending the toolkit. The story is a particularly clean example of basic phage biology yielding therapeutic technology — a recurring pattern in the history of molecular biology.