Bacteriophages and CRISPR

by Ruthie G 

Bacteria are facing a virus epidemic. A deadly one. In fact, 40% of all ocean bacteria are killed every day. There are over 90 species of bacteriophage - viruses specialised to kill bacteria. I want to explore the incredible evolutionary arms race between bacteria and phage, going back millennia - a story which culminates in the discovery of a technology rewriting human history. Modern gene editing, fundamentally altering the DNA which codes for our existence, was born from the phage and the bacterium.

First, let’s get to grips with these bacterial viruses. Phages come in all shapes and sizes. If they can pierce open a bacterium, anything goes; corkscrew, thimble or football shaped. My personal favourite, pictured above, is the creepy, spider-like phage. Once the bacterium is pierced, phages insert their own genetic material into their unfortunate victim. In organisms, the genetic material (often in the form of DNA) codes for characteristics, behaviour, and how an organism is built. So, this phage DNA holds the instructions to produce more phages. 

Finding the phage’s genetic material, the unsuspecting bacterium will assume it’s theirs and incorporate it into their own DNA. Normally, the bacterium reads its DNA for instructions on how to function. Now, when the bacterium reads it, it will be hijacked to make hundreds of new phages. The poor bacterium becomes desperately overcrowded - until it pops open, burst by the sheer number of phages. Now unleashed upon the world, these new phages go on to infect more bacteria. This cycle is an ingenious solution; bacteriophages, and all viruses, are so tiny and low-functioning that they would otherwise be unable to reproduce. Only by using other cells can they secure survival. 

Bacteria, however, are prepared to fight back. A secret weapon is tucked away inside  their DNA- a weapon with profound implications for all life on Earth. Bacteria are mounting a counterattack on the virus’ key firearm: the genetic material which they inject into their victims. If the bacteria can find and destroy this DNA before they read it and inadvertently start producing phages, they can defend themselves. At the heart of this strategy is a segment of bacterial DNA christened CRISPR.

In the CRISPR segment, the bacteria keeps snippets of foreign viral DNA. Short segments of different phage species’ DNA, called spacers, are alternated with regular repeats - identical sequences of normal, bacterial DNA. These mark out the location of each different CRISPR segment, and stop the bacterium from destroying the viral DNA as an immune response. But how does storing short records of different phage’s genetic material combat the phages’ DNA injection strategy?

Here’s the crux: little snippets of phage DNA can be used as a key to find longer segments from the same species. If a bacterium contains a CRISPR record of a species’ DNA, the complete version also contains the smaller segment inside. So, the bacteria can use the segments in CRISPR to search for and destroy any phage genes incorporated into its own DNA during an infection.

However, the bacteria first need foot soldiers. How are they supposed to hunt down viral DNA with just the instructions to find it - the little segments in CRISPR? They need something to operate on those instructions. Enter, RNA and Cas enzymes. RNA has a similar structure to DNA, but is able to copy instructions from DNA and actually move around the cell to act on them. These pair with Cas enzymes, precision cutters which can sever DNA with ease. RNA molecules copy the viral DNA sections from CRISPR and scour the cell’s DNA for a match with their particular virus. 

If they find a replica, they know they’ve located a piece of phage DNA, soon to trigger the cell’s death. The RNA locks onto the located section and the Cas enzymes take this as their trigger to cut. With this small part of the inserted phage DNA severed, the rest is rendered useless. Disaster has been averted; the phage’s dangerous weapon has been deactivated.

The story doesn’t end there, though. Humans, too, have a link to this bacterial defence. Does the phrase CRISPR sound familiar? It’s now well-known as a tool for editing human, animal, and plant DNA. What started as an igneous bacterial method of fighting off phages is now at the cutting edge of science. 

In 2012, a group of scientists made a staggering discovery. After determining the function of CRISPR systems in bacteria, they realised they could be manipulated to serve a different purpose. Fundamentally, the CRISPR set-up involves identifying a gene sequence and cutting it out, deactivating it. They realised if you simply isolate the RNA and Cas enzymes from this system (the foot soldiers), you can program them to target any gene in any organism. Instead of the bacterium’s CRISPR segment giving instructions to find phage DNA, the RNA can be altered to search for a DNA sequence you define. As a result, you could give the RNA/Cas pair instructions to knock out any DNA sequence.

In fact, it’s possible to go beyond just knocking out DNA segments, stopping the characteristics they code for being expressed. Instead, you can replace a DNA segment with a new one - simply by inserting a chosen segment of DNA into the cell. The cell doesn’t like messily rejoining severed DNA, and if a convenient piece which exactly fits into the gap created by the cut appears, it will insert that in. This allows scientists to actually program the DNA of an organism, an incredible finding. 

The impacts of CRISPR’s power are far-reaching and significant. Many argue CRISPR has the potential to change the world for the better. A striking example is genetic disorders which could be fought with CRISPR, such as cystic fibrosis and sickle cell anaemia. These are solely caused by defective sections of DNA in the genetic material of the sufferers, coding for terrible diseases; they often condemn victims to lives of pain, or even death. In the US, an estimated 41% of infant deaths are triggered by genetic disease. 

With CRISPR, the section of DNA causing the disease could be cut out, or replaced, from those born with disorders. It’s a challenge to target faulty DNA when every cell in the body holds a copy of the genetic material, so also the defect. However, there are ways to circumvent this issue, and the possibility of saving innumerable lives is exhilarating. 

However, the argument is nuanced. Many bioethicists and scientists are calling for caution using CRISPR, warning of the catastrophic consequences of moving too hastily. For example, editing of unborn embryos has been a source of debate. Even when well-intentioned, the altering of a cell like an embryo (known as a germline cell) is controversial. By editing them, any future offspring of the embryo will also be passed the altered genes. There is a risk of unintended consequences spiralling out of control. Despite this, it can be argued that  it is simply unethical to condemn unborn babies to a life with a genetic disorder.

Furthermore, there is a danger that CRISPR could be used to alter genes affecting gender, personality, and intelligence, in unborn humans - a practice which would border on eugenics. So, although the usage of CRISPR in the cells of already-born humans holds lots of potential, practices like germline editing raise difficult physiological questions.

We’ve come far from the humble bacteriophage. But, in a roundabout way, we’ve come to understand its impact. The age-old battle between phage and bacteria has driven the development of one of the most fascinating and elegant biological defences: CRISPR. In turn, CRISPR has inspired scientists of today to build a revolutionary technology changing our world. That huge phage epidemic, tearing through bacteria - it shook the very foundations of all life on Earth as well.