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Editing genes like alphabets

The journal, Nature, has selected David R Liu, a researcher at the Massachusetts Institute of Technology, Boston, as the first…

Editing genes like alphabets

DNA

The journal, Nature, has selected David R Liu, a researcher at the Massachusetts Institute of Technology, Boston, as the first of the 10 people who mattered in 2017. Liu has “developed gene-editing tools that are new to nature, and that could one day save lives”, says the write-up in the journal.

In October 2017, the journal had carried a paper by David Liu with Nicole M Gaudelli, Alexis C Komor, Holly A Rees, Michael S Packer, Ahmed H Badran and David I Bryson, working at Harvard University and the Broad Institute of MIT and Harvard, which described a technique of targeting a single component of a vast DNA molecule and making a correction in that component, without disturbing the rest of the molecule.

It is this DNA molecule, of course, which is present in the nucleus of every cell of an organism, which contains the action programme of what proteins the cell will produce, and hence the characteristics of the organism. If there are errors in that programme, naturally, cells do not produce the right proteins, and there is disease or malfunction of the organism. The DNA itself is a chain of millions of units, in the form of two complementary strands in the shape of a helix, but folded and compact, to fit inside the cell nucleus. The millions of units in the DNA are grouped in segments, which are the codes for specific proteins. The segments, in turn, consist of groups of three links in the chain, and each group of three, called a triad, codes for one of 20 possible amino acids, which are the components of proteins.

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A segment of DNA thus entails a large collection, in a specific order, of amino acids from a menu of 20 possible ones. As the segment can be long indeed, a huge sequence of amino acids, and hence a huge number of distinct proteins can be specified by the code. There is also the device of more than one coding for some amino acids, to take care of errors in the structure of the units. Errors, however, do occur and these change the specific structure of proteins, leading to disease with a genetic origin.

A promising method to repair such errors has been with the recent technique, CRISPR, which cuts the DNA at a place that can be specified by the clinician. If this cut is made at the place where an error has crept in, there is a possibility for the error not to persist when the two parts rejoin, resulting in a remedy of the genetic error.
The units that make up the triads, a sequence of which make up the segments of DNA, consists of just four basic kinds, which are named, A,T,G and C. These units occur in the two strands of the DNA in only four possible combinations — G-C or C-G and A-T or T-A. Occurrence of any one in one strand thus determines the units of the other strand, and this is the principle behind each strand being able to build up the complementary strand when a cell divides.

Now, as each unit, called a base pair, can thus be one of four kinds, a triad can be in 4x4x4=64 forms. These are the combinations of base pairs that code, with redundancy built in, for the 20 amino acids of which all proteins consist.
The CRISPR technique has derived from a system that is native to bacteria, in their defence against viruses. Viruses are little more than their own DNA and what they do to the cells that they infect is to monopolise the cells’ resources for their own proliferation. The defence that bacteria employs is to copy a segment of the virus’ DNA into its own DNA and use this template, at a subsequent virus attack, to chop the attacker at the place where the segment appears, with the help of associated segment, called the CAS (CRISPR Associated) gene. CRISPR/CAS is thus a powerful tool which makes use of a portion of a strand of DNA, along with the enzyme, CAS9, to cleave DNA at an identified place.

The authors of the paper in Nature note that about half of all known “point mutations” or single point errors in genes, which cause disease, are because a C in a base pair changes to T and the CG pair becomes a TA pair. The ability to change an AT pair back to a GC pair could thus have great value in correcting genetic features that cause disease. Carrying out such correction with the help of CRISPR/CAS9 and related paths, which involve a break in the DNA, has undesirable outcomes. As the methods only create a break at a specific location and have little control on how the two parts of a divided DNA would rejoin, there could be chance insertion or deletion of a part of the DNA, or the movement of a segment to another part of the DNA. Methods that directly target the error in the DNA structure, by converting one base pair into another, called base editing, have thus been developed.

A number methods have been developed, the paper says, to convert a C into a U (a form related to a T) and lead to permanently converting a CG base pair into a TA. This is also the change that occurs spontaneously. No methods, however, had been developed to convert AT to a GC, which would help reverse a great many CG to TA base pair changes that are associated with disease. This is the task Liu and his group, particularly Nicole Gaudelli and Holly Rees, undertook and is described in the Nature paper. While the conversion of GC to TA has used enzymes found in nature, the reverse conversion required the creation of a new and unknown enzyme in the laboratory.

The Nature citation that recognises Liu’s work says that his lab pioneered the development of new enzymes and the first methods to convert G to T in many organisms. The methods were successfully used in bacteria, yeast, plants, zebra fish, and animals and recently in China to correct a single base pair mutation in a human embryo. But there was no assurance that Liu’s team would be able to find an enzyme that could make it possible to reverse, or change TA to CG. Till the relentless efforts of Nicole Gaudelli, in Liu’s team, nobody could come up with the evasive enzyme.
The enzyme has been tried out with the base, A, in different environments and has even succeeded in correcting a genetic cause of build-up of iron in tissue and organs. The efficiency of base editing was found to average a high 53 per cent and there were no adverse effects. This advance in “single base pair editing”, could be refined to give clinicians access and control over patients’ genetic heritance.

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