From Twitching Worms to Nonbrowning Apples

The tiny worm’s twitch was hardly noticeable, but with that slight shudder science took a giant leap!  A leap big enough to lead to a Nobel Prize that would pave the way to apples that will not brown, onions that will not make you cry, cotton seeds that you can eat and diseases that you can treat.

The 2006 Nobel Prize in Physiology and Medicine was awarded to Professors Andrew Fire of Stanford University and Craig Mello of the University of Massachusetts for their discovery of “RNA interference” and its role in “gene silencing.”  Genes are those segments of the “master molecule of life,” DNA, that speak, but not with words.  Their language is expressed in molecules, specifically ones known as “messenger RNA” or “mRNA.”  The message they carry is the set of instructions for the construction of proteins. 

Life is all about proteins.  Not only are these molecules the building blocks of our tissues, they make up the antibodies that protect us from disease, the receptors that allow cells to communicate with each other, and the enzymes that catalyze virtually every reaction that goes on in our bodies.  But how do cells know which proteins to make?  That’s where the 30,000 or so genes dispersed along the strands of DNA come in.  Each gene holds the instructions for making a particular protein, but the problem is that proteins are synthesized not in the nucleus but in the cytoplasm of a cell.  How then does the message get from the DNA in the nucleus to the protein-making machinery in the cytoplasm?  By means of the appropriately named, “messenger RNA.”  If this process is interfered with, the protein the gene codes for doesn’t get made and the gene is said to have been silenced.

Now back to our little nematode worms.  Some of these creatures make twitching movements because they lack a protein needed for proper muscle function as a result of having a non-functional gene.  Fire and Mello’s breakthrough discovery involved making normal worms twitch by “silencing” the appropriate gene through injection of a special type of RNA (double-stranded RNA).  It turns out that if this tailor-made RNA matches the genetic code of a specific messenger RNA, it will inactivate it, thereby essentially silencing the gene that triggered the formation of that particular messenger RNA. 

Subsequent research showed that this RNA interference machinery can be activated in yet another fashion, without the introduction of any double stranded RNA from the outside.  Sometimes, for proper functioning of our bodies, the synthesis of certain proteins needs to be suppressed, that is, some genes have to be silenced.  Cells accomplish this through making double stranded RNA via an intermediary known as micro-RNA, which in turn is synthesized on instructions encoded in the cells’ DNA.  In other words, DNA contains genes that can silence other genes through RNA interference.

Now with the difficult theoretical stuff out of the way, let’s get down to some practicalities.  The world has no need to remedy muscular problems in worms, but how about producing apples that do not turn brown?  At first this may seem like a frivolous application of RNA interference, but that is not necessarily the case.  A Canadian biotechnology company, Okanagan Specialty Fruits (OSF), has developed a nonbrowning apple by silencing a gene that codes for an enzyme known as polyphenoloxidase (PPO). 

When an apple’s cells are ruptured by bruising, slicing or biting, PPO and oxygen from the air combine with naturally occurring phenols in the apple to trigger a chemical reaction that forms melanin, a brown substance that is thought to protect the apple from attack by microbes.  But the brown discoloration is unappetizing and often results in apples being discarded.  The traditional way of preventing such browning is with lemon or pineapple juice, the acidity of which inactivates polyphenoloxidase.  Commercially packaged apple slices are usually dipped in an antioxidant solution of calcium ascorbate.  Genetically modified apples that do not brown would not require either treatment.  And sliced apples that do not brown would avoid the “yuck factor” and make for a healthy addition to children’s lunches.  Any method that allows for greater apple consumption is attractive.

The exact fashion in which the “Arctic apple,” as it will be known, is genetically modified, is proprietary information, but it is accomplished through RNA interference.  Here is a possible way.  Some apples are naturally very low in polyphenoloxidase because they express a gene that codes for the double stranded RNA that in turn silences the PPO gene.  Through standard genetic modification methods this silencing gene can be copied and inserted into the DNA of other apples with the result that PPO production will be silenced and the apples will not turn brown.

Not everyone is thrilled by the possibility of genetically altering apples in this fashion.  Organic growers worry that pollen from the modified apple trees will spread to their orchard, potentially causing them to lose their organic status.  Okanagan Specialty Fruits argues that apple pollen does not blow around easily and the chance of spreading to a neighbouring orchard is slim.  Some critics, particularly anti GMO activists, have suggested that silencing the PPO gene may have unintended negative consequences, but there is no evidence for this.  That comes as no surprise because there are no novel proteins being formed.  Field trials have shown that the modified apples are like all other apples except that they do not turn brown.

Using RNA interference technology, the lachrymatory factor synthase gene in onions can be silenced so that the nutritional qualities of this vegetable can be enjoyed without weeping.  And how about cotton seed?  The world produces some 44 million tons of high-protein seed every year that cannot be eaten because it contains the poisonous compound gossypol.  Using RNA interference, the gossypol producing gene can be silenced and enough protein to meet the daily requirements of half a billion people can be produced.  But perhaps the most alluring potential of RNA interference lies in tackling genetic diseases.  There have already been some preliminary successes, albeit only in mice, with silencing genes that code for toxic proteins such as the ones found to be present in Huntingdon’s disease, as well as in silencing genes that cause high cholesterol levels. 

In the meantime, the Canadian Food Inspection Agency is considering an application to market the “Arctic apple.”  Whether it makes it to market or not, there is no doubt that the journey from twitching worms to nonbrowning apples has been a fascinating one!  Let’s hope we won’t end up with worms in the apple by silencing the polyphenoloxidase gene.

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Print | posted on Saturday, July 07, 2012 4:09 PM

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