Feb 5, 2010

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Gene silencing

RNA interference (RNAi): – a new, pioneering discovery

RNA interference is a complex molecular biological mechanism for silencing genes. In the cells of plants, fungi and animals this natural process serves to translate into proteins only those genes which the cells in question require. But cells also use the mechanism to protect themselves from foreign RNA, e.g. from viruses.

Used, but not understood.

It is just eight years since the two scientists Andrew Fire and Craig Mello discovered the mechanism of RNA interference, for which they have since been awarded the Nobel Prize.

Even before the discovery of RNA interference, a method of blocking the activity of genes was known and even used: the antisense technique. This involves introducing synthetic complementary (antisense) RNA into the cell. The sense and antisense RNA strands join to form a double strand which cannot be expressed, thereby inhibiting protein production. However, how this mechanism actually worked was still not fully understood. In their studies of nematodes (Caenorhabditis elegans) the two scientists showed that genes with double-stranded RNA (dsRNA) can be switched off more accurately than when antisense RNA was introduced. We now know that the antisense technique is based on the RNA interference mechanism.

RNA interference - a natural defence mechanism

From then on, research focused on decoding this process. In 2001 Thomas Tuschl and Sayda M. Elbashir discovered small interfering RNAs (siRNA) which are involved in this process, in mammalian cells. These are short, double-stranded sections of DNA comprising 21 to 23 base pairs. They occur during RNA interference, when long, double-stranded RNA chains are cleaved into small fragments (siRNA) by the Dicer enzyme. A further enzyme then absorbs a strand of siRNA and forms the Risc complex (RNAi-induced silencing complex). This cuts the m-RNA on the complementary segment. The cut m-RNA is very unstable and quickly breaks down, thereby inhibiting the expression of a specific gene. Since this process takes place after the transcription of DNA, it is known as post-transcriptional gene silencing (PTGS).

It has subsequently been shown that the mechanism works in almost all higher organisms. For this reason it is thought to be a natural regulatory system in cells for protection against viral attacks. Double-stranded RNA occurs during the reproduction of many viruses and is therefore recognised in the cells as a foreign body.

Understanding the processes surrounding RNA interference in plants, animals and humans is still the subject of numerous research projects today.

A powerful tool

In theory, RNA interference can be used to silence the production of every protein. This is where the enormous potential of RNA interference lies: By introducing synthetic siRNA, the sequence of which exactly matches a selected gene, this gene can be accurately silenced.

RNA interference and the antisense technique are also used in various ways in plant research and development:

• In the first genetically modified plants to be made available as food, the FlavrSavr tomato, the antisense technique was used to silence a gene that delays the ripening process in the fruit. This tomato was first launched on the market in the USA in the 1980s, but is no longer grown today.
• In plant research, RNA interference is used as a tool to obtain information about the function of individual genes through specific silencing. Scientists are focusing in particular on genes which influence growth behaviour or stress resistance.
• The antisense technique is used to produce potatoes which produce only one of the two starch components, amylose and amylopectin. This avoids the costly separation of the starches during industrial starch processing. In Europe a potato that produces amylopectin exclusively is about to be approved.
• Various approaches in safety research use RNA interference to prevent the spread of genetically modified plants, e.g. by inhibiting pollen development. If pollen development is suppressed by using RNA interference to switch off a key enzyme for pollen ripening, the plants cannot reproduce and transgenic traits will not be transferred.
• A project with apple plants aims to prevent the formation of transgenic pollen through grafting non-transgenic apple plants onto a transgenic stock. It is hoped to switch off genes in the non-transgenic apple through RNA interference from the transgenic stock. However, the results for the RNA interference with the transgenic apple plants were very contradictory, so that further investigations are necessary.
• A project with apple plants aims to prevent the formation of transgenic pollen through grafting non-transgenic apple plants onto a transgenic stock. It is hoped to switch off genes in the non-transgenic apple through RNA interference from the transgenic stock. However, the results for the RNA interference with the transgenic apple plants were very contradictory, so that further investigations are necessary.