Male sterility

Plants without pollen

Plants disperse their genes via pollen. Targeted suppression of pollen formation could be of particular interest for plants intended for the production of industrial or pharmaceutical substances where gene dispersal is undesirable. Various genetic approaches have been developed to produce ‘male-sterile’ plants, i.e. plants without pollen.

No pollen, no spread: Pollen occurs in the male inflorescence, the filaments (anthers). If pollen production is prevented, the plants are male-sterile - they can no longer reproduce.

Male-sterile plant years have been used in plant breeding for decades. They are unable to produce fertile pollen and are used as mother plants in hybrid breeding for seed production. The aim is to achieve controlled pollination through selected father lines on the male-sterile mother lines. Cytoplasmic male sterility (CMS) is used primarily in maize breeding. A current biosafety research project is investigating whether this naturally occurring male sterility is suitable for biological confinement.

In addition, there are various biotechnology approaches for suppressing pollen formation. All approaches involve combining two methodological steps:

  • A gene present in the plant, the activity of which is required for the function or development of cells (or pollen), is ‘switched off’. It could also be possible to introduce a new gene which disrupts pollen formation in the cell.
  • To ensure that this effect is limited exclusively to the cells involved in pollen production, the relevant gene constructs are linked to tissue-specific promotors. These cause the gene activity to be triggered only in specific cell types.

Male-sterile plants were developed back in the 1990s by PGS (Plant Genetic Systems), a Belgian company, using the barnase-barstar system. At the time, the idea was to be able to producehybrid seed more simply using sterile plants. Genetically modified oilseed rape developed using this concept has been grown in North America for years. In Europe, an oilseed rape line, MS8/RF3, is currently authorised that contains the PGS sterility system in addition to herbicide reisistance to glufosinate.

Example 1: Switching off an anther-specific enzyme

One genetic engineering approach is based on inhibition of the enzyme invertase, which is essential for pollen development. This anther-specific invertase breaks down sucrose (household sugar, a disaccharide) into the monosaccharides fructose and glucose, two important carbon sources for pollen development. Without the breakdown of sucrose, the pollen is deprived of its food source and is unable to develop fully: the plant is sterile.

Two methods can be used to limit or switch off invertase activity: The gene responsible for invertase can be switched off using the antisense technique or, alternatively, the enzyme activity can be directly blocked using an invertase inhibitor.

In a biosafety research project that finished in 2008, the two methods described were applied to oilseed rape and maize. In the case of oilseed rape, the project succeeded in producing male-sterile plants.

Variant: Antisense inhibition

This uses the naturally occurring mechanism of RNA interference. The genetic blueprint for the production of a protein (in this case the enzyme invertase) is expressed at the corresponding site in the genome and a copy (in this case mRNA, blue) is sent to the ribosomes, the protein factories in the cells. This process can be suppressed using special gene constructs (in this case antisense DNA, red). To ensure that this occurs only in cells responsible for pollen development, the antisense construct, e.g. the invertase gene, is equipped with a tissue-specific promoter.

Variant: Enzyme inhibitors

Enzyme activity can be inhibited by substances that have a chemical structure similar to the actual substrate – they too can bind to the enzyme, but are not processed. This means that the relevant enzyme can be individually blocked using specific inhibitors. To suppress pollen development, for instance, the pollen-specific invertase (blue) can be blocked with a substrate resembling sucrose (red). If the gene for the invertase inhibitor is introduced into a plant, it produces the inhibitor which then blocks the invertase: pollen formation is suppressed.

Example 2: Targeted expression of cytotoxins

Targeted expression of cytotoxins, e.g. only in the anthers, can also be a very effective way of suppressing pollen formation. Sterility systems based on a gene from the bacterium Bacillus amyloliquefaciens have been developed successfully in a number of different crops. The toxic effect is triggered by a special enzyme (barnase), which cuts up the RNA molecules in the cells. To ensure that only the male flower parts are affected, the toxin gene is linked to a genetic switch (promoter). This switch activates the gene only in certain cells – the ones needed for the development of the male part of the flower – and ensures that the anthers do not produce pollen.

Plants without pollen

Building on a precursor project, a current biosafety research project is testing various gene constructs for suppressing pollen formation in aspens based on cytotoxins. The barnase gene is one of the genes being used.