April 13, 2021
Written by: Peter Newman
Ssssht. Door opens, storm troopers enter the air lock. Ssssht. Door closes behind them. Ssssht next door opens and storm troopers enter then next section of the death star. This is also how ABC transporters work which is the latest glyphosate resistance mechanism discovered by GRDC-funded researchers, based at AHRI at The University of Western Australia.
There’s a buzz around AHRI at the moment as former visiting PhD scholar Lang Pan and AHRI researcher Qin Yu and others have recently had a paper accepted into the prestigious PNAS journal of the USA. They have achieved this because:
- The researchers have discovered a new, world first, glyphosate resistance mechanism, and
- The resistance mechanism is the same as the way cancer patients develop multi-drug resistant cancer.
This mechanism works by plant cells pumping glyphosate out of the cell into the space between plant cells where it has no activity. The resistant plants have lots of ABC transporters in their cell membrane that grab glyphosate molecules and actively push them out of the cells.
A similar phenomenon has been observed by cancer researchers and doctors. When a patient with a cancer tumour is treated with chemotherapy, the drugs attack the cancer cells and the tumour reduces. All can look well for a while and then sometimes the tumour grows back. It turns out that some of the cancer cells have extra ABC transporters on the cell membrane that pump the cancer drugs out of the cells. These cells survive and the tumour grows back. Now we have a cancer tumour made up of cells with extra ABC transporters, rendering them resistant to a range of cancer drugs. As with resistant weeds, there can be other mechanisms at play that cause multi-drug resistance.
It’s exciting when high level scientists working in disparate fields have their research overlap, allowing them to learn from one other.
Dr Lang Pan was a former visiting PhD student at AHRI, and now Professor at Hunan Agricultural University, China.
What’s better than one world first in a single weed population? Two world firsts!
In 2019 we reported on a population of glyphosate resistant awnless barnyard grass (Echinochloa colona also known as jungle rice) that was discovered near Kununurra, Western Australia which was the world’s first example of metabolic glyphosate resistance.
This same population of barnyard grass is the subject of this research.
Dr Lang Pan when he was completing his research at AHRI in 2017 and Principal Research Fellow, Qin Yu.
RNA seq
The researchers started out using RNA sequencing and found two ABC transporter genes that were over expressed in the resistant population. Over expression simply means as the name suggests – these genes are more common in the resistant plants resulting in plants that have more ABC transporters on their cell membrane.
Transgenic rice
The researchers then created transgenic rice that is identical to normal rice, apart from the over expression of these ABC genes. They then sprayed this rice with glyphosate and found that one of the ABC genes caused resistance. Specifically, it was the EcABCC8 transporter gene.
The diagram below shows the survival of this transgenic rice to glyphosate applied at 540 g ai/ha (on the right) compared to the control plants in the front row.
Transgenic rice over expresses EcABCC8.
As well as rice, the researchers also created transgenic maize and soybean that was resistant to glyphosate through over expression of the EcABCC8 transporter gene.
Burnt leaf tips
Moving glyphosate into the apoplast via these ABCC transporters leaves more glyphosate available for upward movement in the transpiration stream, accumulating glyphosate in the leaf tips. The transgenic rice with increased ABCC transporters where glyphosate was applied had burnt leaf tips compared to the control plants where whole leaf damage was observed.
Making plants more susceptible
In contrast to ABCC8 over-expression, the researchers then took the next step of using CRISPR/Cas9- mediated knockout to see if this enhances rice sensitivity to glyphosate. This is pretty tricky stuff. Essentially, the researchers used CRISPR technology to create a rice plant that has lower than normal ABCC8 transporters on the cell membrane and this increased the susceptibility to glyphosate. So, this technique further proves that ABCC8 is a player for glyphosate resistance.
What are ABC transporters?
They are proteins that are located on cell membranes. ABC transporters help move certain compounds across the cell membrane. Some compounds can simply diffuse across the cell membrane but others need help. This is called active transport. The ABC transporter recognises the compound (in this case glyphosate) and binds to it, then pushes it across the cell membrane with a “power stroke” energised by ATP. Plants have about 120 to 140 different ABC transporters divided into the families ABCA, ABCB, ABCC and so on with many sub families.
In reality, they look a little more like this.
ABC transporter diagram.
Pumping glyphosate out of the cell
Glyphosate enters plant cells through diffusion and floats around in the cytoplasm. The cytoplasm is the ‘soup’ that all of the cell organelles are swimming around in. Glyphosate needs to get into the plastids to find its target enzyme, EPSPS. The pH of the cytoplasm is about 7 (neutral) and glyphosate gets trapped as it disassociates into anions so it is unable to simply diffuse out of the cell. This is why it takes active transport (e.g. from an ABCC transporter) to move it out of the cell. The ABCC transporters sit on the cell membrane and push the glyphosate into the apoplast. The apoplast is the space between the cells that is commonly used to transport liquids and gases. By pushing the glyphosate molecules into the apoplast the concentration of glyphosate in the cytoplasm is reduced and the effectiveness of the herbicide is reduced.
Cytoplasm.
Summary
We are very excited to have this paper accepted into the prestigious journal PNAS (Proceedings of the National Academy of Science of the USA) and to discover that a resistance mechanism in weeds has similarities to that of multi-drug resistance in cancer treatment. This research was made possible by an agreement with the Chinese government to host Chinese students at AHRI and we celebrate the success of student Lang Pan and others in this great achievement.
Paper
Media Enquiries
Paper Author Emeritus Professor Stephen Powles is available for interviews for radio, print and television.
Mobile: 0418 927 181
E-mail: stephen.powles@uwa.edu.au
Posted in: AHRI Insight, Gene discovery for herbicide resistance