LITTLE NINJA microProtein is a new tool for biotechnologists

Environment and sustainability 30. jan 2021 3 min Associate Professor Stephan Wenkel Written by Kristian Sjøgren

Researchers have identified a microProtein that can be used to make crops shorter and bushier and enhance the production of side shoots. This microProtein has the potential to increase crop yields.

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Feeding the future population of the world requires optimally exploiting all the food resources available on the planet. This is especially true for crops such as wheat, oats, rye and all the other grass crops, which together account for a very large proportion of global food production.

Now researchers from the University of Copenhagen have identified LITTLE NINJA, a microProtein that regulates how tall grasses become and how many side shoots they produce. LITTLE NINJA thus also has the potential to regulate how productive these grasses are.

The discovery of LITTLE NINJA gives biotechnologists an extra tool for designing the crops of the future.

“We have identified a microProtein that determines the growth phenotype of plants and helps to regulate a plant’s hormone signalling. LITTLE NINJA has expanded our understanding of what microProteins can do and how we can utilize them biotechnologically,” explains Stephan Wenkel, Associate Professor, Copenhagen Plant Science Centre and Department of Plant and Environmental Sciences, University of Copenhagen.

The research has been published in the Proceedings of the National Academy of Sciences of the United States of America.

Evolution has created microProteins

Understanding the new research results requires understanding what microProteins like LITTLE NINJA are.

MicroProteins are short single-domain proteins that are sequence-related to larger, often multidomain proteins.

During the course of evolution, the size of genomes has increased by processes such as whole-genome duplications or other local amplifications. This has led to increases in the number of genes and the origin of gene families that encode similar proteins. However, in some cases, evolution has also caused some genes to be significantly shortened in the process, resulting in “gene stumps” that produce very small proteins. These microProteins can take on new roles as regulators of the original larger proteins.

This new microProtein may have functions that differ from those of the original larger protein. For example, the larger protein may have a vital function for the organism and might act by binding together in pairs. In this case, the microProtein can bind to the larger protein and modulate its effect.

“In some cases, microProteins can act on the larger proteins like brakes in a car, maintaining the effect of the proteins within optimal levels,” says Stephan Wenkel.

Discovered LITTLE NINJA in a model crop plant

The researchers worked with Brachypodium, a genus of plants in the grass family that is a model organism for grasses. Brachypodium is used in biotechnological research that advances researchers’ knowledge on the genetics of crops that farmers have in their fields.

The researchers set out to discover microProteins in Brachypodium and study whether these microProteins could have useful biotechnological applications in crop plants.

This was done in a computational approach in which the entire genome of Brachypodium was searched for gene sequences that could be identified as stump versions of known proteins.

This exercise resulted in the identification of LITTLE NINJA.

The researchers’ further studies also identified that LITTLE NINJA is a dominant regulator of the jasmonic acid hormone signalling pathway in grasses.

More LITTLE NINJA can make plants more productive

Having identified LITTLE NINJA, the researchers could begin to manipulate the gene encoding for LITTLE NINJA in various crop plants to see how this affects plant growth and function.

The researchers made barley and rice and the model organism Arabidopsis produce more LITTLE NINJA than usual, resulting in shorter and bushier plants with enhanced production of side shoots.

“This is a desired feature from a crop science perspective. Grasses that are shorter and bushier are more wind-resistant. Enhanced production of side shoots also means that they can have more flowers and thus more fruit. Increasing the levels of LITTLE NINJA in grasses might ultimately lead to higher yields,” explains Stephan Wenkel.

Stephan Wenkel further explains that LITTLE NINJA plays a role in hormone signalling in grasses and therefore plays a role in developing flowers and root systems.

An extra tool for biotechnologists

The new results also bring something new to the biotechnological table.

The researchers used CRISPR gene-editing technology to cut a piece of the gene encoding the original larger NINJA protein and transform it into LITTLE NINJA.

Many specific genes have multiple copies of the genetic code, and researchers may be able to transform the code for a larger protein into the code for a microProtein, thereby giving plants new or enhanced biological functions.

“This is the biotechnological perspective. In the past, we did not fully understand how microProteins affect plants and how we can utilize them biotechnologically. We understand that much better now,” says Stephan Wenkel.

Heterologous microProtein expression identifies LITTLE NINJA, a dominant regulator of jasmonic acid signaling” has been published in the Proceedings of the National Academy of Sciences of the United States of America. In 2018, the Novo Nordisk Foundation awarded a grant to Stephan Wenkel for the project Increasing Folate Levels in Crop Plants by MicroProtein Engineering.

In my research group, we are interested in how plants adjust their development to environmental signals. Currently we work on two main topics: Many...

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