Australasian Science Magazine

More chloroplasts in leaf cells do not necessarily lead to more efficient photosynthesis. Credit: Martin Bahmann CC BY-SA 3.0

By John Hamblin

A scientist’s 50-year research journey is finally about to reveal not only that high chlorophyll levels don’t improve wheat yields through more efficient photosynthesis, but that the opposite may be true.

When I was an Honours student 50 years ago I was interested in the relationship between plant growth and energy capture by chlorophyll in a plant’s leaves. Using the model plant Arabidopsis I compared the growth and chlorophyll content of a wildtype line, a variegated mutant and their first generation hybrid (F1).

The advantage of using Arabidopsis was that it has a very short time from seed to plant and back to seed again. This meant that I could grow the parents, hybridise them and then grow the parents and their progeny to see how different levels of chlorophyll affected their growth – all within a term and a half at university.

The results showed that the level of chlorophyll per unit leaf weight was linearly related to the genotypes. Not surprisingly the variegated mutant parent had the lowest level of chlorophyll per unit leaf weight; the hybrid was intermediate; and the wildtype line had the highest level of chlorophyll. The increase was in a nice straight line, showing that the presence of the wildtype chlorophyll gene had what geneticists call an “additive” effect.

However, the effect on growth was completely different. The F1 hybrid was twice as big as its wildtype parent and three times greater than the variegated mutant parent. This is what geneticists call overdominance, and was surprising because one might expect that more chlorophyll would catch more light and lead to greater growth. It didn’t.

Why? My explanation at the time was that the plants had been grown in a small growth chamber with a relatively low light intensity, so the hybrid offspring had just the right level of chlorophyll to capture the available light in the growth chamber without unnecessarily consuming materials required to make more chlorophyll than was needed to achieve maximum growth. It was the “Goldilocks” situation of being “just right”.

Then I went into a career of crop breeding and agronomy followed by administration, and forgot about the thesis. During this time I and many others have spent much time pondering how to get crops to give more yield from given inputs of light, water, fertiliser etc. Then when I retired I wondered why I had kept a copy of my Honours thesis from 45 years ago and threw it out. So too had the university where I had done the project.

Oh dear, because shortly after this a colleague gave me an old copy of New Scientist (11.9.2010) containing an article entitled “Super Crops” that stated:

Plants with excess chlorophyll don’t just shade out rivals though. They also shade their own leaves. What’s more the excess chlorophyll may cause some leaves to absorb light energy faster than it can be used, which can damage them. So plants have evolved a “quenching system” to mop up the surplus energy. All this is costly for the plant, which is one big reason crops yield less food per light unit than they otherwise might. A soybean mutant with half the usual level of chlorophyll can produce 30 per cent more biomass than normal.

This immediately rang a bell for me, and on the web I tracked down Don Ort, who had made this statement to the reporter. The questions that then arose in my mind were whether I should try to repeat the now discarded experiment and, if so, how to do it better?

While Arabidopsis in now the workhorse of plant genetic studies, it does not make much of a meal. Wheat is the major cereal crop grown in Australia, and one of the big three species supplying most of the food energy and protein requirements of humanity. We also understand a great deal about light capture in wheat crops as well as many other facets of its growth and yield. With this in mind I decided to use wheat as my model plant.

Selecting for low levels of leaf chlorophyll seems counter-intuitive, so what are the possible benefits from doing this? As pointed out in the article, leaves act like umbrellas, but instead of keeping rain off they shade leaves below them, and in a crop these are primarily their own leaves deeper in the canopy. This has been known for a long time, and wheat breeders have modified the canopy architecture of plants to let more light pass through to the leaves below.

However, there has been no systematic attempt to make leaves more transparent so that any excess light above what is needed for maximum photosynthesis goes though the leaf, and thus provides more light for shaded leaves further down the plant while avoiding damage to the chloroplasts. The reduced damage to the chloroplasts in the upper leaves also improves their efficiency.

If a leaf absorbs energy faster than it can dissipate it, heat in it will build up. If a leaf gets too hot it dies. To counter this the leaves evaporate water, and just like an evaporative air-conditioner it cools down. This is fine if there is plenty of water.

However, grain filling in wheat occurs as the weather warms up in late spring and early summer. This is the same time that rainfall decreases significantly.

Active photosynthesis is required during grain filling to ensure high crop yields. This needs water, but if more water than necessary is used to prevent leaves from over-heating there is less water available for grain filling. Hence increased leaf transparency may reduce water use early in the season, leaving more available for later and thus increasing the crop’s potential yield.

At the same time chloroplasts consume valuable nutrients. If fewer are needed to achieve maximum photosynthesis, these nutrients will be available for other growth processes.

So there are at least four reasons in relation to chlorophyll levels that may lead to improved crop yields. However, to date no plant material has been specifically produced to test this hypothesis. My current work is focused on developing suitable plant material to test this idea.

The project has several stages. The first was to find wheat parents that differ consistently in their level of chlorophyll per unit leaf area. To do this I turned to both the national wheat gene bank, which supplied a wide range of varieties, as well as to active wheat breeders who supplied me with current high-yielding material.

I grew the full range of genotypes in a series of environments: in the glasshouse, in the field, in winter and summer, at high and low levels of fertiliser, and at different planting dates. The chlorophyll content of the youngest fully expanded leaf was measured over a range of dates, and the results from this initial screening showed that it was possible to identify genotypes that differ consistently and widely in their chlorophyll per unit leaf area. These results have been published in PLOS ONE (http://tinyurl.com/mcpmbzo).

With this background information it is now possible to develop suitable genotypes to look at the growth and yield of similar wheat plants that differ in their level of chlorophyll, and then study the impact of this on their yield.

The two genotypes I used are very different (Fig. 1). P1 is a reduced tillering, very large-eared modern genotype breed in Western Australia that has not yet been released to the public. It has very high levels of chlorophyll. P2 was bred more than 60 years ago in the USA as part of a program to incorporate disease resistance into wheat from wild relatives. It is prostrate, highly tillered with small grains and has only about two-thirds of the amount of chlorophyll per unit leaf area as P1.

I am now growing the two parents (P1 and P2), their first generation progeny (F1) and their second generation progeny (F2), a total of 700 plants (Fig. 2). So far I have only measured the chlorophyll contents of the parents and their F1 hybrids.

The initial results are the same as those I obtained all those years ago. The parents have different levels of chlorophyll, and their F1 progeny are intermediate between the parental values.

Several complete sets of data involving all 700 spaced plants will be analysed early this year. The data sets will cover chlorophyll levels, flowering dates, plant habit and yield components. The data will allow me to get a handle on the genetics of these characters in this cross.

At the same time 1200 true breeding lines are being developed. They will be grown in the field and selected for contrasting chlorophyll levels and agronomic acceptability. The best contrasting lines will be grown in plots over several sites in 2016, and the data will provide the initial confirmation (or otherwise) of the hypothesis that less chlorophyll leads to increased wheat yields.

I just wish I had the original data on Arabidopsis with which to compare the details of the experiments and their results.

John Hamblin is Adjunct Professor at the University of Western Australia’s Institute of Agriculture.