Dying to Live

Studying senescence to increase grain quality in wheat

Jemima Brinton - MECEA 2017 Laureate
Sophie Harrington - MECEA 2019 Laureate

When considering how to increase grain quality in wheat, it might seem counterintuitive to study how wheat dies. After all, it is during photosynthesis that sugars are produced, and nutrients and proteins build up in the green leaves. Yet the process of senescence, by which the photosynthetic tissues of the plant die, is essential in establishing the levels of protein and nutrients in the grain. This is because it is during senescence that large molecules are broken down into smaller parts which are then moved, or remobilised, into the developing grain. Importantly, this process needs to be very tightly controlled to make sure that the breakdown and movement of the nutrients occurs at the best time during the development of the grain. My PhD has focused on understanding how the onset of senescence is regulated at the molecular level in wheat, in the hope that by understanding how senescence is regulated we can also better understand how grain quality is controlled.

While we don’t know much about senescence regulation in wheat yet, we do know that a specific group of genes, the NAC transcription factors (TFs), play an important role in the progression of senescence. Two NAC TFs, NAM-B1 and NAC-S, are known to function as positive regulators of senescence [1,2]. More recently, a study of gene expression during senescence found that many NAC TFs are upregulated during senescence [3]. During my PhD, alongside my work on senescence, I was involved in characterising all the different NAC TFs that are present in wheat [4] and have also investigated the specific properties of protein domains unique to the NAC TFs [5].

senescence figure 1
Figure 1: Senescence involves the breakdown of molecules in the green, photosynthetic tissues such as sugars and proteins, and their movement, or remobilisation, into the developing grain.

I initially focussed on NAM-B1, a gene which positively regulates senescence, and which leads to increased grain protein and nutrient content [1]. While NAM-B1 is not functional in modern cultivars, its homoeologs (the NAM genes) also positively regulate senescence. I wanted to know at what point NAM-B1 can induce senescence in the plant—could it cause a seedling to die, if it was expressed earlier in development? To test this, I adapted a transgenic system for use in wheat which lets us turn on the expression of NAM-B1 at any point in development in an irreversible manner. Surprisingly, we found that even if NAM-B1 expression was turned on just after germination it had no effect on the timing of plant senescence. This indicated that, although NAM-B1 is required for senescence, it is not sufficient on its own to induce senescence prematurely.

This result suggested that NAM-B1 may need to interact with another protein, perhaps another NAC TF, to induce senescence. To test this, we carried out an experiment which allowed us to identify proteins that can interact with NAM-B1. One of the proteins we identified is encoded by a different NAC TF, which we called NAC3. Excitingly, we found that mutants in NAC3 and its homoeolog [6] lead to delayed senescence. This suggests that NAC3 is also a positive regulator of senescence. Could NAC3 be working in tandem with the NAM genes to regulate senescence?

senescence figure 2
Figure 2: Mutants in the transcription factor NAC3 (green, right) are significantly delayed in the onset of senescence compared to wild-type plants (purple, left)

To investigate this further, I used our recently-developed Genie3 network, which predicts downstream targets of TFs [7]. After confirming that the Genie3 network provides biologically relevant predictions, using an independent set of RNA-Seq data [8], I compared the predicted targets of NAM and NAC3. We found that the two TFs share more targets than expected by chance, supporting the hypothesis that NAM and NAC3 may act together to regulate senescence [9].

Moving forward, we aim to combine the information gleaned from the Genie3 network with an analysis of RNA-Seq data from the NAC3 mutants to identify genes regulated by NAC3. We are also developing crosses between mutants in the NAM and the NAC3 genes to investigate whether the genes are acting in the same regulatory pathway. And, crucially, we are investigating what role the NAC3 genes may have on affecting grain quality. This work provides a foundation for further studies into the molecular mechanisms underpinning the regulation of senescence. Techniques such as those here can be applied in concert with more traditional map-based cloning methods to accelerate the identification of regulatory candidates for senescence, grain quality, and other complex traits in wheat.

Sophie Harrington - MECEA 2019 Laureate - JIC Profile, Google Scholar, @sa_harrington

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