Understanding mechanisms of cellular geometry scaling

All life on Earth is made of cells. Intricately organized assemblies of different types of cells make up complex multicellular organisms such as humans or plants. Simpler organisms including yeasts or protozoa live as single cells. Cells come in an astonishing diversity of shapes optimized for their specific function. A cell can grow to different size, depending on the amount of nutrients or other signals in the environment. If specific shape is important, how do cells make sure that it remains the same across different sizes? This is an interesting biological and engineering problem that can be solved by understanding how the intrinsic cellular polarity machinery that generates cell shape by restricting growth to certain sites on the cellular membrane, can decrease or increase the sizes of these growth zones in response to changes in cellular volume. We call this property cellular geometry scaling. It is incredibly widespread in biology but we know very little about it.

We will use a simple unicellular fungal organism called Schizosaccharomyces japonicus (S. japonicus) to understand how cells scale their shape. It is a great system for addressing this problem because it grows fast, is easily amenable to genetic manipulations and, of course, exhibits very robust scaling. We previously found that a regulator of the cellular polarity machinery, a protein called Rga4, was critical for scaling. If S. japonicus does not have Rga4, it cannot scale down when nutrients become scarce and dies, demonstrating that the ability to scale is really important. Now we propose to obtain mechanistic explanation for Rga4 function in scaling. Our program will consist of two interrelated objectives. First, we will understand how cellular polarity changes during scaling and how Rga4 contributes to these changes. We will also probe the relationship between Rga4 and a protein called Tea4 that normally functions to landmark growth at specific membrane sites. In the second objective we will investigate how cells regulate Rga4 by phosphorylation to promote scaling when required. Phosphorylation is one of the most important modifications regulating protein function through changes in conformation and we will additionally look at phosphoregulation of many more cellular proteins to reveal hidden connections between different parts of cellular physiology important for scaling.

It is essential to do this type of fundamental research in a simple organism because it provides important insights into molecular underpinnings of polarized growth and scaling in all eukaryotic cells, from humans to plants. The answers we get may open entirely new possibilities in dealing with devastating fungal pathogens, which rely on changing their size and shape to infect their hosts.

For cells of a specific type, functionally optimal cellular organization is typically maintained across a range of cell sizes. This phenomenon, known as scaling, is widespread in biology but surprisingly poorly understood. We want to understand the molecular mechanisms underlying scaling of cellular polarity, which allows cells to maintain their optimal shape at different cell volumes.

We recently established the fission yeast Schizosaccharomyces japonicus as a genetically tractable model system for understanding both the mechanisms and functional implications of cellular geometry scaling. We discovered that the GTPase activating protein Rga4 was essential for this process. In the proposed work we will test the hypothesis, built on our preliminary data, that scaling requires modulation of Rga4 function by a polarity landmark protein Tea4 and specific phosphoregulation events.

We will pursue two interrelated objectives. First, we will examine contribution of Rga4 to regulating Cdc42 and Rho2 GTPase pathways during scaling and address the molecular function of a cellular polarity landmark Tea4 in regulating Rga4. This set of experiments will allow us to identify the target(s) of Rga4 critical for scaling and interpret phenotypes of mutants obtained in Objective 2. In the second objective, we will focus on identifying specific Rga4 phosphoregulation events key to scaling. To gain insight into the broader regulatory context of scaling and reveal possible epistatic relationships, we will also carry out unbiased phosphoproteomics analyses of scaling. These data will further inform our studies on Rga4 regulation and function.

All in all, the proposed research will lead to discovery of new fundamental mechanisms underlying cell-size dependent morphogenetic plasticity in eukaryotic cells and inform our thinking of the underlying biological and engineering principles of cellular organization.

Training workforce for the UK economy
The proposed program will provide a framework for professional training of a postdoctoral researcher. She will master a highly desirable set of skills for modern biomedical research, such as proteomics, bioinformatics and microdevice engineering. The postdoc will acquire advanced communication and managerial skills, as well as gain more experience in supervising students. This comprehensive training will maximize her value as a skilled employee capable of making important contributions to the UK academia and industry on completion of her training in the PI's lab.

Education and public engagement
The program will allow us to make a lasting impact in the education sector. We will supervise summer and term research projects of undergraduate students from King's (at least 1 per year). In addition, we will offer one-week placements to children who attend state-funded schools within the borough of Camden (1 per year). The first-hand experience in modern research will raise students' awareness of different science professions and enhance their educational opportunities. Of note, the PI's laboratory has an established track record of hosting undergraduate (~35) and school (5) students both in Singapore and the UK, with most students choosing to study biomedical sciences further.

The concept of scale engages imagination, from Gulliver's Travels to Alice in Wonderland to Kandinsky's exploration of geometric rules for painting. Together with a British designer, researcher, and educator Mike Thompson (Thought Collider), we will develop a performative artwork to explore the relationship between size and shape in biological and man-made systems. Our vision is to engage our audience in the creative process, to maximize learning interactions on both sides. We plan for a two-tier approach starting with engagement at the Crick Manby Gallery to prototype, develop and refine our project, eventually taking it to a wider audience through local community (e.g. Somerstown Festival of Cultures or Cally Fest) or the national festivals. This will provide opportunities for conversation and deeper engagement with the artwork and the underlying scientific questions.

Together, these activities will contribute to scientific education in the UK and make fundamental cell biology a part of a wider cultural landscape.

Medicine and agriculture
Our focus on understanding morphogenetic plasticity in a fungal system should deliver long-term impacts in medicine and agriculture. Diseases caused by ascomycete fungi cause significant morbidity and mortality in humans. In plants, ascomycetes are responsible for devastating crop diseases such as rice blast and grey mould. Fungal diseases claim a huge socioeconomic burden, e.g. human fungal diseases in the European Union cost up to 50,000 EUR per patient, depending on the duration of the follow-up (PMID: 24026863). Rice blast typically causes 10-30% loss of grain, staggering numbers given that ~50% of the world's population relies on rice for their primary caloric intake (PMID: 22471698). Understanding biology of fungal pathogenesis is vital from societal and economic viewpoints. Although S. japonicus has a saprophytic lifestyle, it exhibits major morphogenetic plasticity including cellular geometry scaling and nutrient- and stress-triggered yeast-to-hyphae transitions. Fungal pathogens rely on similar morphogenetic events to infect their hosts and we believe that our results will be of use to researchers working on cellular aspects of fungal pathogenesis. To maximize the impact of our work in this field, we will publish our results as open-access papers in journals with widest readerships possible and reach out to relevant academic and industry scientists in London and elsewhere in the UK to discuss areas of common interest with a view of establishing collaborations, e.g. through BBSRC-funded Industrial CASE Partnerships and London Interdisciplinary Doctoral Programme.