Deciphering the limits, mechanisms and evolution of developmental robustness using the paradigm of C. elegans seam cell patterning.

Robustness is the ability of a system to maintain performance in the presence of perturbations. It is a very important principle in engineering, where systems like bridges, aeroplanes or the internet, are designed to withstand a variety of perturbations and retain functionality. Robustness is an equally important property of living systems. However, biological robustness is poorly studied and understood.

It is in fact remarkable how biological systems have evolved to perform reproducibly, in the presence of internal and external perturbations. Development, in particular, is highly robust to genetic and environmental variation and this is instrumental for the transformation of a fertilised egg into a multicellular individual. A very striking example of developmental robustness in humans is the presence of five hand digits, a number that is very robust to variation in the genetic composition among individuals or differences in the environment. However, robust systems are not infallible and make mistakes, so cases of poly- or oligodactyly, although rare, do still occur. Such cases represent developmental errors and allow us to quantify the limits of robustness of a particular developmental trait. Robustness is a property that ensures phenotypic stability, and as such, it is very important for system behaviour and evolution. Disease can also be viewed as a breakdown of robustness mechanisms. Therefore, understanding the general principles of what makes a biological system robust is a fundamental problem in biology.

To find satisfactory answers to similar basic questions, a common practice is to turn to model systems, where we can easily and rapidly perform a lot of fine experiments. Our favourite system is a small nematode, Caenorhabditis elegans, which is well-known for its simple, fast and highly reproducible development. We propose to focus on a group of epidermal cells of C. elegans, the so-called seam cells, and develop this tissue as a model system to study developmental robustness in animals. These cells have attracted substantial attention because they have stem cell properties, so they divide asymmetrically during larval stages and one daughter cell maintains the proliferating status, whereas the other daughter differentiates into a specialised tissue, such as neurons. As a result of these divisions, wild-type C. elegans adult hermaphrodites have 16 seam cells per lateral side. We would like to characterise the limits of the developmental robustness of this system to genetic (mutations) and environmental variation. Our preliminary data indicate that it is possible to identify mutations that increase seam cell number variation without affecting the mean number of seam cells, so we hope to identify what is the nature of these genes that contribute to ensure phenotypic robustness. We will develop and test hypotheses about how the identified genes play a role in phenotypic robustness and study whether their action is specific for the seam cells or more general to systemic robustness at the whole-animal level. Finally, we will address to what extent our results can be extrapolated to other systems, by studying the evolution of robustness mechanisms in C. briggsae, which is a related nematode species to C. elegans.

Our approach will likely enable us to gain insights into the genetic mechanisms and evolution of developmental robustness and derive some general principles about the degree of robustness to quantitative variation in critical regulators and the relative contribution of genes to stabilisation of developmental outcomes. Our goal is to develop broad hypotheses that will be informative to both biomedical scientists interested in robust drug targeting and synthetic biologists who aim at engineering robust biological networks.

Developmental systems are continuously subject to variation, be it genetic or environmental, yet they manage to operate with remarkable reproducibility. Such a property of a system to produce an invariant output in the presence of considerable intrinsic or extrinsic variation is called robustness. Developmental robustness ensures the stability of phenotypic traits, so it is important for the genotype-to-phenotype relationship. It also affects the evolution of a system by permitting the accumulation of genetic variation that is cryptic at the level of the final phenotype. It is conceivable that a breakdown of robustness, leading to an increase in trait variance, can have direct consequences to human health and disease. Therefore, understanding the limits, mechanisms and evolutionary forces underlying developmental robustness is a fundamental problem in biology.

Although recent years have seen an increase in theoretical studies addressing questions on robustness, experimental studies have remained very scarce, especially in multi-cellular eukaryotes. Therefore, for any gene network mediating a robust developmental phenotype, it remains unclear a) what are the genes contributing to phenotypic robustness, b) to what extent these genes represent components of the core developmental network, c) what are the mechanisms underlying the reduction of robustness in developmental mutants and d) how do the mechanisms of robustness evolve. We propose here to study these questions on the epidermal seam cells of C. elegans. We will employ genetic, cell and molecular biology approaches to identify and characterise genes buffering seam cell number variation and study their evolution within C. elegans and C. briggsae. These results will enrich our understanding of the seam cell gene regulatory network and provide new insights into how biological systems buffer the variability of development outcomes.

The most important short-term beneficiaries from this work include the academic community (as described in the academic beneficiaries section above) and the general public. The general public will benefit directly from this project via the following ways. We will host high school students for summer projects and explore possibilities for children science education with the neighboring museums, such as the Natural History and Science Museum in London. We will participate in University Open Access days and attempt to involve our undergraduate students in our research through summer projects and conversations. We will also maintain a lab website where we will explain our research in simple terms, in order to educate and engage the public. We will finally use the Imperial public outreach laboratory and university media offices to communicate important research findings, whenever that is appropriate.

Longer-term beneficiaries are:
1. Biotech industries: Identifying robustness factors and points of fragility for biological systems can provide useful insight into disease and guidance for novel drug targeting. For example, cancer is a complex disease state that exhibits a high level of robustness against therapeutical treatments. This robustness is likely to be achieved by co-opting or "hijacking" intrinsic cellular mechanisms of robustness. Therefore, understanding how a cell or tissue performs a biological function with very high fidelity despite perturbation is likely to be informative for understanding how tumour cells adapt and remain proliferative upon anticancer drug intervention. Moreover, the seam cell robustness paradigm we propose relates to how an organism maintains correct cell numbers for a given tissue, by monitoring proliferation vs differentiation decisions. This concept is at the heart of cancer formation, therefore some identified seam cell regulators are likely to be relevant for cancer research. The estimated time frame for this impact is around 5-10 years from the end of the grant.

2. Bioengineering and synthetic biology companies will benefit from deriving principles and increasing our understanding on the molecular mechanisms underlying biological robustness. Creating robust biological networks is a major challenge in these fields, so our work on the genetic basis of robustness is likely to assist with the design and synthesis of gene regulatory circuits. The estimated time frame for this impact is around 10-15 years from the end of the grant.

3. Engineers, computer scientists and information technologists will also benefit from this work. There are some common principles on how to create robust architectures that are more broadly shared between engineering and biology. These include modularity, functional redundancy and feedback regulatory complexity. Therefore, in depth understanding of biological robustness can facilitate biologically inspired engineering approaches. The estimated time frame for this impact is around 15-20 years from the end of the grant.