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Quantitative developmental genetic analysis of phenotypic buffering and cryptic variation

Periodic Reporting for period 4 - ROBUSTNET (Quantitative developmental genetic analysis of phenotypic buffering and cryptic variation)

Reporting period: 2020-04-01 to 2021-09-30

• What is the problem/issue being addressed?

We investigate the genetic mechanisms of biological robustness, aiming at deriving broad principles about how animals operate consistently. Biological robustness is a poorly studied phenomenon that has recently attracted a lot of attention both from an experimental and theoretical point of view. We often forget how complicated biological systems are, how many perturbations they constantly face and how much stochasticity there is when looking at the underlying molecular processes. It is therefore rather remarkable that biological systems can operate often with extreme consistency. One such example is development in the simple model nematode Caenorhabditis elegans, which is very stereotypical so that the zygote always produces the same number of 959 cells in a reproducible manner with minimal animal-to-animal variation. The majority of previous studies have been performed in unicellular organisms. We use instead a comprehensive system-wide approach using a tractable multi-cellular model organism to break down development robustness of cell numbers and dissect its mechanisms.


• Why is it important for society?


Although system robustness in non-biology fields such as engineering (e.g. robustness of buildings, bridges, aeroplanes or the internet) is man-made, biological robustness arises from selection or as a by-product of evolution and the underlying mechanisms are very little understood. It is currently unclear whether broad principles can be derived that are important for any type of robustness and whether knowledge from one field can guide thinking or future work in another field (e.g. biologically-inspired engineering). Understanding the basis of biological robustness is a fundamental problem that is relevant to biomedical sciences as a lot of diseases can also be studied within the context of defective homeostasis. New approaches in building synthetic biological networks can be influenced by reaching an understanding on principles of biological robustness.



• What are the overall objectives?

Our project has three main aims. First, to develop a new experimental system to study and systematically quantify developmental robustness in C. elegans. Second, to identify a wide-spectrum of genetic determinants of phenotypic robustness and characterise the mechanisms of biological buffering. Third, to explore how differences in the genetic background for example present in divergent nematode populations may affect developmental system traits including robustness to perturbations.
Aim (1): We finalized our work on gene expression profiling in the epidermis and characterised the consequences of environmental perturbations on seam cell number robustness.

Aim (2): We completed our analysis on key mutations recovered from our forward genetic screen focusing on variability of seam cell number as the primary phenotypic read-out. This included phenotypic characterisation of mutant strains using microscopy, imaging and genetics to identify the nature and mechanisms of phenotypic variability as well as experimental validation to establish causality of these mutations for the variable phenotype.

Aim (3): We have completed our work on understanding how the genetic background influences the outcome of developmental mutations. A key highlight was the completion of the genetic characterization of the difference in the expressivity of a mutation in the GATA transcription factor EGL-18 between CB4856 (from Hawaii) and the lab reference N2 (from Bristol). We were able to unpack the high complexity of the genetic architecture discovered and demonstrate that natural variation in a conserved heat shock protein modulates the outcome of a mutation affecting stem cell development through modifying the conserved Wnt signalling pathway.
A major achievement in Aim 1 was the development of a new method (Targeted DamID) that allows researchers to study trancriptomes in a cell-type-specific manner in intact animals without cell isolation. Major achievements in Aim 2 include the completion of work on understanding how mutations in the fusogen eff-1 and the N-acetyltransferase nath-10 affect phenotypic variability through modulating the Wnt signaling pathway. Discoveries of the genetic basis of cryptic genetic variation in Aim 3 have broad implications for Wnt-dependent stem cell biology and disease.Overall we produced a big collection of mutants and new data on phenotypic variability. Variable phenotypes are in general more difficult to isolate and study so the project was successful in delivering these results. We anticipate our findings to be impactful within our field with potential wider implications as highlighted above.