Fast developments in synthetic biology now enable introduction of entire novel-to-nature pathways into industrial microorganisms, thus opening the door to a novel bio-based industry. However, successful scale-up of novel-to-market products from bench-top proof-of-principle to full-scale processes requires a profound understanding of cellular and sub-cellular mechanisms and their interaction with industrial process conditions. In particular, successful industrial production requires optimal performance of engineered strains under the dynamic, slow-growth process conditions that occur in large-scale industrial bioreactors. These differ strongly from the conditions in conventional laboratory analysis.The RoBoYeast project aims to reduce time-to-market and to improve productivity of microbial synthetic pathways by identifying key physiological and molecular determinants of performance under industrially relevant conditions and to integrate these at an early stage in strain development programs. The project is characterized by multi-scale analysis and integration of results from industrial-scale into lab-scale studies.RoBoYeast aims to improve productivity and cellular robustness in large-scale fermentations of the novel-to-nature production in Saccharomyces cerevisiae of arachidonic and eicosapentaenoic acid, two health-promoting poly-unsaturated fatty acids with excellent market perspectives as nutraceuticals. Biosynthetic pathways of these compounds are energy (ATP) intensive, which adds on the challenge of constructing robust strains that withstand typical production stresses without loss of productivity and product yield. To accelerate project progress, key findings of a likewise challenging resveratrol progress, already developed by the SME partner, can be used. RoBoYeast intends to identify and improve robustness mechanism not only for the benefit of this project but also as a blueprint for optimization of the production of other ATP-intensive products.