Ophelia S Venturelli | bioRxiv Science

bioRxiv2016

Programming mRNA decay to modulate synthetic circuit resource allocation

Synthetic biomolecular networks embedded in host-cells compete with cellular processes for limited intracellular resources. Resource utilization is a major variable that impacts synthetic circuit behavior. Here we show that intracellular resources could be diverted from cellular operations to a synthetic circuit by programming global mRNA decay using the sequence-dependent endoribonuclease MazF. Synthetic circuit genes were protected from MazF activity by recoding the gene sequence to eliminate recognition sites, while preserving the amino acid sequence. The expression of a protected fluorescent reporter and the metabolic flux of a high-value metabolite gluconate were significantly enhanced using this genome-scale control strategy. Proteomics measurements discovered a key host translation factor in need of protection to optimize the resource redistribution activity. A dynamic computational model demonstrated that the MazF mRNA-decay feedback loop achieved proportional control of MazF levels in an optimal operating regime. RNA-seq time-series measurements of MazF-induced cells elucidated the dynamic shifts in the transcript abundance and discovered regulatory design elements that could be used to expand the control mechanisms of the MazF resource allocator. Together, these results demonstrated that manipulation of resource allocation is a tunable parameter that can be used to redirect resources away from cellular processes to synthetic circuits to enhance target functions.

Synthetic Biology↗

bioRxiv2014

Population diversification in a yeast metabolic program promotes anticipation of environmental shifts

Delineating the strategies by which cells contend with combinatorial changing environments is crucial for understanding cellular regulatory organization. When presented with two carbon sources, microorganisms first consume the carbon substrate that supports the highest growth rate (e.g. glucose) and then switch to the secondary carbon source (e.g. galactose), a paradigm known as the Monod model. Sequential sugar utilization has been attributed to transcriptional repression of the secondary metabolic pathway, followed by activation of this pathway upon depletion of the preferred carbon source. In this work, we challenge this notion. Although Saccharomyces cerevisiae cells consume glucose before galactose, we demonstrate that the galactose regulatory pathway is activated in a fraction of the cell population hours before glucose is fully consumed. This early activation reduces the time required for the population to transition between the two metabolic programs and provides a fitness advantage that might be crucial in competitive environments. Importantly, these findings define a new paradigm for the response of microbial populations to combinatorial carbon sources.

Systems Biology↗