The inner physicochemical
heterogeneity of bacterial cells generates
three-dimensional (3D)-dependent variations of resources for effective
expression of given chromosomally located genes. This fact has been
exploited for adjusting the most favorable parameters for implanting
a complex device for optogenetic control of biofilm formation in the
soil bacteriumPseudomonas putida.
To this end, a DNA segment encoding a superactive variant of theCaulobacter crescendusdiguanylate cyclase PleD expressed
under the control of the cyanobacterial light-responsive CcaSR system
was placed in a mini-Tn5 transposon vector and randomly inserted through
the chromosome of wild-type and biofilm-deficient variants ofP. putidalacking thewspgene cluster.
This operation delivered a collection of clones covering a whole range
of biofilm-building capacities and dynamic ranges in response to green
light. Since the phenotypic output of the device depends on a large
number of parameters (multiple promoters, RNA stability, translational
efficacy, metabolic precursors, protein folding, etc.), we argue that
random chromosomal insertions enable sampling the intracellular milieu
for an optimal set of resources that deliver a preset phenotypic specification.
Results thus support the notion that the context dependency can be
exploited as a tool for multiobjective optimization, rather than a
foe to be suppressed in Synthetic Biology constructs.