Manipulation of topoisomerase expression inhibits cell division but not growth and reveals a distinctive promoter structure in Synechocystis.

In cyanobacteria DNA supercoiling varies over the diurnal cycle and is integrated with temporal programs of transcription and replication. We manipulated DNA supercoiling in Synechocystis sp. PCC 6803 by CRISPRi-based knockdown of gyrase subunits and overexpression of topoisomerase I (TopoI). Cell division was blocked but cell growth continued in all strains. The small endogenous plasmids were only transiently relaxed, then became strongly supercoiled in the TopoI overexpression strain. Transcript abundances showed a pronounced 5'/3' gradient along transcription units, incl. the rRNA genes, in the gyrase knockdown strains. These observations are consistent with the basic tenets of the homeostasis and twin-domain models of supercoiling in bacteria. TopoI induction initially led to downregulation of G+C-rich and upregulation of A+T-rich genes. The transcriptional response quickly bifurcated into six groups which overlap with diurnally co-expressed gene groups. Each group shows distinct deviations from a common core promoter structure, where helically phased A-tracts are in phase with the transcription start site. Together, our data show that major co-expression groups (regulons) in Synechocystis all respond differentially to DNA supercoiling, and suggest to re-evaluate the long-standing question of the role of A-tracts in bacterial promoters.

Silica nanoparticles disrupt OPT-2/PEP-2-dependent trafficking of nutrient peptides in the intestinal epithelium.

Despite of the increasing application of silica nanoparticles and identification of oral exposure as a major entry portal, we lack understanding of nanosilica effects in the gut. Thus, we investigated biointeractions of nanosilica with single intestinal cells. The invertebrate nematode Caenorhabditis elegans was chosen as model organism with a tractable intestine and realistic target organism of nanomaterials in the environment. We found that nanosilica impairs the intestinal uptake of oligopeptides. Downstream to absorption by the apical OPT-2/PEP-2 transporter dipeptides were trapped in aberrant vesicles that grow over time and reach diameters of ≥6 µm. The peptide vesicles do not correspond to known organelles such as gut granules and form independently of related gene products GLO-1 or GLO-3. Formation of aberrant peptide vesicles also occurred independently of insulin/IGF-I receptor (DAF-2) signaling and daf-2 loss of function mutants showed specific vesicle patterns including distinct localization along the apical membrane of single intestinal cells. As malnutrition of exposed C. elegans manifested in reduced growth and a petite phenotype similar to OPT-2/PEP-2 transporter deficient mutants, we conclude that nanosilica-induced peptide vesicles represent a new compartment of di- and tripeptide trapping which disrupts hydrolysis of nutrient peptides and metabolism.