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Guynet, C.

Publications and source records attributed to Guynet, C..

2 recordsLinked to original sources

An easy-to-use microfluidic mechano-chemostat for tissues and organisms reveals that confined growth is accompanied with increased macromolecular crowding

Conventional culture conditions are oftentimes insufficient to study tissues, organisms, or 3D multicellular assemblies. They lack both dynamic chemical and mechanical control over the microenvironment. While specific microfluidic devices have been developed to address chemical control, they are often hard to use and do not allow the control of compressive forces. Here, we present a set of microfluidic devices which all rely on the use of sliding elements consisting of microfabricated rods that can be inserted inside a microfluidic device. Sliding elements enable the creation of reconfigurable sealed culture chambers for the study of whole organisms or model micro-tissues. By confining the micro-tissues, we studied the biophysical impact of growth-induced pressure and showed that this mechanical stress is associated with an increase in macromolecular crowding, shedding light on this understudied type of mechanical stress. Our mechano-chemostat is an easy-to-use microfluidic device that allows the long-term culture of biological samples and can be used to study both the impact of specific conditions as well as the consequences of mechanical compression.

biophysics↗

Characterization of the DNA binding domain of StbA, a key protein of a new type of DNA segregation system

Low-copy-number plasmids require sophisticated genetic devices to achieve efficient segregation of plasmid copies during cell division. Plasmid R388 uses a unique segregation mechanism, based on StbA, a small multifunctional protein. StbA is the key protein in a segregation system not involving a plasmid-encoded NTPase partner, it regulates the expression of several plasmid operons, and it is the main regulator of plasmid conjugation. The mechanisms by which StbA, together with the centromere-like sequence stbS, achieves segregation, is largely uncharacterized. To better understand the molecular basis of R388 segregation, we determined the crystal structure of the conserved N-terminal domain of StbA to 1.9 [A] resolution. It folds into an HTH DNA-binding motif, structurally related to that of the PadR subfamily II of transcriptional regulators. StbA is organized in two domains. Its N-terminal domain carries the specific stbS DNA binding activity. A truncated version of StbA, deleted of its C-terminal domain, displays only partial activities in vivo, indicating that the non-conserved C-terminal domain is required for efficient segregation and subcellular plasmid positioning. The structure of StbA DNA-binding domain also provides some insight into how StbA monomers cooperate to repress transcription by binding to the stbDR and to form the segregation complex with stbS.

microbiology↗