SummaryGenes encoding cell-surface proteins regulated by SigB are stably expressed in Staphylococcus aureus small-colony variants (SCVs) isolated from cystic fibrosis (CF) patients. Our hypothesis is that CF-isolated SCVs are locked into a colonization state by sustaining the expression of adhesins such as fibronectin-binding proteins (FnBPs) throughout growth. Force spectroscopy was used to study the fibronectin-FnBPs interaction among strains varying for their SigB activity. The fibronectin-FnBPs interaction was described by a strength of 1000 Ϯ 400 pN (pulling rate of 2 mm s . A CF-isolated SCV highly expressed fnbA throughout growth and showed a sustained capacity to bind fibronectin, whereas a prototypic strain showed a reduced frequency of fibronectin-binding during the stationary growth phase when its fnbA gene was down-regulated. Reduced expression of fnbA was observed in sigB mutants, which was associated with an overall decrease adhesion to fibronectin. These results suggest that the fibronectin-FnBPs interaction plays a role in the formation of a mechanically resistant adhesion of S. aureus to host tissues and supports the hypothesis that CF-isolated SCVs are locked into a colonization state as a result of a sustained SigB activity.

The exploration of molecular processes governing physiological functions has significantly benefited from the emergence of novel nanoscaled techniques. Atomic force microscopy in force measurement mode can be used to investigate a multitude of cellular events in individual living cells with great sensitivity. Precise regions of the plasma membrane can be examined in relation to particular signalling pathways activated by a mechanical stimulus. Similarly, subtle cellular movements induced by biochemical activation of specific membrane receptors can be detected in real time with excellent temporal and spatial resolution. The possibility to challenge locally and mechanically cell surface receptors also provides information on the control exerted by a cell over individual adhesion sites. Overall, this information is vital for an in-depth understanding of mechanisms related to cellular movement and morphological regulation. Lastly, atomic force microscope-based nanomanipulations on living cells have recently been proposed as a tool to influence and monitor cellular homeostasis by introducing specific molecular entities into or extracting them from the cytoplasm of individual cells. This review provides detailed examples on how such atomic force microscopy experiments can be conducted to investigate processes relevant to cell physiology.

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Interaction between hyaluronic acid (HA) present at the surface of tubular epithelial cells and calcium oxalate monohydrate (COM) crystals is thought to play an important role in kidney stone formation. AFM-based force spectroscopy, where HA is covalently attached to AFM-probes, was used to quantify the interaction between HA and the surfaces of COM crystals. The work of adhesion of the HA-probe as well as the rupture force of single HA molecules were quantified in order to understand the molecular regulation of HA binding to COM crystals. Our results reveal that HA adsorbs to the crystal surface in physiological conditions. We also observed increased adhesion when the pH is lowered to a value that increases the risk of kidney stone formation. HA adhesion to the COM crystal surface can be suppressed by citrate, a physiological inhibitor of stone retention currently used in the treatment and prevention of kidney stone formation. Interestingly, we also observed preferential binding of HA onto the [100] face versus the [010] face, suggesting a major contribution of the [100] faces in the crystal retention process at the surface of tubular epithelial cells and the promotion of stone formation. Our results clearly establish a direct role for the glycosaminoglycan HA present at the surface of kidney tubular epithelium in the process of COM crystal retention.