The glycophorin A transmembrane sequence within integrin αvß3 creates a non-signaling integrin with low basal affinity that is strongly adhesive under force.

Integrin heterodimeric cell adhesion and signaling receptors bind ligands of the extracellular matrix and relay signals bidirectionally across cell membranes. Thereby, integrins adopt multiple conformational and functional states that control ligand binding affinity and linkage to cytosolic/cytoskeletal proteins. Here, we designed an integrin chimera encompassing the strongly dimerizing transmembrane domain (TMD) of glycophorin A (GpA) in the context of the otherwise unaltered integrin αvß3. We hypothesized that this chimera should have a low basal affinity to soluble ligand but should be force-activatable. By cellular expression of this chimera, we found a decreased integrin affinity to a soluble peptide ligand and inhibited intracellular signaling. However, under external forces applied by an atomic force microscope or by a spinning disc device causing shear forces, the mutant caused stronger cell adhesion than the wild-type integrin. Our results demonstrate that the signaling- and migration-incapable integrin αvß3-TMD mutant TMD-GpA shows the characteristics of a primed integrin state, which is of low basal affinity in the absence of forces, but may form strong bonds in the presence of forces. Thus, TMD-GpA may mimic a force-activatable signaling intermediate.

Probing the interaction forces of prostate cancer cells with collagen I and bone marrow derived stem cells on the single cell level.

Adhesion of metastasizing prostate carcinoma cells was quantified for two carcinoma model cell lines LNCaP (lymph node-specific) and PC3 (bone marrow-specific). By time-lapse microscopy and force spectroscopy we found PC3 cells to preferentially adhere to bone marrow-derived mesenchymal stem cells (SCP1 cell line). Using atomic force microscopy (AFM) based force spectroscopy, the mechanical pattern of the adhesion to SCP1 cells was characterized for both prostate cancer cell lines and compared to a substrate consisting of pure collagen type I. PC3 cells dissipated more energy (27.6 aJ) during the forced de-adhesion AFM experiments and showed significantly more adhesive and stronger bonds compared to LNCaP cells (20.1 aJ). The characteristic signatures of the detachment force traces revealed that, in contrast to the LNCaP cells, PC3 cells seem to utilize their filopodia in addition to establish adhesive bonds. Taken together, our study clearly demonstrates that PC3 cells have a superior adhesive affinity to bone marrow mesenchymal stem cells, compared to LNCaP. Semi-quantitative PCR on both prostate carcinoma cell lines revealed the expression of two Col-I binding integrin receptors, α1ß1 and α2ß1 in PC3 cells, suggesting their possible involvement in the specific interaction to the substrates. Further understanding of the exact mechanisms behind this phenomenon might lead to optimized therapeutic applications targeting the metastatic behavior of certain prostate cancer cells towards bone tissue.

Identifying discrete states of a biological system using a novel step detection algorithm.

Identification of discrete states is a common task when studying biological systems on microscopic scales. Here, we present a novel step detection algorithm that is ideally suited to locate steplike features separating adjacent plateaus, even if they are smooth and hidden by noise. It can be adjusted to detect very low or narrow steps that cannot be recognized by conventional methods. We demonstrate the applicability of the technique on various experimental data and show strong evidence of sub-10-pN steps in atomic force spectroscopy measurements performed with living lymphocytes.