Contributions of the lower dimer to supramolecular actin patterning revealed by TIRF microscopy.

Two distinct dimers are formed during the initial steps of actin polymerization. The first one, referred to as the 'lower dimer' (LD) was discovered many years ago by means of chemical crosslinking. Owing to its transient nature, a biological relevance had long been precluded when, using LD-specific antibodies, we detected LD-like contacts in actin assemblies that are associated with the endolysosomal compartment in a number of different cell lines. Moreover, immunofluorescence showed the presence of LD-related structures at the cell periphery of migrating fibroblasts, in the nucleus, and in association with the centrosome of interphase cells. Here, we explore contributions of the LD to the assembly of supramolecular actin structures in real time by total internal reflection fluorescence (TIRF) microscopy. Our data shows that while LD on its own cannot polymerize under filament forming conditions, it is able to incorporate into growing F-actin filaments. This incorporation of LD triggers the formation of X-shaped filament assemblies with barbed ends that are pointing in the same direction in the majority of cases. Similarly, an increased frequency of junction sites was observed when filaments were assembled in the presence of oxidized actin. This data suggests that a disulfide bridge between Cys374 residues might stabilize LD-contacts. Based on our findings, we propose two possible models for the molecular mechanism underlying the supramolecular actin patterning in LD-related structures.

The nanomechanical signature of breast cancer.

Cancer initiation and progression follow complex molecular and structural changes in the extracellular matrix and cellular architecture of living tissue. However, it remains poorly understood how the transformation from health to malignancy alters the mechanical properties of cells within the tumour microenvironment. Here, we show using an indentation-type atomic force microscope (IT-AFM) that unadulterated human breast biopsies display distinct stiffness profiles. Correlative stiffness maps obtained on normal and benign tissues show uniform stiffness profiles that are characterized by a single distinct peak. In contrast, malignant tissues have a broad distribution resulting from tissue heterogeneity, with a prominent low-stiffness peak representative of cancer cells. Similar findings are seen in specific stages of breast cancer in MMTV-PyMT transgenic mice. Further evidence obtained from the lungs of mice with late-stage tumours shows that migration and metastatic spreading is correlated to the low stiffness of hypoxia-associated cancer cells. Overall, nanomechanical profiling by IT-AFM provides quantitative indicators in the clinical diagnostics of breast cancer with translational significance.

Synthetic protein targeting by the intrinsic biorecognition functionality of poly(ethylene glycol) using PEG antibodies as biohybrid molecular adaptors.

Biointerfaces capable of biological recognition and specificity are sought after for conferring bioinspired functionality onto synthetic biomaterials systems. This is important for biosensing, bioseparations, and biomedical materials. Here, we demonstrate how intrinsic polymer-protein interactions between highly localized polyethylene glycol (PEG) brushes and PEG-binding antibodies can be used for sorting specific biomolecules from complex bulk biological fluids to synthetic nanoscale targets. A principal feature lies with the antifouling property of PEG that prevents unspecific binding. Exclusive access is provided by anti-PEG, which acts as a biohybrid molecular adaptor that sifts out and targets specific IgG "cargo" from solution to the PEG. The PEG can be reversibly washed and targeted in blood serum, which suggests potential benefits in technological applications. Moreover, anti-PEG binding triggers a stimuli-responsive conformational collapse in the PEG brush, thereby imparting an intrinsic "smart" biorecognition functionality to the PEG that can considerably impact its use as an antifouling biomaterial.