The role of Vimentin in Regulating Cell Invasive Migration in Dense Cultures of Breast Carcinoma Cells.

Cell migration and mechanics are tightly regulated by the integrated activities of the various cytoskeletal networks. In cancer cells, cytoskeletal modulations have been implicated in the loss of tissue integrity and acquisition of an invasive phenotype. In epithelial cancers, for example, increased expression of the cytoskeletal filament protein vimentin correlates with metastatic potential. Nonetheless, the exact mechanism whereby vimentin affects cell motility remains poorly understood. In this study, we measured the effects of vimentin expression on the mechano-elastic and migratory properties of the highly invasive breast carcinoma cell line MDA231. We demonstrate here that vimentin stiffens cells and enhances cell migration in dense cultures, but exerts little or no effect on the migration of sparsely plated cells. These results suggest that cell-cell interactions play a key role in regulating cell migration, and coordinating cell movement in dense cultures. Our findings pave the way toward understanding the relationship between cell migration and mechanics in a biologically relevant context.

Phosphorylation-Induced Mechanical Regulation of Intrinsically Disordered Neurofilament Proteins.

The biological function of protein assemblies has been conventionally equated with a unique three-dimensional protein structure and protein-specific interactions. However, in the past 20 years it has been found that some assemblies contain long flexible regions that adopt multiple structural conformations. These include neurofilament proteins that constitute the stress-responsive supportive network of neurons. Herein, we show that the macroscopic properties of neurofilament networks are tuned by enzymatic regulation of the charge found on the flexible protein regions. The results reveal an enzymatic (phosphorylation) regulation of macroscopic properties such as orientation, stress response, and expansion in flexible protein assemblies. Using a model that explains the attractive electrostatic interactions induced by enzymatically added charges, we demonstrate that phosphorylation regulation is far richer and versatile than previously considered.

Neurofilaments Function as Shock Absorbers: Compression Response Arising from Disordered Proteins.

What can cells gain by using disordered, rather than folded, proteins in the architecture of their skeleton? Disordered proteins take multiple coexisting conformations, and often contain segments which act as random-walk-shaped polymers. Using x-ray scattering we measure the compression response of disordered protein hydrogels, which are the main stress-responsive component of neuron cells. We find that at high compression their mechanics are dominated by gaslike steric and ionic repulsions. At low compression, specific attractive interactions dominate. This is demonstrated by the considerable hydrogel expansion induced by the truncation of critical short protein segments. Accordingly, the floppy disordered proteins form a weakly cross-bridged hydrogel, and act as shock absorbers that sustain large deformations without failure.

Probing the interactions of intrinsically disordered proteins using nanoparticle tags.

The structural plasticity of intrinsically disordered proteins serves as a rich area for scientific inquiry. Such proteins lack a fix three-dimensional structure but can interact with multiple partners through numerous weak bonds. Nevertheless, this intrinsic plasticity possesses a challenging hurdle in their characterization. We underpin the intermolecular interactions between intrinsically disordered neurofilaments in various hydrated conditions, using grafted gold nanoparticle (NP) tags. Beyond its biological significance, this approach can be applied to modify the surface interaction of NPs for the creation of future tunable "smart" hybrid biomaterials.

Neurofilament assembly and function during neuronal development.

Studies on the assembly of neuronal intermediate filaments (IFs) date back to the early work of Alzheimer. Developing neurons express a series of IF proteins, sequentially, at distinct stages of mammalian cell differentiation. This correlates with altered morphologies during the neuronal development, including axon outgrowth, guidance and conductivity. Importantly, neuronal IFs that fail to properly assemble into a filamentous network are a hallmark of neurodegenerative diseases such as amyotrophic lateral sclerosis, Alzheimer's, and Parkinson's disease. Traditional structural methodologies fail to fully describe neuronal IF assembly, interactions and resulting function due to IFs structural plasticity, particularly in their C-terminal domains. We review here current progress in the field of neuronal-specific IFs, a dominant component affecting the cytoskeletal structure and function of neurons.

Order and disorder in intermediate filament proteins.

Intermediate filaments (IFs), important components of the cytoskeleton, provide a versatile, tunable network of self-assembled proteins. IF proteins contain three distinct domains: an α-helical structured rod domain, flanked by intrinsically disordered head and tail domains. Recent studies demonstrated the functional importance of the disordered domains, which differ in length and amino-acid sequence among the 70 different human IF genes. Here, we investigate the biophysical properties of the disordered domains, and review recent findings on the interactions between them. Our analysis highlights key components governing IF functional roles in the cytoskeleton, where the intrinsically disordered domains dictate protein-protein interactions, supramolecular assembly, and macro-scale order.

Met kinetic signature derived from the response to HGF/SF in a cellular model predicts breast cancer patient survival.

To determine the signaling pathways leading from Met activation to metastasis and poor prognosis, we measured the kinetic gene alterations in breast cancer cell lines in response to HGF/SF. Using a network inference tool we analyzed the putative protein-protein interaction pathways leading from Met to these genes and studied their specificity to Met and prognostic potential. We identified a Met kinetic signature consisting of 131 genes. The signature correlates with Met activation and with response to anti-Met therapy (p<0.005) in in-vitro models. It also identifies breast cancer patients who are at high risk to develop an aggressive disease in six large published breast cancer patient cohorts (p<0.01, N>1000). Moreover, we have identified novel putative Met pathways, which correlate with Met activity and patient prognosis. This signature may facilitate personalized therapy by identifying patients who will respond to anti-Met therapy. Moreover, this novel approach may be applied for other tyrosine kinases and other malignancies.