Taxanes convert regions of perturbed microtubule growth into rescue sites

Microtubules are polymers of tubulin dimers, and conformational transitions in the microtubule lattice drive microtubule dynamic instability and affect various aspects of microtubule function. The exact nature of these transitions and their modulation by anti -cancer drugs such as Taxol and epothilone, which can stabilize microtubules but also perturb their growth, are poorly understood. Here, we directly visualize the action of fluorescent Taxol and epothilone derivatives and show that microtubules can transition to a state that triggers cooperative drug binding to form regions with altered lattice conformation. Such regions emerge at growing microtubule ends that are in a pre-catastrophe state and inhibit microtubule growth and shortening. Electron microscopy and in vitro dynamics data indicate that taxane accumulation zones represent incomplete tubes that can persist, incorporate tubulin dimers and repeatedly induce microtubule rescues. Thus, taxanes modulate the material properties of microtubules by converting destabilized growing microtubule ends into regions resistant to depolymerization

End-binding proteins sensitize microtubules to the action of microtubule-targeting agents

Microtubule-targeting agents (MTAs) are widely used for treatment of cancer and other diseases, and a detailed understanding of the mechanism of their action is important for the development of improved microtubule-directed therapies. Although there is a large body of data on the interactions of different MTAs with purified tubulin andmicrotubules,much less is known about how the effects of MTAs are modulated by microtubule-associated proteins. Among the regulatory factors with a potential to have a strong impact on MTA activity are the microtubule plus end-tracking proteins, which controlmultiple aspects of microtubule dynamic instability. Here,we reconstituted microtubule dynamics in vitro to investigate the influence of end-binding proteins (EBs), the core components of the microtubule plus end-tracking protein machinery, on the effects that MTAs exert onmicrotubule plus-end growth.We found that EBs promote microtubule catastrophe induction in the presence of all MTAs tested. Analysis of microtubule growth times supported the view that catastrophes aremicrotubule age dependent. This analysis indicated that MTAs affect microtubule aging in multiple ways: destabilizing MTAs, such as colchicine and vinblastine, accelerate aging in an EB-dependentmanner,whereas stabilizingMTAs, such as paclitaxel and peloruside A, induce not only catastrophes but also rescues and can reverse the aging process

Organization and dynamics of the cortical complexes controlling insulin secretion in beta-cells

Insulin secretion in pancreatic beta-cells is regulated by cortical complexes that are enriched at the sites of adhesion to extracellular matrix facing the vasculature. Many components of these complexes, including bassoon, RIM, ELKS and liprins, are shared with neuronal synapses. Here, we show that insulin secretion sites also contain the non-neuronal proteins LL5 beta (also known as PHLDB2) and KANK1, which, in migrating cells, organize exocytotic machinery in the vicinity of integrin-based adhesions. Depletion of LL5 beta or focal adhesion disassembly triggered by myosin II inhibition perturbed the clustering of secretory complexes and attenuated the first wave of insulin release. Although previous analyses in vitro and in neurons have suggested that secretory machinery might assemble through liquid-liquid phase separation, analysis of endogenously labeled ELKS in pancreatic islets indicated that its dynamics is inconsistent with such a scenario. Instead, fluorescence recovery after photobleaching and single-molecule imaging showed that ELKS turnover is driven by binding and unbinding to low-mobility scaffolds. Both the scaffold movements and ELKS exchangewere stimulated by glucose treatment. Our findings help to explain how integrin-based adhesions control spatial organization of glucose-stimulated insulin release.Nephrolog

Investigation of the complex antibiotic INA-5812

A concentrate with the antimicrobial activity has been isolated from the culture broth of Streptomyces roseoflavus INA-Ac-5812. Its further fractionation by reversed-phase HPLC has resulted in six fractions. It has been established by MALDI-TOF and ESI-MSn precision mass-spectrometry methods that the main components of the complex antibiotic are several closely related compounds, presumably of a glycopeptide nature. The fraction containing an individual component with a mass of 1845.788 Da has been characterized by UV/Vis absorbance and fluorescence spectra, amino acid analysis, and derivatization with tris(2,6-dimethoxyphenyl)methyl cation. The activity of fractions against pathogenic microbes has been studied. The results allow the supposition that the INA-5812 antibiotic complex is a glyco- or lipoglycopeptide antibiotic of a new type, which is very promising for further study. © 2016, Pleiades Publishing, Ltd

Investigation of the complex antibiotic INA-5812

A concentrate with the antimicrobial activity has been isolated from the culture broth of Streptomyces roseoflavus INA-Ac-5812. Its further fractionation by reversed-phase HPLC has resulted in six fractions. It has been established by MALDI-TOF and ESI-MSn precision mass-spectrometry methods that the main components of the complex antibiotic are several closely related compounds, presumably of a glycopeptide nature. The fraction containing an individual component with a mass of 1845.788 Da has been characterized by UV/Vis absorbance and fluorescence spectra, amino acid analysis, and derivatization with tris(2,6-dimethoxyphenyl)methyl cation. The activity of fractions against pathogenic microbes has been studied. The results allow the supposition that the INA-5812 antibiotic complex is a glyco- or lipoglycopeptide antibiotic of a new type, which is very promising for further study. © 2016, Pleiades Publishing, Ltd