Gel-Based Chemical Cross-Linking Analysis of 20S Proteasome Subunit-Subunit Interactions in Breast Cancer

The ubiquitin-proteasome system plays a pivotal role in breast tumorigenesis by controlling transcription factors, thus promoting cell cycle growth, and degradation of tumor suppressor proteins. However, breast cancer patients have failed to benefit from proteasome inhibitor treatment partially due to proteasome heterogeneity, which is poorly understood in malignant breast neoplasm. Chemical crosslinking is an increasingly important tool for mapping protein three-dimensional structures and proteinprotein interactions. In the present study, two cross-linkers, bis（sulfosuccinimidyl） suberate（BS3） and its water-insoluble analog disuccinimidyl suberate（DSS）, were used to map the subunit-subunit interactions in 20 S proteasome core particle（CP） from MDA-MB-231 cells. Different types of gel electrophoresis technologies were used. In combination with chemical cross-linking and mass spectrometry, we applied these gel electrophoresis technologies to the study of the noncovalent interactions among 20 S proteasome subunits. Firstly, the CP subunit isoforms were profiled. Subsequently, using native/SDSPAGE, it was observed that 0.5 mmol/L BS^3 was a relatively optimal cross-linking concentration for CP subunit-subunit interaction study. 2-DE analysis of the cross-linked CP revealed that α1 might preinteract with α2, and α3 might pre-interact with α4. Moreover, there were different subtypes of α1α2 and α3α4 due to proteasome heterogeneity. There was no significant difference in cross-linking pattern for CP subunits between BS3 and DSS. Taken together, the gel-based characterization in combination with chemical cross-linking could serve as a tool for the study of subunit interactions within a multi-subunit protein complex. The heterogeneity of 20 S proteasome subunit observed in breast cancer cells may provide some key information for proteasome inhibition strategy.

3D printing of architectured graphene-based aerogels by cross-linking GO inks with adjustable viscoelasticity for energy storage devices

Three-dimensional(3D)functional graphenebased architecture with superior electrical conductivity and good mechanical strength has promising applications in energy storage and electrics.Viscoelasticity-adjustable inks make it possible to achieve desired 3D architectures with interconnected and continuous interior networks by microextrusion printing.In this work,ultra-low-concentration graphene oxide(GO)inks of~15 mg·ml-1 have been obtained and demonstrated in direct 3D printing with a facile cross-linking(direct ink writing).The rheological behavior of the GO strategy by cations,which is the lowest concentration to achieve direct ink writing inks,could be adjusted from 1×10^(4) to 1×10^(5) Pa·s^(-1) with different concentrations of cations due to strong cross-linking networks between GO sheets and cations.Meanwhile,the specific strength and electrical conductivity of 3D-printed graphene architecture are notably enhanced,reaching up to 51.7×10^(3) N·m·kg^(-1)and 119 S·m^(-1),which are superior to conventional graphene aerogels.Furthermore,3D printing graphene-based architecture assembled in micro-superc apacitor exhibits excellent electrochemical performance,which can be ascribed to the effective ion transportation through the interconnected networks.The strategy demonstrated is useful in the design of complex-shaped,graphene-based architectures for scalable manufacturing of practical energy storage applications.