Detailed observations of transcriptional, translational, and post-translational events in the human brain are essential to improving our understanding of its development, function, and vulnerability to disease. Here, we exploited label-free quantitative tandem mass-spectrometry proteomics to create an in-depth proteomic survey of adult human brain regions. Integration of protein data with existing whole-transcriptome sequencing (RNA-seq) from the BrainSpan project revealed varied patterns of protein:RNA relationships with generally increased magnitudes of protein abundance differences between brain regions compared to RNA. Many of the differences amplified in protein data were reflective of cyto-architectural and functional variation between brain regions. Comparing structurally similar cortical regions revealed significant differences in the abundance of receptor-associated and resident plasma membrane proteins that were not readily observed in the RNA expression data.

This study identifies an unusual sulfur-based chemical as a novel and specific inhibitor of the tyrosine phosphatase STEP and shows that it can improve the cognitive function of a mouse model of Alzheimer's disease.

USP7 is a deubiquitinating enzyme that plays a pivotal role in multiple oncogenic pathways and therefore is a desirable target for new anti-cancer therapies. However, the lack of structural information about the USP7-inhibitor interactions has been a critical gap in the development of potent inhibitors. USP7 is unique among USPs in that its active site is catalytically incompetent, and is postulated to rearrange into a productive conformation only upon binding to ubiquitin. Surprisingly, we found that ubiquitin alone does not induce an active conformation in solution. Using a combination of nuclear magnetic resonance, mass spectrometry, computational modeling, and cell-based assays, we found that DUB inhibitors P22077 and P50429 covalently modify the catalytic cysteine of USP7 and induce a conformational switch in the enzyme associated with active site rearrangement. This work represents the first experimental insights into USP7 activation and inhibition and provides a structural basis for rational development of potent anti-cancer therapeutics.

The arrangement and stacking of noncovalently contiguous double-helical sections are increasingly invoked in single-stranded DNA and RNA tertiary structure. These tertiary structures of nucleic acids are defined by their double stranded regions, and their orientation in the molecular frame constitutes an important component of the nucleic acid structure. A direct view of these tertiary structures can be obtained by fluorescence polarization anisotropy of bound ethidium bromide (EB). The orientation of the dye in the molecular frame of the nucleic acid yields the orientation of the helix. The complete anisotropy function for EB intercalated in genome-derived DNA duplexes was derived by Allison and Schurr (1979) and accounts for base-pair twisting and DNA bending. Single-stranded ribozymes, ribosomal and transfer RNAs, and model DNA junctions contain double-stranded regions shorter than 35 bp in length, for which bending is not significant. We developed and experimentally verified an expression of the anisotropy function for short DNA duplexes which is theoretically compatible with the existing theory, originally developed for long nucleic acids (Schurr et al., 1992). Simulations showed that for DNA duplexes shorter than 35 bp, our expression of the anisotropy function is equivalent to Schurr's and is consistent with experiments carried out on eight DNA duplexes. Modeling the eight duplexes as cylinders, we calculate a duplex diameter of 1.91 +/- 0.15 nm when EB makes a 90 degrees angle with the DNA helix axis and undergoes anisotropic wobbling and 1.97 +/- 0.15 nm when EB makes a 70.5 degrees angle and undergoes isotropic wobbling, respectively. We used this treatment to establish the conformation of five DNA oligonucleotides made of single and tethered hairpins, some designed to exhibit coaxial stacking. Analysis of the fluorescence anisotropy decays shows that the tethered hairpins take an extended rather than parallel conformation. It also shows that the DNA oligonucleotides made of two tethered hairpins exhibit freedom compatible with two independent hairpins. When the linker between hairpins is shortened, the two hairpins are not independent anymore as probed by fluorescence anisotropy, suggesting coaxial stacking of the two helices.