Proteomic analysis of brain proteins in APP/PS-1 human double mutant knock-in mice with increasing amyloid β-peptide deposition: insights into the effects of in vivo treatment with N-acetylcysteine as a potential therapeutic intervention in mild cognitive impairment and Alzheimer's disease

Proteomics analyses were performed on the brains of wild-type (WT) controls and an Alzheimer's disease (AD) mouse model, APP/PS-1 human double mutant knock-in mice. Mice were given either drinking water or water supplemented with N-acetylcysteine (NAC) (2 mg/kg body weight) for a period of five months. The time periods of treatment correspond to ages prior to Aβ deposition (i.e. 4-9 months), resembling human mild cognitive impairment (MCI), and after Aβ deposition (i.e. 7-12 months), more closely resembling advancing stages of AD. Substantial differences exist between the proteomes of WT and APP/PS-1 mice at 9 or 12 months, indicating that Aβ deposition and oxidative stress lead to downstream changes in protein expression. Altered proteins are involved in energy-related pathways, excitotoxicity, cell cycle signaling, synaptic abnormalities, and cellular defense and structure. Overall, the proteomic results support the notion that NAC may be beneficial for increasing cellular stress responses in WT mice and for influencing the levels of energy- and mitochondria-related proteins in APP/PS-1 mice.     

Accelerating protein docking in ZDOCK using an advanced 3D convolution library

Computational prediction of the 3D structures of molecular interactions is a challenging area, often requiring significant computational resources to produce structural predictions with atomic-level accuracy. This can be particularly burdensome when modeling large sets of interactions, macromolecular assemblies, or interactions between flexible proteins. We previously developed a protein docking program, ZDOCK, which uses a fast Fourier transform to perform a 3D search of the spatial degrees of freedom between two molecules. By utilizing a pairwise statistical potential in the ZDOCK scoring function, there were notable gains in docking accuracy over previous versions, but this improvement in accuracy came at a substantial computational cost. In this study, we incorporated a recently developed 3D convolution library into ZDOCK, and additionally modified ZDOCK to dynamically orient the input proteins for more efficient convolution. These modifications resulted in an average of over 8.5-fold improvement in running time when tested on 176 cases in a newly released protein docking benchmark, as well as substantially less memory usage, with no loss in docking accuracy. We also applied these improvements to a previous version of ZDOCK that uses a simpler non-pairwise atomic potential, yielding an average speed improvement of over 5-fold on the docking benchmark, while maintaining predictive success. This permits the utilization of ZDOCK for more intensive tasks such as docking flexible molecules and modeling of interactomes, and can be run more readily by those with limited computational resources.     

Vertebrate host protective immunity drives genetic diversity and antigenic polymorphism in Schistosoma mansoni

Schistosomes are gonochoric blood parasites with a complex life cycle responsible for a disease of considerable medical and veterinary importance in tropical and subtropical regions. Understanding the evolution of schistosome genetic diversity is clearly of fundamental importance to interpreting schistosomiasis epidemiology and disease transmission patterns of this parasite. In this article, we investigated the putative role of the host immune system in the selection of male genetic diversity. We demonstrated the link between genetic dissimilarity and the protective effect among male worms. We then compared the proteomes of three male clones with different genotypes and differing by their capacity to protect against reinfection. The identified differences correspond mainly to antigens known or supposed to be involved in the induction of protective immunity. These results underline the role played by host immune system in the selection of schistosome genetic diversity that is linked to antigenic diversity. We discuss the evolutionary consequences in the context of schistosome infection.     

Nutrient stress does not cause retrograde transport of cytoplasmic tRNA to the nucleus in evolutionarily diverse organisms

Intracellular trafficking of tRNA was long thought to be a one-way trip from the site of biogenesis in the nucleus to the translation machinery in the cytoplasm. This view has recently been challenged, however, by the discovery that tRNA can move retrograde from the cytoplasm back to the nucleus in Saccharomyces cerevisiae and rat hepatoma H4IIE cells during nutrient stress and in S. cerevisiae after intron-containing pre-tRNAs are spliced in the cytoplasm. Contrary to studies reported, we present data suggesting that nutrient stress does not cause retrograde transport of cytoplasmic tRNAs to the nucleus in rat hepatoma H4IIE cells, human HeLa and HEK293 cells, and the yeasts Kluyveromyces lactis and S. cerevisiae. However, the efficiency of nuclear re-export of retrograded spliced tRNA was severely affected in S. cerevisiae and two other Saccharomyces species deprived of nutrient. Collectively, the data suggest that nutrient stress does not cause nuclear import of cytoplasmic tRNA; instead, nutrient stress specifically regulates nuclear re-export of retrograded spliced tRNAs but not nuclear export of tRNAs made from intronless pre-tRNAs in Saccharomyces species. Furthermore, we provide evidence suggesting that Mtr10p and the Gsp1pGTP/Gsp1pGDP cycle are not involved in nuclear tRNA import in S. cerevisiae during nutrient stress.     

Distal Loop Flexibility of a Regulatory Domain Modulates Dynamics and Activity of C-Terminal Src Kinase (Csk)

The Src family of tyrosine kinases (SFKs) regulate numerous aspects of cell growth and differentiation and are under the principal control of the C-terminal Src Kinase (Csk). Csk and SFKs share a modular design with the kinase domain downstream of the N-terminal SH2 and SH3 domains that regulate catalytic function and membrane localization. While the function of interfacial segments in these multidomain kinases are well-investigated, little is known about how surface sites and long-range, allosteric coupling control protein dynamics and catalytic function. The SH2 domain of Csk is an essential component for the down-regulation of all SFKs. A unique feature of the SH2 domain of Csk is the tight turn in place of the canonical CD loop in a surface site far removed from kinase domain interactions. In this study, we used a combination of experimental and computational methods to probe the importance of this difference by constructing a Csk variant with a longer SH2 CD loop to mimic the flexibility found in homologous kinase SH2 domains. Our results indicate that while the fold and function of the isolated domain and the full-length kinase are not affected by loop elongation, native protein dynamics that are essential for efficient catalysis are perturbed. We also identify key motifs and routes through which the distal SH2 site might influence catalysis at the active site. This study underscores the sensitivity of intramolecular signaling and catalysis to native protein dynamics that arise from modest changes in allosteric regions while providing a potential strategy to alter intrinsic activity and signaling modulation.     

Transglutaminase 2 expression in acute myeloid leukemia: Association with adhesion molecule expression and leukemic blast motility

Acute myeloid leukemia (AML) is a heterogenous disease with differential oncogene association, outcome and treatment regimens. Treatment strategies for AML have improved outcome but despite increased molecular biological information AML is still associated with poor prognosis. Proteomic analysis on the effects of a range of leukemogenic oncogenes showed that the protein transglutaminase 2 (TG2) is expressed at greater levels as a consequence of oncogenic transformation. Further analysis of this observation was performed with 511 AML samples using reverse phase proteomic arrays, demonstrating that TG2 expression was higher at relapse than diagnosis in many cases. In addition elevated TG2 expression correlated with increased expression of numerous adhesion proteins and many apoptosis regulating proteins, two processes related to leukemogenesis. TG2 has previously been linked to drug resistance in cancer and given the negative correlation between TG2 levels and peripheral blasts observed increased TG2 levels may lead to the protection of the leukemic stem cell due to increased adhesion/reduced motility. TG2 may therefore form part of a network of proteins that define poor outcome in AML patients and potentially offer a target to sensitize AML stem cells to drug treatment.