Alterations In Regulatory Proteins Research Articles

Proteasomes of Autophagy-Deficient Cells Exhibit Alterations in Regulatory Proteins and a Marked Reduction in Activity.

Autophagy and the ubiquitin proteasome system are the two major processes for the clearance and recycling of proteins and organelles in eukaryotic cells. Evidence is accumulating that there is extensive crosstalk between the two pathways, but the underlying mechanisms are still unclear. We previously found that autophagy 9 (ATG9) and 16 (ATG16) proteins are crucial for full proteasomal activity in the unicellular amoeba Dictyostelium discoideum. In comparison to AX2 wild-type cells, ATG9-and ATG16- cells displayed a 60%, and ATG9-/16- cells a 90%, decrease in proteasomal activity. Mutant cells also showed a significant increase in poly-ubiquitinated proteins and contained large ubiquitin-positive protein aggregates. Here, we focus on possible reasons for these results. Reanalysis of published tandem mass tag-based quantitative proteomic results of AX2, ATG9-, ATG16-, and ATG9-/16- cells revealed no change in the abundance of proteasomal subunits. To identify possible differences in proteasome-associated proteins, we generated AX2 wild-type and ATG16- cells expressing the 20S proteasomal subunit PSMA4 as GFP-tagged fusion protein, and performed co-immunoprecipitation experiments followed by mass spectrometric analysis. The results revealed no significant differences in the abundance of proteasomes between the two strains. However, we found enrichment as well as depletion of proteasomal regulators and differences in the ubiquitination of associated proteins for ATG16-, as compared to AX2 cells. Recently, proteaphagy has been described as a means to replace non-functional proteasomes. We propose that autophagy-deficient D. discoideum mutants suffer from inefficient proteaphagy, which results in the accumulation of modified, less-active, and also of inactive, proteasomes. As a consequence, these cells exhibit a dramatic decrease in proteasomal activity and deranged protein homeostasis.

Functional characterization of somatic mutations in cancer using network-based inference of protein activity.

Identifying the multiple dysregulated oncoproteins that contribute to tumorigenesis in a given patient is crucial for developing personalized treatment plans. However, accurate inference of aberrant protein activity in biological samples is still challenging as genetic alterations are only partially predictive and direct measurements of protein activity are generally not feasible. To address this problem we introduce and experimentally validate a new algorithm, VIPER (Virtual Inference of Protein-activity by Enriched Regulon analysis), for the accurate assessment of protein activity from gene expression data. We use VIPER to evaluate the functional relevance of genetic alterations in regulatory proteins across all TCGA samples. In addition to accurately inferring aberrant protein activity induced by established mutations, we also identify a significant fraction of tumors with aberrant activity of druggable oncoproteins—despite a lack of mutations, and vice-versa. In vitro assays confirmed that VIPER-inferred protein activity outperforms mutational analysis in predicting sensitivity to targeted inhibitors.

Overexpression of sigma factor SigB improves temperature and butanol tolerance of Synechocystis sp. PCC6803

Among phenotypes of interest for an industrial cyanobacteria host are improved tolerance to temperature, salt, and solvent stress. Cellular responses to many stresses are controlled by the network of sensory receptors and downstream regulatory proteins. We applied transcription factor engineering to Synechocystis and tested mutant strains for tolerance to temperature and the biofuel 1-butanol. Histidine kinases (Hik), response regulators (Rre), and an RNA polymerase sigma factor (SigB) were overexpressed or deleted. Overexpression of SigB increased both temperature and butanol tolerance and lowered the intracellular concentration of reactive oxygen species. This report demonstrates that alteration of regulatory proteins in a cyanobacterium can be a useful tool to improve stress tolerance.

Attenuation of the development of murine solid leukemia tumor by physical exercise.

The active involvement of physical exercise in the evolution of a variety of cancers is well documented. However, its role in solid leukemia tumor development is essentially unknown. Solid leukemia tumor cells were transplanted into 21 hybrid BDF1 control mice, exercise-trained mice that did not exercise during leukemia and exercise-trained mice that exercised during leukemia. The tumor size of the continuously exercising group was ~50% of that of control and exercise-terminated animals 18 days after the transplantation. The activity of antioxidant enzymes and the levels of lipid peroxidation and 8-hydroxy-2'-deoxyguanosine were not different in the tumors of the three groups. The level of carbonylated proteins was smaller in tumors of continuously exercising animals. The mutant form of cell regulatory protein p53 and vascular endothelial growth factor were present in similar amounts in the tumor cells of each group. On the other hand, the protooncogene Ras and I-kappaB proteins were present in higher concentrations in tumors of continuously exercising rats. The present data suggest that exercise during leukemia attenuates the development of tumors in mice. The selective alteration of regulatory proteins might play a role in the beneficial effects of exercise during leukemia.