Genetic screening of the breast and ovarian cancer susceptibility gene BRCA1 has uncovered a large number of variants of uncertain clinical significance. Here, we use biochemical and cell-based ...transcriptional assays to assess the structural and functional defects associated with a large set of 117 distinct BRCA1 missense variants within the essential BRCT domain of the BRCA1 protein that have been documented in individuals with a family history of breast or ovarian cancer. In the first method, we used limited proteolysis to assess the protein folding stability of each of the mutants compared with the wild-type. In the second method, we used a phosphopeptide pull-down assay to assess the ability of each of the variants to specifically interact with a peptide containing a pSer-X-X-Phe motif, a known functional target of the BRCA1 BRCT domain. Finally, we used transcriptional assays to assess the ability of each BRCT variant to act as a transcriptional activation domain in human cells. Through a correlation of the assay results with available family history and clinical data, we define limits to predict the disease risk associated with each variant. Forty-two of the variants show little effect on function and are likely to represent variants with little or no clinical significance; 50 display a clear functional effect and are likely to represent pathogenic variants; and the remaining 25 variants display intermediate activities. The excellent agreement between the structure/function effects of these mutations and available clinical data supports the notion that functional and structure information can be useful in the development of models to assess cancer risk.
Phage P1 is a temperate phage which makes the lytic or lysogenic decision upon infecting bacteria. During the lytic cycle, progeny phages are produced and the cell lyses, and in the lysogenic cycle, ...P1 DNA exists as a low-copy-number plasmid and replicates autonomously. Previous studies at the bulk level showed that P1 lysogenization was independent of
ultiplicity
f
nfection (MOI; the number of phages infecting a cell), whereas lysogenization probability of the paradigmatic phage λ increases with MOI. However, the mechanism underlying the P1 behavior is unclear. In this work, using a fluorescent reporter system, we demonstrated this P1 MOI-independent lysogenic response at the single-cell level. We further observed that the activity of the major repressor of lytic functions (C1) is a determining factor for the final cell fate. Specifically, the repression activity of P1, which arises from a combination of C1, the anti-repressor Coi, and the corepressor Lxc, remains constant for different MOI, which results in the MOI-independent lysogenic response. Additionally, by increasing the distance between phages that infect a single cell, we were able to engineer a λ-like, MOI-dependent lysogenization upon P1 infection. This suggests that the large separation of coinfecting phages attenuates the effective communication between them, allowing them to make decisions independently of each other. Our work establishes a highly quantitative framework to describe P1 lysogeny establishment. This system plays an important role in disseminating antibiotic resistance by P1-like plasmids and provides an alternative to the lifestyle of phage λ.
Phage P1 has been shown potentially to play an important role in disseminating antibiotic resistance among bacteria during lysogenization, as evidenced by the prevalence of P1 phage-like elements in animal and human pathogens. In contrast to phage λ, a cell fate decision-making paradigm, P1 lysogenization was shown to be independent of MOI. In this work, we built a simple genetic model to elucidate this MOI independency based on the gene-regulatory circuitry of P1. We also proposed that the effective communication between coinfecting phages contributes to the lysis-lysogeny decision-making of P1 and highlighted the significance of spatial organization in the process of cell fate determination in a single-cell environment. Finally, our work provides new insights into different strategies acquired by viruses to interact with their bacterial hosts in different scenarios for their optimal survival.
The BRCT repeats in BRCA1 are essential for its tumor suppressor activity and interact with phosphorylated protein targets containing the sequence pSer-X-X-Phe, where X indicates any residue. The ...structure of the tandem BRCA1 BRCT repeats bound to an optimized phosphopeptide reveals that the N-terminal repeat harbors a conserved BRCT phosphoserine-binding pocket, while the interface between the repeats forms a hydrophobic groove that recognizes the phenylalanine. Crystallographic and biochemical data suggest that the structural integrity of both binding sites is essential for peptide recognition. The diminished peptide-binding capacity observed for cancer-associated BRCA1-BRCT variants may explain the enhanced cancer risks associated with these mutations.
Photocatalysis under low energy light is of great importance for study of environmental pollution impact. There is an increasing demand for preparing integrated photocatalysts to show their potential ...in persistent organic compound degradation under visible and near-infrared light. In this work, we have reported the structural combination of three functional components of NaYF
4
:Yb,Er, TiO
2
, Cu into photocatalytic nanocomposites for near-infrared light (NIR) photocatalysis. Uniform and monodisperse NaYF
4
:Yb,Er nanocubes prepared by hydrothermolysis were used as a photon converter and they were combined with TiO
2
semiconductor and Cu co-catalysts by sequential deposition to fabricate NIR-active photocatalysts. The Cu/TiO
2
/NaYF
4
:Yb,Er nanocomposites were investigated for the organic degradation under NIR light. The Cu/TiO
2
/NaYF
4
:Yb,Er photocatalysts were photoactive with NIR light and full decomposition of methylene blue was reached for 90 min. These nanocomposites exhibit the NIR photocatalytic response caused by the NIR light absorption and photon electron transfer over NaYF
4
:Yb,Er, TiO
2
, and Cu components.
Most cancer-associated BRCA1 mutations identified to date result in the premature translational termination of the protein, highlighting a crucial role for the C-terminal, BRCT repeat region in ...mediating BRCA1 tumor suppressor function. However, the molecular and genetic effects of missense mutations that map to the BRCT region remain largely unknown. Using a protease-based assay, we directly assessed the sensitivity of the folding of the BRCT domain to an extensive set of truncation and single amino acid substitutions derived from breast cancer screening programs. The protein can tolerate truncations of up to 8 amino acids, but further deletion results in drastic BRCT folding defects. This molecular phenotype can be correlated with an increased susceptibility to disease. A cross-validated computational assessment of the BRCT mutation data base suggests that as much as half of all BRCT missense mutations contribute to BRCA1 loss of function and disease through protein-destabilizing effects. The coupled use of proteolytic methods and computational predictive methods to detect mutant BRCA1 conformations at the protein level will augment the efficacy of current BRCA1 screening protocols, especially in the absence of clinical data that can be used to discriminate deleterious BRCT missense mutations from benign polymorphisms.
Bioinspired materials have aroused great interest as their inherent biocompatible and structural characteristics have given rise to sustainable applications. In this work, we have reported the phase ...and morphology transformation of chitosan from crystalline nanofibrils into amorphous sheets for fabricating sustainable materials. Acetylation-induced aqueous dissolution of native chitosan nanofibrils affords water-soluble chitosan as a biopolymeric liquid. Water-soluble chitosan macromolecules self-aggregate into amorphous sheets on solidification, presenting an interesting way to inspire new structures of chitosan assemblies. Through control over gelation, lyophilization, and self-assembled confinement of water-soluble chitosan, we have fabricated novel chitosan materials including filaments, aerogels, microspheres, and plastics that are promising for sustainable use.
Bioinspired materials: chitosan filaments, aerogels, microspheres, and plastics have been designed by transforming crystalline nanofibrils into amorphous sheets on solidification.
This study focuses on the factors that impact on assessment of student satisfaction with the quality of training services at the College of Commerce in the period from 2014 to 2019. A survey ...questionnaire was sent to 249 student respondents. The survey period was from April 2019 to the end of May 2019. The research results show that there are 7 elements: Trust, Empathy, Tangible medium, Response, Service capacity. Based on the research results, recommendations for personnel policies have been proposed to improve student satisfaction with the quality of training services at the Da Nang Commercial College in the 2019-2030 periods.
A key step in the signaling cascade responsible for activation of the transcription factor NF‐κB involves Lys63‐linked polyubiquitination of TRAF6. Covalent attachment of ubiquitin (Ub) to TRAF6, and ...subsequent poly(Ub) chain synthesis, is catalyzed by the hUev1a–hUbc13 heterodimer. hUbc13 is a catalytically competent E2 enzyme, and hUev1a is an E2‐like protein that binds substrate Ub. The hUev1a–hUbc13 heterodimer is targeted to TRAF6 through interactions between hUbc13 and the N‐terminal RING domain from TRAF6. Nuclear magnetic resonance (NMR) spectroscopy was used to determine the solution state structure of the RING domain from human TRAF6, and the interaction between hUbc13 and TRAF6 was characterized using NMR chemical shift mapping. The main‐chain dynamics of the RING domain from TRAF6 were studied using 15N NMR relaxation. Analysis of the main‐chain dynamics data indicates that residues within the α‐helix and β‐sheet of the RING domain are as rigid as regions of canonical secondary structure in larger proteins, consistent with the biological role of RING‐domain E3 proteins, which requires that the E3 contain a recognition site for recruitment of E2 ubiquitin conjugation enzymes.
Modification of proteins by post-translational covalent attachment of a single, or chain, of ubiquitin molecules serves as a signaling mechanism for a number of regulatory functions in eukaryotic ...cells. For example, proteins tagged with lysine-63 linked polyubiquitin chains are involved in error-free DNA repair. The catalysis of lysine-63 linked polyubiquitin chains involves the sequential activity of three enzymes (E1, E2, and E3) that ultimately transfer a ubiquitin thiolester intermediate to a protein target. The E2 responsible for catalysis of lysine-63 linked polyubiquitination is a protein heterodimer consisting of a canonical E2 known as Ubc13, and an E2-like protein, or ubiquitin conjugating enzyme variant (UEV), known as Mms2. We have determined the solution structure of the complex formed by human Mms2 and ubiquitin using high resolution, solution state nuclear magnetic resonance (NMR) spectroscopy. The structure of the Mms2-Ub complex provides important insights into the molecular basis underlying the catalysis of lysine-63 linked polyubiquitin chains.
Simultaneous integration of photon emission and biocompatibility into nanoparticles is an interesting strategy to develop applications of advanced optical materials. In this work, we present the ...synthesis of biocompatible optical nanocomposites from the combination of near-infrared luminescent lanthanide nanoparticles and water-soluble chitosan. NaYF4:Yb,Er upconverting nanocrystal guests and water-soluble chitosan hosts are prepared and integrated together into biofunctional optical composites. The control of aqueous dissolution, gelation, assembly, and drying of NaYF4:Yb,Er nanocolloids and chitosan liquids allowed us to design novel optical structures of spongelike aerogels and beadlike microspheres. Well-defined shape and near-infrared response lead upconverting nanocrystals to serve as photon converters to couple with plasmonic gold (Au) nanoparticles. Biocompatible chitosan-stabilized Au/NaYF4:Yb,Er nanocomposites are prepared to show their potential use in biomedicine as we find them exhibiting a half-maximal effective concentration (EC50) of 0.58 mg mL–1 for chitosan-stabilized Au/NaYF4:Yb,Er nanorods versus 0.24 mg mL–1 for chitosan-stabilized NaYF4:Yb,Er after 24 h. As a result of their low cytotoxicity and upconverting response, these novel materials hold promise to be interesting for biomedicine, analytical sensing, and other applications.
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