Fluorescent amino acids as versatile building blocks for chemical biology

Fluorophores have transformed the way we study biological systems, enabling non-invasive studies in cells and intact organisms, which increase our understanding of complex processes at the molecular level. Fluorescent amino acids have become an essential chemical tool because they can be used to construct fluorescent macromolecules, such as peptides and proteins, without disrupting their native biomolecular properties. Fluorescent and fluorogenic amino acids with unique photophysical properties have been designed for tracking protein–protein interactions in situ or imaging nanoscopic events in real time with high spatial resolution. In this Review, we discuss advances in the design and synthesis of fluorescent amino acids and how they have contributed to the field of chemical biology in the past 10 years. Important areas of research that we review include novel methodologies to synthesize building blocks with tunable spectral properties, their integration into peptide and protein scaffolds using site-specific genetic encoding and bioorthogonal approaches, and their application to design novel artificial proteins, as well as to investigate biological processes in cells by means of optical imaging. [Figure not available: see fulltext.]

Multiple sea-ice states and abrupt MOC transitions in a general circulation ocean model

Sea ice has been suggested, based on simple models, to play an important role in past glacial–interglacial oscillations via the so-called “sea-ice switch” mechanism. An important requirement for this mechanism is that multiple sea-ice extents exist under the same land ice configuration. This hypothesis of multiple sea-ice extents is tested with a state-of-the-art ocean general circulation model coupled to an atmospheric energy–moisture-balance model. The model includes a dynamic-thermodynamic sea-ice module, has a realistic ocean configuration and bathymetry, and is forced by annual mean forcing. Several runs with two different land ice distributions represent present-day and cold-climate conditions. In each case the ocean model is initiated with both ice-free and fully ice-covered states. We find that the present-day runs converge approximately to the same sea-ice state for the northern hemisphere while for the southern hemisphere a difference in sea-ice extent of about three degrees in latitude between the different runs is observed. The cold climate runs lead to meridional sea-ice extents that are different by up to four degrees in latitude in both hemispheres. While approaching the final states, the model exhibits abrupt transitions from extended sea-ice states and weak meridional overturning circulation, to less extended sea ice and stronger meridional overturning circulation, and vice versa. These transitions are linked to temperature changes in the North Atlantic high-latitude deep water. Such abrupt changes may be associated with Dansgaard–Oeschger events, as proposed by previous studies. Although multiple sea ice states have been observed, the difference between these states is not large enough to provide a strong support for the sea-ice-switch mechanism

Bacillus anthracis Spore Surface Protein BclA Mediates Complement Factor H Binding to Spores and Promotes Spore Persistence

Spores of Bacillus anthracis, the causative agent of anthrax, are known to persist in the host lungs for prolonged periods of time, however the underlying mechanism is poorly understood. In this study, we demonstrated that BclA, a major surface protein of B. anthracis spores, mediated direct binding of complement factor H (CFH) to spores. The surface bound CFH retained its regulatory cofactor activity resulting in C3 degradation and inhibition of downstream complement activation. By comparing results from wild type C57BL/6 mice and complement deficient mice, we further showed that BclA significantly contributed to spore persistence in the mouse lungs and dampened antibody responses to spores in a complement C3-dependent manner. In addition, prior exposure to BclA deletion spores (ΔbclA) provided significant protection against lethal challenges by B. anthracis, whereas the isogenic parent spores did not, indicating that BclA may also impair protective immunity. These results describe for the first time an immune inhibition mechanism of B. anthracis mediated by BclA and CFH that promotes spore persistence in vivo. The findings also suggested an important role of complement in persistent infections and thus have broad implications

Genome-wide analysis of antiviral signature genes in porcine macrophages at different activation statuses

Macrophages (MФs) can be polarized to various activation statuses, including classical (M1), alternative (M2), and antiviral states. To study the antiviral activation status of porcine MФs during porcine reproductive and respiratory syndrome virus (PRRSV) infection, we used RNA Sequencing (RNA-Seq) for transcriptomic analysis of differentially expressed genes (DEGs). Sequencing assessment and quality evaluation showed that our RNA-Seq data met the criteria for genome-wide transcriptomic analysis. Comparisons of any two activation statuses revealed more than 20,000 DEGs that were normalized to filter out 153–5,303 significant DEGs [false discovery rate (FDR) ≤0.001, fold change ≥2] in each comparison. The highest 5,303 significant DEGs were found between lipopolysaccharide- (LPS) and interferon (IFN)γ-stimulated M1 cells, whereas only 153 significant DEGs were detected between interleukin (IL)-10-polarized M2 cells and control mock-activated cells. To identify signature genes for antiviral regulation pertaining to each activation status, we identified a set of DEGs that showed significant up-regulation in only one activation state. In addition, pathway analyses defined the top 20–50 significantly regulated pathways at each activation status, and we further analyzed DEGs pertinent to pathways mediated by AMP kinase (AMPK) and epigenetic mechanisms. For the first time in porcine macrophages, our transcriptomic analyses not only compared family-wide differential expression of most known immune genes at different activation statuses, but also revealed transcription evidence of multiple gene families. These findings show that using RNA-Seq transcriptomic analyses in virus-infected and status-synchronized macrophages effectively profiled signature genes and gene response pathways for antiviral regulation, which may provide a framework for optimizing antiviral immunity and immune homeostasis