Redox control of protein function involves oxidation and reduction of amino acid residues, but the mechanisms and regulators involved are insufficiently understood. Here, we report that in ...conjunction with Mical proteins, methionine-R-sulfoxide reductase B1 (MsrB1) regulates mammalian actin assembly via stereoselective methionine oxidation and reduction in a reversible, site-specific manner. Two methionine residues in actin are specifically converted to methionine-R-sulfoxide by Mical1 and Mical2 and reduced back to methionine by selenoprotein MsrB1, supporting actin disassembly and assembly, respectively. Macrophages utilize this redox control during cellular activation by stimulating MsrB1 expression and activity as a part of innate immunity. We identified the regulatory role of MsrB1 as a Mical antagonist in orchestrating actin dynamics and macrophage function. More generally, our study shows that proteins can be regulated by reversible site-specific methionine-R-sulfoxidation.
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•Mical and MsrB regulate actin through stereospecific methionine oxidation/reduction•Met oxidation depolymerizes actin and reduction promotes actin repolymerization•Selenoprotein MsrB1 regulates actin polymerization in response to LPS stimulation•Proteins can be regulated by reversible site-specific methionine-R-sulfoxidation
Diffuse alveolar hemorrhage (DAH), although rare, is a life-threatening complication of systemic lupus erythematosus (SLE). Little is known about the pathophysiology of DAH in humans, although ...increasingly neutrophils, NETosis and inflammatory monocytes have been shown to play an important role in the pristane-induced model of SLE which develops lung hemorrhage and recapitulates many of the pathologic features of human DAH. Using this experimental model, we asked whether endoplasmic reticulum (ER) stress played a role in driving the pathology of pulmonary hemorrhage and what role infiltrating neutrophils had in this process. Analysis of lung tissue from pristane-treated mice showed genes associated with ER stress and NETosis were increased in a time-dependent manner and reflected the timing of CD11b
Ly6G
neutrophil accumulation in the lung. Using precision cut lung slices from untreated mice we observed that neutrophils isolated from the peritoneal cavity of pristane-treated mice could directly induce the expression of genes associated with ER stress, namely
and
. Mice which had myeloid-specific deletion of PAD4 were generated and treated with pristane to assess the involvement of PAD4 and PAD4-dependent NET formation in pristane-induced lung inflammation. Specific deletion of PAD4 in myeloid cells resulted in decreased expression of ER stress genes in the pristane model, with accompanying reduction in IFN-driven genes and pathology. Lastly, coculture experiments of human neutrophils and human lung epithelial cell line (BEAS-2b) showed neutrophils from SLE patients induced significantly more ER stress and interferon-stimulated genes in epithelial cells compared to healthy control neutrophils. These results support a pathogenic role of neutrophils and NETs in lung injury during pristane-induced DAH through the induction of ER stress response and suggest that overactivation of neutrophils in SLE and NETosis may underlie development of DAH.
Cardiac fibrosis, denoted by the deposition of extracellular matrix, manifests with a variety of diseases such as hypertension, diabetes, and myocardial infarction. Underlying this pathological ...extracellular matrix secretion is an expansion of fibroblasts. The mouse is now a common experimental model system for the study of cardiovascular remodeling and elucidation of fibroblast responses to cardiac growth and stress is vital for understanding disease processes. Here, using diverse but fibroblast specific markers, we report murine fibroblast distribution and proliferation in early postnatal, adult, and injured hearts. We find that perinatal fibroblasts and endothelial cells proliferate at similar rates. Furthermore, regardless of the injury model, fibroblast proliferation peaks within the first week after injury, a time window similar to the period of the inflammatory phase. In addition, fibroblast densities remain high weeks after the initial insult. These results provide detailed information regarding fibroblast distribution and proliferation in experimental methods of heart injury.
•Perinatal cardiac fibroblast proliferation parallels cardiomyocyte proliferation•Resident cardiac fibroblasts expand in distinct patterns after cardiac injury.•Fibroblast proliferation peaks in the first week after injury, then returns to basal levels.•The fibroblast proliferative phase is similar to the reported inflammatory phase.
Nostocyclopeptides A1 and A2 are novel cyclic heptapeptides produced by the terrestrial cyanobacterium
Nostoc sp. ATCC53789 that possess a unique imino linkage in the macrocyclic ring. Herein we ...report the cloning, sequencing, annotation, and biochemical analysis of the 33-kb nostocyclopeptide (
ncp) biosynthetic gene cluster, which includes seven open reading frames predicted to be involved in the biosynthesis and transport of these natural products. The genetic architecture and domain organization of the
ncpA-B nonribosomal peptide synthetase (NRPS) is co-linear in arrangement with respect to the putative order of the biosynthetic assembly of the cyclic peptide. A reductase domain identified at the C-terminal end of the NRPS NcpB is predicted to catalyze an NAD(P)H-mediated hydride transfer to the heptapeptidyl-
S-enzyme intermediate NH
2-Tyr-Gly-DGln-Ile-Ser-mPro-Leu/Phe-
S-NRPS to yield a linear heptapeptide aldehyde that is subsequently captured intramolecularly with the amino group of the
N-terminal amino acid residue tyrosine to form a stable imine bond. While a few C-terminal reductases associated with NRPSs have been identified, the
ncp reductase is the first to mediate imine macrocyclization involving peptide N- and C-termini. Biochemical analysis of the NcpA1 and NcpB1 adenylation domains coupled with the recent characterization of the (2
S,4
S)-5-hydroxyleucine dehydrogenase NcpD, which is involved in the biosynthesis of the nonproteinogenic amino acid residue
l-4-methylproline from
l-leucine, support the involvement of this cluster in nostocyclopeptide biosynthesis.
The cloning, sequencing, annotation and biochemical analysis of the nostopeptolide (
nos) biosynthetic gene cluster from the terrestrial cyanobacterium
Nostoc sp. GSV224 is described. Nostopeptolides ...A1 and A2 are cyclic peptide-polyketide hybrid natural products possessing nine amino acid residues, a butyric acid group, and an internal acetate-derived unit that are linked by peptide and ester bonds. The
nos gene cluster includes eight ORFs encompassing 40 kb and includes most of the genes predicted to be involved in the biosynthesis and transport of this group of nonapeptolides. The genetic architecture and domain organization of the
nos synthetase, a mixed non-ribosomal peptide synthetase-polyketide synthase, is co-linear in arrangement with respect to the putative order of the biosynthetic assembly of the lipopeptolide. Biochemical analysis of the NosA1, NosC1 and NosD1 adenylation domains coupled with the recent characterization of the
nosE and
nosF gene products, which are involved in the biosynthesis of the rare non-proteinogenic amino acid residue
l-4-methylproline from
l-leucine, support the involvement of this gene cluster in nostopeptolide biosynthesis.
Objective
Mitochondria are found in the extracellular space in rheumatoid arthritis (RA). However, whether mitochondria are a source of autoantigens in RA has not been carefully addressed. Thus, we ...undertook this study to investigate the presence and significance of antimitochondrial antibodies (AMAs) in patients with RA.
Methods
AMAs were measured in serum samples from 3 cross‐sectional cohorts of RA patients (n = 95, n = 192, and n = 117) and healthy individuals (n = 38, n = 72, and n = 50) using a flow cytometry–based assay. Further, AMAs were detected using an anti–mitofusin‐1 (anti–MFN‐1) IgG enzyme‐linked immunosorbent assay and Western blot analysis. A longitudinal inception cohort, followed up for a median of 8 years, was used to study disease progression.
Results
AMA levels were elevated in RA patients from all 3 cohorts as compared to healthy individuals (P < 0.001, P < 0.05, and P < 0.01), with a range of 14–26% positivity. Levels of anti–MFN‐1 antibodies correlated with AMA levels (r = 0.31, P = 0.006) and were elevated in RA patients as compared to healthy individuals (P < 0.001). The presence of AMAs was associated with erosive disease (P < 0.05) and interstitial lung disease (P < 0.01). Further, AMA levels were found to predict erosive disease (odds ratio OR 4.59, P = 0.006) and joint space narrowing (OR 3.08, P = 0.02) independent of anti–citrullinated protein antibodies. Finally, anti–MFN‐1 antibodies identified seronegative patients developing erosive disease (OR 9.33; P = 0.02).
Conclusion
Our findings demonstrate the presence of novel autoantibodies targeting mitochondria in the setting of RA. AMAs were used to stratify patients based on disease phenotype and to predict development of erosive disease, including in patients with seronegative disease. Our results highlight the essential role of mitochondria in the pathogenesis of RA and suggest a possible benefit of therapies targeting mitochondrial‐mediated inflammation and clearance in these patients.
BACKGROUND: Reconstructing the evolutionary history of nervous systems requires an understanding of their architecture and development across diverse taxa. The spiralians encompass diverse body plans ...and organ systems, and within the spiralians, annelids exhibit a variety of morphologies, life histories, feeding modes and associated nervous systems, making them an ideal group for studying evolution of nervous systems. RESULTS: We describe nervous system development in the annelid Capitella teleta (Blake JA, Grassle JP, Eckelbarger KJ. Capitella teleta, a new species designation for the opportunistic and experimental Capitella sp. I, with a review of the literature for confirmed records. Zoosymposia. 2009;2:25–53) using whole-mount in situ hybridization for a synaptotagmin 1 homolog, nuclear stains, and cross-reactive antibodies against acetylated α-tubulin, 5-HT and FMRFamide. Capitella teleta is member of the Sedentaria (Struck TH, Paul C, Hill N, Hartmann S, Hosel C, Kube M, et al. Phylogenomic analyses unravel annelid evolution. Nature. 2011;471:95–8) and has an indirectly-developing, lecithotrophic larva. The nervous system of C. teleta shares many features with other annelids, including a brain and a ladder-like ventral nerve cord with five connectives, reiterated commissures, and pairs of peripheral nerves. Development of the nervous system begins with the first neurons differentiating in the brain, and follows a temporal order from central to peripheral and from anterior to posterior. Similar to other annelids, neurons with serotonin-like-immunoreactivity (5HT-LIR) and FMRFamide-like-immunoreactivity (FMRF-LIR) are found throughout the brain and ventral nerve cord. A small number of larval-specific neurons and neurites are present, but are visible only after the central nervous system begins to form. These larval neurons are not visible after metamorphosis while the rest of the nervous system is largely unchanged in juveniles. CONCLUSIONS: Most of the nervous system that forms during larvogenesis in C. teleta persists into the juvenile stage. The first neurons differentiate in the brain, which contrasts with the early formation of peripheral, larval-specific neurons found in some spiralian taxa with planktotrophic larvae. Our study provides a clear indication that certain shared features among annelids - e.g., five connectives in the ventral nerve cord - are only visible during larval stages in particular species, emphasizing the need to include developmental data in ancestral character state reconstructions. The data provided in this paper will serve as an important comparative reference for understanding evolution of nervous systems, and as a framework for future molecular studies of development.
All proposed gravitational explanations of the Pioneer anomaly must crucially face the Equivalence Principle. Thus, if Pioneers 10 and 11 were influenced by anomalous gravitational effects in regions ...containing other Solar System bodies, then those bodies should likewise be influenced, irrespective of their shape, composition or mass. Although the lack of any observed influence upon planetary orbits severely constrains such explanations, here we aim to construct by computer modeling, hypothetical gravitating annuli having no gravitational impact on planetary orbits from Mercury to Neptune. One model has a central zone, free of radial gravitation in the annular plane, and an ‘onset’ beyond Saturn’s orbit, where sunward annular gravitation increases to match the Pioneer anomaly data. Sharp nulls are included so that Uranus and Neptune escape this influence. Such models can be proportionately reduced in mass: a 1 % contribution to the anomaly requires an annulus of approximately 1 Earth mass. It is thus possible to comply with the JPL assessment of newly recovered data attributing 80 %, or more, of the anomaly to spacecraft heat, which appears to allow small contributions from other causes. Following the possibility of an increasing Kuiper belt density at great ranges, another model makes an outward small anomalous gravitation in the TNO region, tallying with an observed slight indication of such an effect, suggesting that New Horizons may slightly accelerate in this region.
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