A 14C age calibration curve for the last 60 ka: the Greenland-Hulu U/Th timescale and its impact on understanding the Middle to Upper Paleolithic transition in Western Eurasia

This paper combines the data sets available today for ¹⁴C-age calibration of the last 60 ka. By stepwise synchronization of paleoclimate signatures, each of these sets of ¹⁴C-ages is compared with the U/Th-dated Chinese Hulu Cave speleothem records, which shows global paleoclimate change in high temporal resolution. By this synchronization we have established an absolute-dated Greenland-Hulu chronological framework, against which global paleoclimate data can be referenced, extending the ¹⁴C-age calibration curve back to the limits of the radiocarbon method. Based on this new, U/Th-based Greenland_Hulu chronology, we confirm that the radiocarbon timescale underestimates calendar ages by several thousand years during most of Oxygen Isotope Stage 3. Major atmospheric ¹⁴C variations are observed for the period of the Middle to Upper Paleolithic transition, which has significant implications for dating the demise of the last Neandertals. The early part of “the transition” (with ¹⁴C ages > 35.0 ka ¹⁴C BP) coincides with the Laschamp geomagnetic excursion. This period is characterized by highly-elevated atmospheric ¹⁴C levels. The following period ca. 35.0-32.5 ka ¹⁴C BP shows a series of distinct large-scale ¹⁴C age inversions and extended plateaus. In consequence, individual archaeological ¹⁴C dates older than 35.0 ka ¹⁴C BP can be age-calibrated with relatively high precision, while individual dates in the interval 35.0-32.5 ka ¹⁴C BP are subject to large systematic age-‘distortions,’ and chronologies based on large data sets will show apparent age-overlaps of up to ca. 5,000 cal years. Nevertheless, the observed variations in past ¹⁴C levels are not as extreme as previously proposed (“Middle to Upper Paleolithic dating anomaly”), and the new chronological framework leaves ample room for application of radiocarbon dating in the age-range 45.0-25.0 ka ¹⁴C BP at high temporal resolution.     

Direct and quantitative single-cell analysis of human immunodeficiency virus type 1 reactivation from latency

The ability of human immunodeficiency virus type 1 (HIV-1) to establish latent infections in cells has received renewed attention owing to the failure of highly active antiretroviral therapy to eradicate HIV-1 in vivo. Despite much study, the molecular bases of HIV-1 latency and reactivation are incompletely understood. Research on HIV-1 latency would benefit from a model system that is amenable to rapid and efficient analysis and through which compounds capable of regulating HIV-1 reactivation may be conveniently screened. We describe a novel reporter system that has several advantages over existing in vitro systems, which require elaborate, expensive, and time-consuming techniques to measure virus production. Two HIV-1 molecular clones (NL4-3 and 89.6) were engineered to express enhanced green fluorescent protein (EGFP) under the control of the viral long terminal repeat without removing any viral sequences. By using these replication-competent viruses, latently infected T-cell (Jurkat) and monocyte/macrophage (THP-1) lines in which EGFP fluorescence and virus expression are tightly coupled were generated. Following reactivation with agents such as tumor necrosis factor alpha, virus expression and EGFP fluorescence peaked after 4 days and over the next 3 weeks each declined in a synchronized manner, recapitulating the establishment of latency. Using fluorescence microscopy, flow cytometry, or plate-based fluorometry, this system allows immediate, direct, and quantitative real-time analysis of these processes within single cells or in bulk populations of cells. Exploiting the single-cell analysis abilities of this system, we demonstrate that cellular activation and virus reactivation following stimulation with proinflammatory cytokines can be uncoupled.     

Protein kinase C delta regulates the phosphorylation of heat shock protein 27 in human hepatocellular carcinoma

We have recently reported that attenuated phosphorylation of heat shock protein (HSP) 27 correlates with tumor progression in patients with hepatocellular carcinoma (HCC). In the present study, we investigated what kind of kinase regulates phosphorylation of HSP27 in human HCC-derived HuH7 cells. 12-O-tetradecanoylphorbol-13-acetate (TPA) and 1-oleoyl-2-acetylglycerol, direct activators of protein kinase C (PKC), markedly strengthened the phosphorylation of HSP27. Bisindorylmaleimide I, an inhibitor of PKC, suppressed the TPA-induced levels of HSP27 phosphorylation in addition to its basal levels. Knock down of PKCdelta suppressed HSP27 phosphorylation, as well as p38 mitogen-activated protein kinase (MAPK) phosphorylation. SB203580, an inhibitor of p38 MAPK, suppressed the TPA-induced HSP27 phosphorylation. Our results strongly suggest that activation of PKCdelta regulates the phosphorylation of HSP27 via p38 MAPK in human HCC.     

Evidence for convergent evolution in the antimicrobial peptide system in anuran amphibians

Amphibians are characterized in part by their highly specialized and glandular skin that enables key physiological functions such as cutaneous respiration and defense against a variety of micro- and macroscopic predators via toxic components (e.g., alkaloids and bufodienolids), biogenic amines, neuropeptides and antimicrobial peptides (AMPs). To date, DNA sequence information regarding AMP genes in anurans is restricted to only a few anuran families and largely to “higher frogs” (Neobatrachia). Here, we analyze the DNA information for the signal sequences of the AMP precursors in anuran amphibians available to the end of 2009 in an explicit phylogenetic framework to characterize the evolution of this large, diverse gene family. Comparison of cDNA sequences suggests that there are at least three different motifs within the signal peptide sequence of the AMP-precursor corresponding to the evolutionary lineages Neobatrachia, Bombinatoridae (Bombina spp.) and Pipidae (Xenopus laevis). The signal sequences are strongly conserved within each lineage (as previously noted for Neobatrachia), but highly divergent between them. Together with the lack of a linear relationship between the degree of sequence divergence and evolutionary time, we hypothesize that the anuran AMP system has evolved convergently on at least three occasions. However, additional sampling, especially among the largely poorly sampled non-neobatrachian lineages, is required to confirm this hypothesis and could reveal the existence of additional signal sequence motifs.     

Compound K, a metabolite of ginsenosides, induces cardiac protection mediated nitric oxide via Akt/PI3K pathway

AimsCompound K (C-K; 20-O-d-glucopyranosyl-20(S)-protopanaxadiol) is a novel ginsenoside metabolite formed by intestinal bacteria and does not occur naturally in ginseng. In this study, we investigated whether administration of C-K has protective effects on myocardial ischemia-reperfusion injury and its potential mechanisms.Main methodsWe used in vivo mouse models of ischemia-reperfusion injury and performed biochemical assays in excised hearts.Key findingsC-K reduced infarct size compared with the control group after ischemia-reperfusion. Immunoblot analysis showed that C-K significantly enhanced protein kinase B (Akt) and endothelial nitric oxide synthase (eNOS) activity. Wortmannin, a phosphoinositide 3-kinase (PI3K) inhibitor, blocked cardiac protection in vivo and attenuated phosphorylation of Akt and eNOS. Additionally, the hearts of C-K pretreated mice showed inhibition of mitochondrial swelling induced by Ca²⁺.SignificanceThis study showed that Compound K pretreatment has protective effects on myocardial ischemia-reperfusion injury, partly by mediating the activation of PI3K pathway and phosphorylation of Akt and eNOS.     

Distinction of G0 cells from senescent cells in cultures of non-cycling human fetal lung fibroblasts by anti-MAP-1 monoclonal antibody staining

On staining with a monoclonal antibody raised against microtubule-associated protein-1 (MAP-1), dot-like structures were seen in the nuclei of interphase cells, but not in those of non-cycling G0-arrested cells. Dots were also not seen in the nuclei of non-cycling senescent human cells (IMR-90). A SV40-DNA-transformed subline of IMR-90 with a limited growth potential showed progressive decrease of cells with nuclei containing dots in the final stage of their lifespan. The dots appeared in G0-arrested IMR-90 cells when these cells were incubated in medium of high osmotic pressure for 3 min. In contrast, no dots appeared in senescent cells or X-ray-irradiated young cells when they were incubated in medium of high osmotic pressure. Thus irreversibly non-cycling cells could be distinguished from G0-phase cells on the level of whole cultures. The results suggest that senescent cells lose their division potential by entering an irreversible cell-cycle stage differing from G0.     

Yop fusions to tightly folded protein domains and their effects on Yersinia enterocolitica type III secretion

Yersinia enterocolitica organisms secrete Yop proteins via the type III pathway. Translational fusion of yop genes to ubiquitin or dihydrofolate reductase results in hybrid proteins that cannot be secreted. The folding of hybrids prevents their own transport, but it does not hinder the type III secretion of other Yops.     

An Atypical Tubulin Kinase Mediates Stress-Induced Microtubule Depolymerization in Arabidopsis

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Environment-mediated accumulation of diacyl lipoproteins over their triacyl counterparts in Staphylococcus aureus

Bacterial lipoproteins are believed to exist in only one specific lipid-modified structure, such as the diacyl form or the triacyl form, in each bacterium. In the case of Staphylococcus aureus, recent extensive matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry analysis revealed that S. aureus lipoproteins exist in the α-aminoacylated triacyl form. Here, we discovered conditions that induce the accumulation of diacyl lipoproteins that lack α-aminoacylation in S. aureus. The accumulation of diacyl lipoproteins required a combination of conditions, including acidic pH and a post-logarithmic-growth phase. High temperatures and high salt concentrations additively accelerated the accumulation of the diacyl lipoprotein form. Following a post-logarithmic-growth phase where S. aureus MW2 cells were grown at pH 6, SitC lipoprotein was found almost exclusively in its diacyl structure rather than in its triacyl structure. This is the first report showing that the environment mediates lipid-modified structural alterations of bacterial lipoproteins.