Alzheimer's disease and non-demented high pathology control nonagenarians: comparing and contrasting the biochemistry of cognitively successful aging

The amyloid cascade hypothesis provides an economical mechanistic explanation for Alzheimer's disease (AD) dementia and correlated neuropathology. However, some nonagenarian individuals (high pathology controls, HPC) remain cognitively intact while enduring high amyloid plaque loads for decades. If amyloid accumulation is the prime instigator of neurotoxicity and dementia, specific protective mechanisms must enable these HPC to evade cognitive decline. We evaluated the neuropathological and biochemical differences existing between non-demented (ND)-HPC and an age-matched cohort with AD dementia. The ND-HPC selected for our study were clinically assessed as ND and possessed high amyloid plaque burdens. ELISA and Western blot analyses were used to quantify a group of proteins related to APP/Aβ/tau metabolism and other neurotrophic and inflammation-related molecules that have been found to be altered in neurodegenerative disorders and are pivotal to brain homeostasis and mental health. The molecules assumed to be critical in AD dementia, such as soluble or insoluble Aβ40, Aβ42 and tau were quantified by ELISA. Interestingly, only Aβ42 demonstrated a significant increase in ND-HPC when compared to the AD group. The vascular amyloid load which was not used in the selection of cases, was on the average almost 2-fold greater in AD than the ND-HPC, suggesting that a higher degree of microvascular dysfunction and perfusion compromise was present in the demented cohort. Neurofibrillary tangles were less frequent in the frontal cortices of ND-HPC. Biochemical findings included elevated vascular endothelial growth factor, apolipoprotein E and the neuroprotective factor S100B in ND-HPC, while anti-angiogenic pigment epithelium derived factor levels were lower. The lack of clear Aβ-related pathological/biochemical demarcation between AD and ND-HPC suggests that in addition to amyloid plaques other factors, such as neurofibrillary tangle density and vascular integrity, must play important roles in cognitive failure.     

Protein kinase CK2 links polyamine metabolism to MAPK signalling in Drosophila

MAPK signalling is a complex process not only requiring the core components Raf, MEK and Erk, but also many proteins like the scaffold protein KSR and several kinases to specifically localize, modulate and fine-tune the outcome of the pathway in a cell context specific manner. In mammals, protein kinase CK2 was shown to bind to the scaffold protein KSR and to phosphorylate Raf proteins at a conserved serine residue in the negative-charge regulatory (N−) region, thereby facilitating maximal activity of the MAPK signalling pathway. In this work we show that in Drosophila CK2 is also bound to KSR. However, despite the presence of a corresponding serine residue in the N-region of DRaf, CK2-mediated phosphorylation of DRaf takes place on a serine residue at the N-terminus and is required for Erk activation. Previous work identified polyamines as regulators of CK2 kinase activity. The main cellular source of polyamines is the catabolism of amino acids. Evidence is provided that phosphorylation of DRaf by CK2 is modulated by polyamines, with spermine being the most potent inhibitor of the reaction. We suggest that CK2 is able to monitor intracellular polyamine levels and translates this information to modulate MAPK signalling.     

Survival rates in a small hibernator,the edible dormouse: a comparison across Europe

Understanding how local environmental factors lead to temporal variability of vital rates and to plasticity of life history tactics is one of the central questions in population ecology. We used long‐term capture‐recapture data from five populations of a small hibernating rodent, the edible dormouse Glis glis, collected over a large geographical range across Europe, to determine and analyze both seasonal patterns of local survival and their relation to reproductive activity. In all populations studied, survival was lowest in early summer, higher in late summer and highest during hibernation in winter. In reproductive years survival was always lower than in non‐reproductive years, and females had higher survival rates than males. Very high survival rates during winter indicate that edible dormice rarely die from starvation due to insufficient energy reserves during the hibernation period. Increased mortality in early summer was most likely caused by high predation risk and unmet energy demands. Those effects have probably an even stronger impact in reproductive years, in which dormice were more active. Although these patterns could be found in all areas, there were also considerable differences in average survival rates, with resulting differences in mean lifetime reproductive success between populations. Our results suggest that edible dormice have adapted their life history strategies to maximize lifetime reproductive success depending on the area specific frequency of seeding events of trees producing energy‐rich seeds.     

Stepwise and large-magnitude negative shift in δ13Ccarb preceded the main marine mass extinction of the Permian–Triassic crisis interval

Large perturbations to the global carbon cycle occurred during the Permian–Triassic boundary mass extinction, the largest extinction event of the Phanerozoic Eon (542 Ma to present). Controversy concerning the pattern and mechanism of variations in the marine carbonate carbon isotope record of the Permian–Triassic crisis interval (PTCI) and their relationship to the marine mass extinction has not been resolved to date. Herein, high-resolution carbonate carbon isotope profiles (δ¹³C_carb), accompanied by lithofacies, were generated for four sections with microbialite (Taiping, Zuodeng, Cili, and Chongyang) in South China to better constrain patterns and controls on δ¹³C_carb variation in the PTCI and to test hypotheses about the temporal relationship between perturbations to the global carbon cycle and the marine mass extinction event. All four study sections exhibit a stepwise negative shift in δ¹³C_carb during the Late Permian–Early Triassic, with the shift preceding the end-Permian crisis being larger (> 3‰) than that following it (1–2‰). The pre-crisis shifts in δ¹³C_carb are widely correlatable and, hence, represent perturbations to the global carbon cycle. The comparatively smaller shifts following the crisis demonstrate that the marine mass extinction event itself had at most limited influence on the global carbon cycle, and that both Late Permian δ¹³C_carb shifts and the mass extinction must be attributed to some other cause. Their origin cannot be uniquely determined from C-isotopic data alone but appears to be most compatible with a mechanism based on episodic volcanism in combination with collapse of terrestrial ecosystems and soil erosion.     

The human TTAGGG repeat factors 1 and 2 bind to a subset of interstitial telomeric sequences and satellite repeats

The study of the proteins that bind to telomeric DNA in mammals has provided a deep understanding of the mechanisms involved in chromosome-end protection. However, very little is known on the binding of these proteins to nontelomeric DNA sequences. The TTAGGG DNA repeat proteins 1 and 2 (TRF1 and TRF2) bind to mammalian telomeres as part of the shelterin complex and are essential for maintaining chromosome end stability. In this study, we combined chromatin immunoprecipitation with high-throughput sequencing to map at high sensitivity and resolution the human chromosomal sites to which TRF1 and TRF2 bind. While most of the identified sequences correspond to telomeric regions, we showed that these two proteins also bind to extratelomeric sites. The vast majority of these extratelomeric sites contains interstitial telomeric sequences (or ITSs). However, we also identified non-ITS sites, which correspond to centromeric and pericentromeric satellite DNA. Interestingly, the TRF-binding sites are often located in the proximity of genes or within introns. We propose that TRF1 and TRF2 couple the functional state of telomeres to the long-range organization of chromosomes and gene regulation networks by binding to extratelomeric sequences.     

Quantitative in vivo analyses reveal calcium-dependent phosphorylation sites and identifies a novel component of the Toxoplasma invasion motor complex

Apicomplexan parasites depend on the invasion of host cells for survival and proliferation. Calcium-dependent signaling pathways appear to be essential for micronemal release and gliding motility, yet the target of activated kinases remains largely unknown. We have characterized calcium-dependent phosphorylation events during Toxoplasma host cell invasion. Stimulation of live tachyzoites with Ca²⁺-mobilizing drugs leads to phosphorylation of numerous parasite proteins, as shown by differential 2-DE display of ³²[P]-labeled protein extracts. Multi-dimensional Protein Identification Technology (MudPIT) identified ∼546 phosphorylation sites on over 300 Toxoplasma proteins, including 10 sites on the actomyosin invasion motor. Using a Stable Isotope of Amino Acids in Culture (SILAC)-based quantitative LC-MS/MS analyses we monitored changes in the abundance and phosphorylation of the invasion motor complex and defined Ca²⁺-dependent phosphorylation patterns on three of its components - GAP45, MLC1 and MyoA. Furthermore, calcium-dependent phosphorylation of six residues across GAP45, MLC1 and MyoA is correlated with invasion motor activity. By analyzing proteins that appear to associate more strongly with the invasion motor upon calcium stimulation we have also identified a novel 15-kDa Calmodulin-like protein that likely represents the MyoA Essential Light Chain of the Toxoplasma invasion motor. This suggests that invasion motor activity could be regulated not only by phosphorylation but also by the direct binding of calcium ions to this new component.     

The SARS-coronavirus-host interactome: identification of cyclophilins as target for pan-coronavirus inhibitors

Coronaviruses (CoVs) are important human and animal pathogens that induce fatal respiratory, gastrointestinal and neurological disease. The outbreak of the severe acute respiratory syndrome (SARS) in 2002/2003 has demonstrated human vulnerability to (Coronavirus) CoV epidemics. Neither vaccines nor therapeutics are available against human and animal CoVs. Knowledge of host cell proteins that take part in pivotal virus-host interactions could define broad-spectrum antiviral targets. In this study, we used a systems biology approach employing a genome-wide yeast-two hybrid interaction screen to identify immunopilins (PPIA, PPIB, PPIH, PPIG, FKBP1A, FKBP1B) as interaction partners of the CoV non-structural protein 1 (Nsp1). These molecules modulate the Calcineurin/NFAT pathway that plays an important role in immune cell activation. Overexpression of NSP1 and infection with live SARS-CoV strongly increased signalling through the Calcineurin/NFAT pathway and enhanced the induction of interleukin 2, compatible with late-stage immunopathogenicity and long-term cytokine dysregulation as observed in severe SARS cases. Conversely, inhibition of cyclophilins by cyclosporine A (CspA) blocked the replication of CoVs of all genera, including SARS-CoV, human CoV-229E and -NL-63, feline CoV, as well as avian infectious bronchitis virus. Non-immunosuppressive derivatives of CspA might serve as broad-range CoV inhibitors applicable against emerging CoVs as well as ubiquitous pathogens of humans and livestock.     

Na+,K+-pump stimulation improves contractility in isolated muscles of mice with hyperkalemic periodic paralysis

In patients with hyperkalemic periodic paralysis (HyperKPP), attacks of muscle weakness or paralysis are triggered by K(+) ingestion or rest after exercise. Force can be restored by muscle work or treatment with β(2)-adrenoceptor agonists. A missense substitution corresponding to a mutation in the skeletal muscle voltage-gated Na(+) channel (Na(v)1.4, Met1592Val) causing human HyperKPP was targeted into the mouse SCN4A gene (mutants). In soleus muscles prepared from these mutant mice, twitch, tetanic force, and endurance were markedly reduced compared with soleus from wild type (WT), reflecting impaired excitability. In mutant soleus, contractility was considerably more sensitive than WT soleus to inhibition by elevated [K(+)](o). In resting mutant soleus, tetrodotoxin (TTX)-suppressible (22)Na uptake and [Na(+)](i) were increased by 470 and 58%, respectively, and membrane potential was depolarized (by 16 mV, P < 0.0001) and repolarized by TTX. Na(+),K(+) pump-mediated (86)Rb uptake was 83% larger than in WT. Salbutamol stimulated (86)Rb uptake and reduced [Na(+)](i) both in mutant and WT soleus. Stimulating Na(+),K(+) pumps with salbutamol restored force in mutant soleus and extensor digitorum longus (EDL). Increasing [Na(+)](i) with monensin also restored force in soleus. In soleus, EDL, and tibialis anterior muscles of mutant mice, the content of Na(+),K(+) pumps was 28, 62, and 33% higher than in WT, respectively, possibly reflecting the stimulating effect of elevated [Na(+)](i) on the synthesis of Na(+),K(+) pumps. The results confirm that the functional disorders of skeletal muscles in HyperKPP are secondary to increased Na(+) influx and show that contractility can be restored by acute stimulation of the Na(+),K(+) pumps. Calcitonin gene-related peptide (CGRP) restored force in mutant soleus but caused no detectable increase in (86)Rb uptake. Repeated excitation and capsaicin also restored contractility, possibly because of the release of endogenous CGRP from nerve endings in the isolated muscles. These observations may explain how mild exercise helps locally to prevent severe weakness during an attack of HyperKPP.