Aging alters the metabolic flux signature of the ER‐unfolded protein response in vivo in mice

Age is a risk factor for numerous diseases, including neurodegenerative diseases, cancers, and diabetes. Loss of protein homeostasis is a central hallmark of aging. Activation of the endoplasmic reticulum unfolded protein response (UPR^ER) includes changes in protein translation and membrane lipid synthesis. Using stable isotope labeling, a flux “signature” of the UPR^ER in vivo in mouse liver was developed by inducing ER stress with tunicamycin and measuring rates of both proteome‐wide translation and de novo lipogenesis. Several changes in protein synthesis across ontologies were noted with age, including a more dramatic suppression of translation under ER stress in aged mice as compared with young mice. Binding immunoglobulin protein (BiP) synthesis rates and mRNA levels were increased more in aged than young mice. De novo lipogenesis rates decreased under ER stress conditions in aged mice, including both triglyceride and phospholipid fractions. In young mice, a significant reduction was seen only in the triglyceride fraction. These data indicate that aged mice have an exaggerated metabolic flux response to ER stress, which may indicate that aging renders the UPR^ER less effective in resolving proteotoxic stress.     

Influence of Genetic Variation on Plasma Protein Levels in Older Adults Using a Multi-Analyte Panel

Proteins, widely studied as potential biomarkers, play important roles in numerous physiological functions and diseases. Genetic variation may modulate corresponding protein levels and point to the role of these variants in disease pathophysiology. Effects of individual single nucleotide polymorphisms (SNPs) within a gene were analyzed for corresponding plasma protein levels using genome-wide association study (GWAS) genotype data and proteomic panel data with 132 quality-controlled analytes from 521 Caucasian participants in the Alzheimer’s Disease Neuroimaging Initiative (ADNI) cohort. Linear regression analysis detected 112 significant (Bonferroni threshold p = 2.44×10⁻⁵) associations between 27 analytes and 112 SNPs. 107 out of these 112 associations were tested in the Indiana Memory and Aging Study (IMAS) cohort for replication and 50 associations were replicated at uncorrected p<0.05 in the same direction of effect as those in the ADNI. We identified multiple novel associations including the association of rs7517126 with plasma complement factor H-related protein 1 (CFHR1) level at p<1.46×10⁻⁶⁰, accounting for 40 percent of total variation of the protein level. We serendipitously found the association of rs6677604 with the same protein at p<9.29×10⁻¹¹². Although these two SNPs were not in the strong linkage disequilibrium, 61 percent of total variation of CFHR1 was accounted for by rs6677604 without additional variation by rs7517126 when both SNPs were tested together. 78 other SNP-protein associations in the ADNI sample exceeded genome-wide significance (5×10⁻⁸). Our results confirmed previously identified gene-protein associations for interleukin-6 receptor, chemokine CC-4, angiotensin-converting enzyme, and angiotensinogen, although the direction of effect was reversed in some cases. This study is among the first analyses of gene-protein product relationships integrating multiplex-panel proteomics and targeted genes extracted from a GWAS array. With intensive searches taking place for proteomic biomarkers for many diseases, the role of genetic variation takes on new importance and should be considered in interpretation of proteomic results.     

Use of a Molecular Decoy to Segregate Transport from Antigenicity in the FrpB Iron Transporter from Neisseria meningitidis

FrpB is an outer membrane transporter from Neisseria meningitidis, the causative agent of meningococcal meningitis. It is a member of the TonB-dependent transporter (TBDT) family and is responsible for iron uptake into the periplasm. FrpB is subject to a high degree of antigenic variation, principally through a region of hypervariable sequence exposed at the cell surface. From the crystal structures of two FrpB antigenic variants, we identify a bound ferric ion within the structure which induces structural changes on binding which are consistent with it being the transported substrate. Binding experiments, followed by elemental analysis, verified that FrpB binds Fe³⁺ with high affinity. EPR spectra of the bound Fe³⁺ ion confirmed that its chemical environment was consistent with that observed in the crystal structure. Fe³⁺ binding was reduced or abolished on mutation of the Fe³⁺-chelating residues. FrpB orthologs were identified in other Gram-negative bacteria which showed absolute conservation of the coordinating residues, suggesting the existence of a specific TBDT sub-family dedicated to the transport of Fe³⁺. The region of antigenic hypervariability lies in a separate, external sub-domain, whose structure is conserved in both the F3-3 and F5-1 variants, despite their sequence divergence. We conclude that the antigenic sub-domain has arisen separately as a result of immune selection pressure to distract the immune response from the primary transport function. This would enable FrpB to function as a transporter independently of antibody binding, by using the antigenic sub-domain as a ‘molecular decoy’ to distract immune surveillance.     

The Flexibility of Alternating dA—dT Sequences

Abstract

The flexibility of alternating poly (dA—dT) has been investigated by the technique of transient electric dichroism. Rotational relaxation times, which are very sensitive to changes in the end-to-end length of flexible polymers, are determined from the field free dichroism decay curves of four, well defined fragments of poly (dA—dT) ranging in size from 136 to 270 base pairs. Persistence lengths, calculated from the results of Hagerman and Zimm (Biopolymers (1981) 29, 1481–1502), are in the range 200–250 A. This makes alternating dA—dT sequences about twice as flexible as naturally occurring, “random” sequence DNA. Considering a bend around a nucleosome, for example, this difference in persistence length translates to an energy difference between poly (dA—dT) and random sequence DNA of 0. 17 kT/base pair or 1 kcal per 10 base pair stretch. This energy difference is sufficiently large to suggest that dA—dT sequences could serve as markers in DNA packaging, for example, at sites where DNA must tightly bend to accommodate structures.     

Supernumerary teeth in a subadult rhino mandible (<Emphasis Type="Italic">Stephanorhinus hundsheimensis</Emphasis>) from the middle Pleistocene of Mosbach in Wiesbaden (Germany)

A well preserved subadult rhino mandible from Mosbach 2 can be attributed toStephanorhinus hundsheimensis based on a metrical and morphological analysis. A comparison to tooth eruption of livingDiceros bicornis suggests an individual age for the animal of about 7 years at death. The described mandible shows a significant tooth anomaly: two teeth occupy the p3 position on each side of the mandible. Comparisons with three younger juvenileStephanorhinus hundsheimensis from Mosbach 2 show the sequence of tooth eruption for the species and allow us to determine that the anomalous teeth are not persistent milk teeth but are supernumerary teeth, which are morphologically intermediate between normal p2 and p3. The animal’s occlusion was compromised to some degree by the anomaly, and the functional disadvantage may have been critical during a harsh period.      

The stationary-phase sigma factor σS (RpoS) is required for a sustained acid tolerance response in virulent Salmonella typhimurium

The acid tolerance response (ATR) of log-phase Salmonella typhimurium is induced by acid exposures below pH 4.5 and will protect cells against more extreme acid. Two systems are evident: a transiently induced system dependent on the iron regulator Fur that provides a moderate degree of acid tolerance and a more effective sustained ATR that requires the alternate sigma factor σ^S encoded by rpoS. Differences between the acid responses of virulent S. typhimurium and the attenuated laboratory strain LT2 were attributed to disparate levels of RpoS caused by different translational starts. The sustained ATR includes seven newly identified acid shock proteins (ASPs) that are dependent upon σ^S for their synthesis. It is predicted that one or more of these ASPs is essential for the sustained system. The sustained ATR also provided cross-protection to a variety of other environmental stresses (heat, H₂O₂ and osmolarity); however, adaptation to the other stresses did not provide significant acid tolerance. Therefore, in addition to starvation, acid shock serves as an important signal for inducing general stress resistance. Consistent with this model, σ^S proved to be induced by acid shock. Our results also revealed a connection between the transient and sustained ATR systems. Mutations in the regulator atbR are known to cause the overproduction of ten proteins, of which one or more can suppress the acid tolerance defect of an rpoS mutant. One member of the AtbR regulon, designated atrB, was found to be co-regulated by σ^S and AtbR. Both regulators had a negative effect on atrB expression. The results suggest AtrB serves as a link between the sustained and transient ATR systems. When σ^S concentration are low, a compensatory increase in AtrB is required to engage the transiently induced, RpoS-independent system of acid tolerance. Results also suggest different acid-sensitive targets occur in log-phase versus stationary-phase cells.