Tertiary contacts control switching of the SAM-I riboswitch

Riboswitches are non-coding RNAs that control gene expression by sensing small molecules through changes in secondary structure. While secondary structure and ligand interactions are thought to control switching, the exact mechanism of control is unknown. Using a novel two-piece assay that competes the anti-terminator against the aptamer, we directly monitor the process of switching. We find that the stabilization of key tertiary contacts controls both aptamer domain collapse and the switching of the SAM-I riboswitch from the aptamer to the expression platform conformation. Our experiments demonstrate that SAM binding induces structural alterations that indirectly stabilize the aptamer domain, preventing switching toward the expression platform conformer. These results, combined with a variety of structural probing experiments performed in this study, show that the collapse and stabilization of the aptamer domain are cooperative, relying on the sum of key tertiary contacts and the bimodal stability of the kink-turn motif for function. Here, ligand binding serves to shift the equilibrium of aptamer domain structures from a more open toward a more stable collapsed form by stabilizing tertiary interactions. Our data show that the thermodynamic landscape for riboswitch operation is finely balanced to allow large conformational rearrangements to be controlled by small molecule interactions.     

Dissecting the influence of Mg2+ on 3D architecture and ligand-binding of the guanine-sensing riboswitch aptamer domain

Long-range tertiary interactions determine the three-dimensional structure of a number of metabolite-binding riboswitch RNA elements and were found to be important for their regulatory function. For the guanine-sensing riboswitch of the Bacillus subtilis xpt-pbuX operon, our previous NMR-spectroscopic studies indicated pre-formation of long-range tertiary contacts in the ligand-free state of its aptamer domain. Loss of the structural pre-organization in a mutant of this RNA (G37A/C61U) resulted in the requirement of Mg²⁺ for ligand binding. Here, we investigate structural and stability aspects of the wild-type aptamer domain (Gsw) and the G37A/C61U-mutant (Gsw^loop) of the guanine-sensing riboswitch and their Mg²⁺-induced folding characteristics to dissect the role of long-range tertiary interactions, the link between pre-formation of structural elements and ligand-binding properties and the functional stability. Destabilization of the long-range interactions as a result of the introduced mutations for Gsw^loop or the increase in temperature for both Gsw and Gsw^loop involves pronounced alterations of the conformational ensemble characteristics of the ligand-free state of the riboswitch. The increased flexibility of the conformational ensemble can, however, be compensated by Mg²⁺. We propose that reduction of conformational dynamics in remote regions of the riboswitch aptamer domain is the minimal pre-requisite to pre-organize the core region for specific ligand binding.     

Metal-ion binding and metal-ion induced folding of the adenine-sensing riboswitch aptamer domain

Divalent cations are important in the folding and stabilization of complex RNA structures. The adenine-sensing riboswitch controls the expression of mRNAs for proteins involved in purine metabolism by directly sensing intracellular adenine levels. Adenine binds with high affinity and specificity to the ligand binding or aptamer domain of the adenine-sensing riboswitch. The X-ray structure of this domain in complex with adenine revealed an intricate RNA-fold consisting of a three-helix junction stabilized by long-range base-pairing interactions and identified five binding sites for hexahydrated Mg²⁺-ions. Furthermore, a role for Mg²⁺-ions in the ligand-induced folding of this RNA was suggested. Here, we describe the interaction of divalent cations with the RNA–adenine complex in solution as studied by high-resolution NMR spectroscopy. Paramagnetic line broadening, chemical shift mapping and intermolecular nuclear Overhauser effects (NOEs) indicate the presence of at least three binding sites for divalent cations. Two of them are similar to those in the X-ray structure. The third site, which is important for the folding of this RNA, has not been observed previously. The ligand-free state of the RNA is conformationally heterogeneous and contains base-pairing patterns detrimental to ligand binding in the absence of Mg²⁺, but becomes partially pre-organized for ligand binding in the presence of Mg²⁺. Compared to the highly similar guanine-sensing riboswitch, the folding pathway for the adenine-sensing riboswitch aptamer domain is more complex and the influence of Mg²⁺ is more pronounced.     

L-type Ca2+ channel function is linked to dystrophin expression in mammalian muscle

Background

In dystrophic mdx skeletal muscle, aberrant Ca²⁺ homeostasis and fibre degeneration are found. The absence of dystrophin in models of Duchenne muscular dystrophy (DMD) has been connected to altered ion channel properties e.g. impaired L-type Ca²⁺ currents. In regenerating mdx muscle, ‘revertant’ fibres restore dystrophin expression. Their functionality involving DHPR-Ca²⁺-channels is elusive.

Methods and Results

We developed a novel ‘in-situ’ confocal immuno-fluorescence and imaging technique that allows, for the first time, quantitative subcellular dystrophin-DHPR colocalization in individual, non-fixed, muscle fibres. Tubular DHPR signals alternated with second harmonic generation signals originating from myosin. Dystrophin-DHPR colocalization was substantial in wt fibres, but diminished in most mdx fibres. Mini-dystrophin (MinD) expressing fibres successfully restored colocalization. Interestingly, in some aged mdx fibres, colocalization was similar to wt fibres. Most mdx fibres showed very weak membrane dystrophin staining and were classified ‘mdx-like’. Some mdx fibres, however, had strong ‘wt-like’ dystrophin signals and were identified as ‘revertants’. Split mdx fibres were mostly ‘mdx-like’ and are not generally ‘revertants’. Correlations between membrane dystrophin and DHPR colocalization suggest a restored putative link in ‘revertants’. Using the two-micro-electrode-voltage clamp technique, Ca²⁺-current amplitudes (i_max) showed very similar behaviours: reduced amplitudes in most aged mdx fibres (as seen exclusively in young mdx mice) and a few mdx fibres, most likely ‘revertants’, with amplitudes similar to wt or MinD fibres. Ca²⁺ current activation curves were similar in ‘wt-like’ and ‘mdx-like’ aged mdx fibres and are not the cause for the differences in current amplitudes. i_max amplitudes were fully restored in MinD fibres.

Conclusions

We present evidence for a direct/indirect DHPR-dystrophin interaction present in wt, MinD and ‘revertant’ mdx fibres but absent in remaining mdx fibres. Our imaging technique reliably detects single isolated ‘revertant’ fibres that could be used for subsequent physiological experiments to study mechanisms and therapy concepts in DMD.     

Structural and functional insight into the mechanism of an alkaline exonuclease from Laribacter hongkongensis

Alkaline exonuclease and single-strand DNA (ssDNA) annealing proteins (SSAPs) are key components of DNA recombination and repair systems within many prokaryotes, bacteriophages and virus-like genetic elements. The recently sequenced β-proteobacterium Laribacter hongkongensis (strain HLHK9) encodes putative homologs of alkaline exonuclease (LHK-Exo) and SSAP (LHK-Bet) proteins on its 3.17 Mb genome. Here, we report the biophysical, biochemical and structural characterization of recombinant LHK-Exo protein. LHK-Exo digests linear double-stranded DNA molecules from their 5′-termini in a highly processive manner. Exonuclease activities are optimum at pH 8.2 and essentially require Mg²⁺ or Mn²⁺ ions. 5′-phosphorylated DNA substrates are preferred over dephosphorylated ones. The crystal structure of LHK-Exo was resolved to 1.9 Å, revealing a ‘doughnut-shaped’ toroidal trimeric arrangement with a central tapered channel, analogous to that of λ-exonuclease (Exo) from bacteriophage-λ. Active sites containing two bound Mg²⁺ ions on each of the three monomers were located in clefts exposed to this central channel. Crystal structures of LHK-Exo in complex with dAMP and ssDNA were determined to elucidate the structural basis for substrate recognition and binding. Through structure-guided mutational analysis, we discuss the roles played by various active site residues. A conserved two metal ion catalytic mechanism is proposed for this class of alkaline exonucleases.     

HMGB‐1 induces c‐kit+ cell microvascular rolling and adhesion via both toll‐like receptor‐2 and toll‐like receptor‐4 of endothelial cells

High-mobility group box 1 (HMGB-1) is a strong chemo-attractive signal for both inflammatory and stem cells. The aim of this study is to evaluate the mechanisms regulating HMGB-1–mediated adhesion and rolling of c-kit⁺ cells and assess whether toll-like receptor-2 (TLR-2) and toll-like receptor-4 (TLR-4) of endothelial cells or c-kit⁺ cells are implicated in the activation of downstream migration signals to peripheral c-kit⁺ cells. Effects of HMGB-1 on the c-kit⁺ cells/endothelial interaction were evaluated by a cremaster muscle model in wild-type (WT), TLR-2 (−/−) and Tlr4 (LPS-del) mice. The mRNA and protein expression levels of endothelial nitric oxide synthase were determined by quantitative real-time PCR and immunofluorescence staining. Induction of crucial adhesion molecules for rolling and adhesion of stem cells and leukocytes were monitored in vivo and in vitro. Following local HMGB-1 administration, a significant increase in cell rolling was detected (32.4 ± 7.1% in ‘WT’ versus 9.9 ± 3.2% in ‘control’, P < 0.05). The number of firmly adherent c-kit⁺ cells was more than 13-fold higher than that of the control group (14.6 ± 5.1 cells/mm² in ‘WT’ versus 1.1 ± 1.0 cells/mm² in ‘control’, P < 0.05). In knockout animals, the fraction of rolling cells did not differ significantly from control levels. Firm endothelial adhesion was significantly reduced in TLR-2 (−/−) and Tlr4 (LPS-del) mice compared to WT mice (1.5 ± 1.4 cells/mm² in ‘TLR-2 (−/−)’ and 2.4 ± 1.4 cells/mm² in ‘Tlr4 (LPS-del)’ versus 14.6 ± 5.1 cells/mm² in ‘WT’, P < 0.05). TLR-2 (−/−) and Tlr4 (LPS-del) stem cells in WT mice did not show significant reduction in rolling and adhesion compared to WT cells. HMGB-1 mediates c-kit⁺ cell recruitment via endothelial TLR-2 and TLR-4.     

Characterization of MicA interactions suggests a potential novel means of gene regulation by small non-coding RNAs

MicA is a small non-coding RNA that regulates ompA mRNA translation in Escherichia coli. MicA has an inhibitory function, base pairing to the translation initiation region of target mRNAs through short sequences of complementarity, blocking their ribosome-binding sites. The MicA structure contains two stem loops, which impede its interaction with target mRNAs, and it is thought that the RNA chaperone protein Hfq, known to be involved in MicA regulation of ompA, may structurally remodel MicA to reveal the ompA-binding site for cognate pairing. To further characterize these interactions, we undertook biochemical and biophysical studies using native MicA and a ‘stabilized’ version, modified to mimic the conformational state of MicA where the ompA-binding site is exposed. Our data corroborate two proposed roles for Hfq: first, to bring both MicA and ompA into close proximity, and second, to restructure MicA to allow exposure of the ompA-binding site for pairing, thereby demonstrating the RNA chaperone function of Hfq. Additionally, at accumulated MicA levels, we identified a Mg²⁺-dependent self-association that occludes the ompA-recognition region. We discuss the potential contribution of an Mg²⁺-mediated conformational switch of MicA for the regulation of MicA function.     

The infection processes of Sclerotinia sclerotiorum in cotyledon tissue of a resistant and a susceptible genotype of Brassica napus

Background and Aims

Sclerotinia sclerotiorum can attack >400 plant species worldwide. Very few studies have investigated host–pathogen interactions at the plant surface and cellular level in resistant genotypes of oilseed rape/canola (Brassica napus).

Methods

Infection processes of S. sclerotiorum were examined on two B. napus genotypes, one resistant cultivar ‘Charlton’ and one susceptible ‘RQ001-02M2’ by light and scanning electron microscopy from 2 h to 8 d post-inoculation (dpi).

Key Results

The resistant ‘Charlton’ impeded fungal growth at 1, 2 and 3 dpi, suppressed formation of appresoria and infection cushions, caused extrusion of protoplast from hyphal cells and produced a hypersensitive reaction. At 8 dpi, whilst in ‘Charlton’ pathogen invasion was mainly confined to the upper epidermis, in the susceptible ‘RQ001-02M2’, colonization up to the spongy mesophyll cells was evident. Calcium oxalate crystals were found in the upper epidermis and in palisade cells in susceptible ‘RQ001-02M2’ at 6 dpi, and throughout leaf tissues at 8 dpi. In resistant ‘Charlton’, crystals were not observed at 6 dpi, whereas at 8 dpi they were mainly confined to the upper epidermis. Starch deposits were also more prevalent in ‘RQ001-02M2’.

Conclusions

This study demonstrates for the first time at the cellular level that resistance to S. sclerotiorum in B. napus is a result of retardation of pathogen development, both on the plant surface and within host tissues. The resistance mechanisms identified in this study will be useful for engineering disease-resistant genotypes and for developing markers for screening for resistance against this pathogen.     

Frugivory and spatial patterns of seed deposition by carnivorous mammals in anthropogenic landscapes: a multi-scale approach

Background

Knowledge about how frugivory and seed deposition are spatially distributed is valuable to understand the role of dispersers on the structure and dynamics of plant populations. This may be particularly important within anthropogenic areas, where either the patchy distribution of wild plants or the presence of cultivated fleshy-fruits may influence plant-disperser interactions.

Methodology/Principal Findings

We investigated frugivory and spatial patterns of seed deposition by carnivorous mammals in anthropogenic landscapes considering two spatial scales: ‘landscape’ (∼10 km²) and ‘habitat type’ (∼1–2 km²). We sampled carnivore faeces and plant abundance at three contrasting habitats (chestnut woods, mosaics and scrublands), each replicated within three different landscapes. Sixty-five percent of faeces collected (n = 1077) contained seeds, among which wild and cultivated seeds appeared in similar proportions (58% and 53%) despite that cultivated fruiting plants were much less abundant. Seed deposition was spatially structured among both spatial scales being different between fruit types. Whereas the most important source of spatial variation in deposition of wild seeds was the landscape scale, it was the habitat scale for cultivated seeds. At the habitat scale, seeds of wild species were mostly deposited within mosaics while seeds of cultivated species were within chestnut woods and scrublands. Spatial concordance between seed deposition and plant abundance was found only for wild species.

Conclusions/Significance

Spatial patterns of seed deposition by carnivores differed between fruit types and seemed to be modulated by the fleshy-fruited plant assemblages and the behaviour of dispersers. Our results suggest that a strong preference for cultivated fruits by carnivores may influence their spatial foraging behaviour and lower their dispersal services to wild species. However, the high amount of seeds removed within and between habitats suggests that carnivores must play an important role – often overlooked – as ‘restorers’ and ‘habitat shapers’ in anthropogenic areas.     

Cation-dependent cleavage of the duplex form of the subtype-B HIV-1 RNA dimerization initiation site

The crystal structure of subtype-B HIV-1 genomic RNA Dimerization Initiation Site duplex revealed chain cleavage at a specific position resulting in 3′-phosphate and 5′-hydroxyl termini. A crystallographic analysis showed that Ba²⁺, Mn²⁺, Co²⁺ and Zn²⁺ bind specifically on a guanine base close to the cleaved position. The crystal structures also point to a necessary conformational change to induce an ‘in-line’ geometry at the cleavage site. In solution, divalent cations increased the rate of cleavage with pH/pKa compensation, indicating that a cation-bound hydroxide anion is responsible for the cleavage. We propose a ‘Trojan horse’ mechanism, possibly of general interest, wherein a doubly charged cation hosted near the cleavage site as a ‘harmless’ species is further transformed in situ into an ‘aggressive’ species carrying a hydroxide anion.     

Incoming nucleotide binds to Klenow ternary complex leading to stable physical sequestration of preceding dNTP on DNA

Klenow–DNA complex is known to undergo a rate-limiting, protein conformational transition from an ‘open’ to ‘closed’ state, upon binding of the ‘correct’ dNTP at the active site. In the ‘closed’ state, Mg²⁺ mediates a rapid chemical step involving nucleophilic displacement of pyrophosphate by the 3′ hydroxyl of the primer terminus. The enzyme returns to the ‘open’ state upon the release of PPi and translocation permits the next round of reaction. To determine whether Klenow can translocate to the next site on the addition of the next dNTP, without the preceding chemical step, we studied the ternary complex (Klenow–DNA–dNTP) in the absence of Mg²⁺. While the ternary complex is proficient in chemical addition of dNTPs in Mg²⁺, as revealed by primer extensions, the same in Mg²⁺-deficient conditions lead to non-covalent (physical) sequestration of first two ‘correct’ dNTPs in the ternary complex. Moreover, the second dNTP traps the first one in the DNA-helix of the ternary complex. Such a dNTP–DNA complex is found to be stable even after the dissociation of Klenow. This reveals the novel state of the dNTP–DNA complex where the complementary base is stacked in a DNA-helix non-covalently, without the phosphodiester linkage. Further, shuttling of the DNA between the polymerase and the exonuclease site mediates the release of such a DNA complex. Interestingly, Klenow in such a Mg²⁺-deficient ternary complex exhibits a ‘closed’ conformation.     

A magnesium-induced RNA conformational switch at the internal ribosome entry site of hepatitis C virus genome visualized by atomic force microscopy

The 5′ untranslated region of hepatitis C virus (HCV) genomic RNA contains an internal ribosome entry site (IRES) element, composed of domains II–IV, which is required for cap-independent translation initiation. Little information on the 3D structure of the whole functional HCV IRES is still available. Here, we use atomic force microscopy to visualize the HCV IRES conformation in its natural sequence context, which includes the upstream domain I and the essential, downstream domains V and VI. The 574 nt-long molecule analyzed underwent an unexpected, Mg²⁺-induced switch between two alternative conformations: from ‘open’, elongated morphologies at 0–2 mM Mg²⁺ concentration to a ‘closed’, comma-shaped conformation at 4–6 mM Mg²⁺. This sharp transition, confirmed by gel-shift analysis and partial RNase T1 cleavage, was hindered by the microRNA miR-122. The comma-shaped IRES-574 molecules visualized at 4–6 mM Mg²⁺ in the absence of miR-122 showed two arms. Our data support that the first arm would contain domain III, while the second one would be composed of domains (I–II)+(V–VI) thanks to a long-range RNA interaction between the I-II spacer and the basal region of domain VI. This reinforces the previously described structural continuity between the HCV IRES and its flanking domains I, V and VI.     

HIV-1 DIS stem loop forms an obligatory bent kissing intermediate in the dimerization pathway

The HIV-1 dimerization initiation sequence (DIS) is a conserved palindrome in the apical loop of a conserved hairpin motif in the 5′-untranslated region of its RNA genome. DIS hairpin plays an important role in genome dimerization by forming a ‘kissing complex’ between two complementary hairpins. Understanding the kinetics of this interaction is key to exploiting DIS as a possible human immunodeficiency virus (HIV) drug target. Here, we present a single-molecule Förster resonance energy transfer (smFRET) study of the dimerization reaction kinetics. Our data show the real-time formation and dissociation dynamics of individual kissing complexes, as well as the formation of the mature extended duplex complex that is ultimately required for virion packaging. Interestingly, the single-molecule trajectories reveal the presence of a previously unobserved bent intermediate required for extended duplex formation. The universally conserved A272 is essential for the formation of this intermediate, which is stabilized by Mg²⁺, but not by K⁺ cations. We propose a 3D model of a possible bent intermediate and a minimal dimerization pathway consisting of three steps with two obligatory intermediates (kissing complex and bent intermediate) and driven by Mg²⁺ ions.     

Fluctuating Helical Asymmetry and Morphology of Snails (Gastropoda) in Divergent Microhabitats at 'Evolution Canyons I and II,' Israel

Background

Developmental instability of shelled gastropods is measured as deviations from a perfect equiangular (logarithmic) spiral. We studied six species of gastropods at ‘Evolution Canyons I and II’ in Carmel and the Galilee Mountains, Israel, respectively. The xeric, south-facing, ‘African’ slopes and the mesic, north-facing, ‘European’ slopes have dramatically different microclimates and plant communities. Moreover, ‘Evolution Canyon II’ receives more rainfall than ‘Evolution Canyon I.’

Methodology/Principal Findings

We examined fluctuating asymmetry, rate of whorl expansion, shell height, and number of rotations of the body suture in six species of terrestrial snails from the two ‘Evolution Canyons.’ The xeric ‘African’ slope should be more stressful to land snails than the ‘European’ slope, and ‘Evolution Canyon I’ should be more stressful than ‘Evolution Canyon II.’ Only Eopolita protensa jebusitica showed marginally significant differences in fluctuating helical asymmetry between the two slopes. Contrary to expectations, asymmetry was marginally greater on the ‘European’ slope. Shells of Levantina spiriplana caesareana at ‘Evolution Canyon I,’ were smaller and more asymmetric than those at ‘Evolution Canyon II.’ Moreover, shell height and number of rotations of the suture were greater on the north-facing slopes of both canyons.

Conclusions/Significance

Our data is consistent with a trade-off between drought resistance and thermoregulation in snails; Levantina was significantly smaller on the ‘African’ slope, for increasing surface area and thermoregulation, while Eopolita was larger on the ‘African’ slope, for reducing water evaporation. In addition, ‘Evolution Canyon I’ was more stressful than Evolution Canyon II’ for Levantina.     

Modulation of P2X4 pore closure by magnesium,potassium, and ATP

The P2X4 receptor plays a prominent role in cellular responses to extracellular ATP. Through classical all-atom molecular dynamics (MD) simulations totaling 24 μs we have investigated how metal-complexed ATP stabilizes the channel's open state and prevents its closing. We have identified two metal-binding sites, Mg²⁺ and potassium K⁺, one at the intersection of the three subunits in the ectodomain (MBS1) and the second one near the ATP-binding site (MBS2), similar to those characterized in Gulf Coast P2X. Our data indicate that when Mg²⁺ and K⁺ ions are complexed with ATP, the channel is locked into an open state. Interestingly, irrespective of the number of bound ATP molecules, Mg²⁺ ions bound to the MBS2 impeded the collapse of the open-state protein to a closed state by stabilizing the ATP-protein interactions. However, when Mg²⁺ in the MBS2 was replaced with K⁺ ions, as might be expected when in equilibrium with an extracellular solution, the interactions between the subunits were weakened and the pore collapsed. This collapse was apparent when fewer than two ATPs were bound to MBS2 in the presence of K⁺. Therefore, the different capacities of common cations to stabilize the channel may underlie a mechanism governing P2X4 channel gating in physiological systems. This study therefore provides structural insights into the differential modulation of ATP activation of P2X4 by Mg²⁺ and K⁺.     

Signal percolation through plants and the shape of the calcium signature

Plants respond to almost any kind of external stimulus with transients in their cytoplasmic free calcium concentration ([Ca²⁺]_c). A huge variety of kinetics recorded by optical techniques has been reported in the past. This variety has been credited the specificity needed to explain how information about incoming stimuli is evaluated by the organism and turned into the right physiological responses which provide advantages for survival and reproduction. A physiological response often takes place away from the site of stimulation. This requires cell-to-cell communication. Hence, responding cells are not necessarily directly stimulated but rather receive an indirect stimulus via cell-to-cell communication. It appears unlikely that the ‘[Ca²⁺]_c signature’ in the primarily stimulated cell is conveyed over long distances via cell-to-cell communication from the ‘receptor cells’ to the ‘effector cells’. Here, a novel aspect is highlighted to explain the variety of [Ca²⁺] kinetics seen by integrating methods of [Ca²⁺]_c recording. Plants can generally be seen as cellular automata with specific morphology and capable for cell-to-cell communication. Just a few rules are needed to demonstrate how waves of [Ca²⁺]_c-increases percolate through the organism and thereby deliver a broad variety of ‘signatures’. Modelling intercellular signalling may be a possible way to find explanations for different kinds of signal transmission, signal amplification, wave formation, oscillations and stimulus-response coupling. The basic examples presented here show that care has to be taken when interpreting cellular ‘[Ca²⁺]_c signatures’ recorded by optical techniques which integrate over a big number of cells or even whole plants.Key words: cellular automata, cell-to-cell communication, cytoplasmic calcium, modelling, percolation, signature     

Meta‐analysis on blood transcriptomic studies identifies consistently coexpressed protein–protein interaction modules as robust markers of human aging

The bodily decline that occurs with advancing age strongly impacts on the prospects for future health and life expectancy. Despite the profound role of age in disease etiology, knowledge about the molecular mechanisms driving the process of aging in humans is limited. Here, we used an integrative network-based approach for combining multiple large-scale expression studies in blood (2539 individuals) with protein–protein Interaction (PPI) data for the detection of consistently coexpressed PPI modules that may reflect key processes that change throughout the course of normative aging. Module detection followed by a meta-analysis on chronological age identified fifteen consistently coexpressed PPI modules associated with chronological age, including a highly significant module (P = 3.5 × 10⁻³⁸) enriched for ‘T-cell activation’ marking age-associated shifts in lymphocyte blood cell counts (R² = 0.603; P = 1.9 × 10⁻¹⁰). Adjusting the analysis in the compendium for the ‘T-cell activation’ module showed five consistently coexpressed PPI modules that robustly associated with chronological age and included modules enriched for ‘Translational elongation’, ‘Cytolysis’ and ‘DNA metabolic process’. In an independent study of 3535 individuals, four of five modules consistently associated with chronological age, underpinning the robustness of the approach. We found three of five modules to be significantly enriched with aging-related genes, as defined by the GenAge database, and association with prospective survival at high ages for one of the modules including ASF1A. The hereby-detected age-associated and consistently coexpressed PPI modules therefore may provide a molecular basis for future research into mechanisms underlying human aging.     

Association of the variants in the PPARG gene and serum lipid levels: a meta‐analysis of 74 studies

Considerable studies have been carried out to investigate the relationship between the polymorphisms of PPARG (Pro12Ala, C161T and C1431T) and serum lipid levels, but the results were inconclusive. Hence, we conducted a meta-analysis to clarify the association. MEDLINE, EMBASE and the Cochrane Library databases were searched systematically. The subgroup analysis was performed based on ethnicity. Seventy-four studies with 54,953 subjects were included in this meta-analysis. In Pro12Ala, the group with the ‘PP’ (C/C genotype) genotype group had lower levels of total cholesterol (TC) (mean difference, MD: −0.02, P < 0.00001; I² = 28%), low-density lipoprotein cholesterol (LDL-C) (MD: −0.02, P < 0.00001; I² = 30%) and higher levels of triglyceride (TG) (MD: 0.06, P < 0.00001; I² = 30%) than the combined ‘PA+AA’ (PA = C/G genotype, AA = G/G genotype) genotype group in Asian population, and the group with the ‘PP’ genotype had higher levels of TG (MD: 0.07, P < 0.02; I² = 67%) than the combined ‘PA+AA’ genotype group in non-Asian population. No statistically significant differences in the levels of TC, TG, high-density lipoprotein cholesterol, LDL-C were detected between different genotypes in C161T(Asian or non-Asian) and C1431T(Asian) polymorphisms. This meta-analysis was a renewed and confirmed study to assess the association between PPARG polymorphisms and serum lipid levels in Asian and non-Asian populations. There is a prominent association between Pro12Ala polymorphism and the levels of TC, LDL-C and TG in Asian population. No statistically significant differences in serum lipid levels were detected between different genotypes in C161T and C1431T polymorphisms.     

Magnesium-binding architectures in RNA crystal structures: validation,binding preferences,classification and motif detection

The ubiquitous presence of magnesium ions in RNA has long been recognized as a key factor governing RNA folding, and is crucial for many diverse functions of RNA molecules. In this work, Mg²⁺-binding architectures in RNA were systematically studied using a database of RNA crystal structures from the Protein Data Bank (PDB). Due to the abundance of poorly modeled or incorrectly identified Mg²⁺ ions, the set of all sites was comprehensively validated and filtered to identify a benchmark dataset of 15 334 ‘reliable’ RNA-bound Mg²⁺ sites. The normalized frequencies by which specific RNA atoms coordinate Mg²⁺ were derived for both the inner and outer coordination spheres. A hierarchical classification system of Mg²⁺ sites in RNA structures was designed and applied to the benchmark dataset, yielding a set of 41 types of inner-sphere and 95 types of outer-sphere coordinating patterns. This classification system has also been applied to describe six previously reported Mg²⁺-binding motifs and detect them in new RNA structures. Investigation of the most populous site types resulted in the identification of seven novel Mg²⁺-binding motifs, and all RNA structures in the PDB were screened for the presence of these motifs.