Tracking Changes in SARS-CoV-2 Spike: Evidence that D614G Increases Infectivity of the COVID-19 Virus.

A SARS-CoV-2 variant carrying the Spike protein amino acid change D614G has become the most prevalent form in the global pandemic. Dynamic tracking of variant frequencies revealed a recurrent pattern of G614 increase at multiple geographic levels: national, regional, and municipal. The shift occurred even in local epidemics where the original D614 form was well established prior to introduction of the G614 variant. The consistency of this pattern was highly statistically significant, suggesting that the G614 variant may have a fitness advantage. We found that the G614 variant grows to a higher titer as pseudotyped virions. In infected individuals, G614 is associated with lower RT-PCR cycle thresholds, suggestive of higher upper respiratory tract viral loads, but not with increased disease severity. These findings illuminate changes important for a mechanistic understanding of the virus and support continuing surveillance of Spike mutations to aid with development of immunological interventions.

The origin recognition complex requires chromatin tethering by a hypervariable intrinsically disordered region that is functionally conserved from sponge to man.

The first step toward eukaryotic genome duplication is loading of the replicative helicase onto chromatin. This 'licensing' step initiates with the recruitment of the origin recognition complex (ORC) to chromatin, which is thought to occur via ORC's ATP-dependent DNA binding and encirclement activity. However, we have previously shown that ATP binding is dispensable for the chromatin recruitment of fly ORC, raising the question of how metazoan ORC binds chromosomes. We show here that the intrinsically disordered region (IDR) of fly Orc1 is both necessary and sufficient for recruitment of ORC to chromosomes in vivo and demonstrate that this is regulated by IDR phosphorylation. Consistently, we find that the IDR confers the ORC holocomplex with ATP-independent DNA binding activity in vitro. Using phylogenetic analysis, we make the surprising observation that metazoan Orc1 IDRs have diverged so markedly that they are unrecognizable as orthologs and yet we find that these compositionally homologous sequences are functionally conserved. Altogether, these data suggest that chromatin is recalcitrant to ORC's ATP-dependent DNA binding activity, necessitating IDR-dependent chromatin tethering, which we propose poises ORC to opportunistically encircle nucleosome-free regions as they become available.

U6 snRNA m6A modification is required for accurate and efficient splicing of C. elegans and human pre-mRNAs.

pre-mRNA splicing is a critical feature of eukaryotic gene expression. Both cis- and trans-splicing rely on accurately recognising splice site sequences by spliceosomal U snRNAs and associated proteins. Spliceosomal snRNAs carry multiple RNA modifications with the potential to affect different stages of pre-mRNA splicing. Here, we show that the conserved U6 snRNA m6A methyltransferase METT-10 is required for accurate and efficient cis- and trans-splicing of C. elegans pre-mRNAs. The absence of METT-10 in C. elegans and METTL16 in humans primarily leads to alternative splicing at 5' splice sites with an adenosine at +4 position. In addition, METT-10 is required for splicing of weak 3' cis- and trans-splice sites. We identified a significant overlap between METT-10 and the conserved splicing factor SNRNP27K in regulating 5' splice sites with +4A. Finally, we show that editing endogenous 5' splice site +4A positions to +4U restores splicing to wild-type positions in a mett-10 mutant background, supporting a direct role for U6 snRNA m6A modification in 5' splice site recognition. We conclude that the U6 snRNA m6A modification is important for accurate and efficient pre-mRNA splicing.

Identification and functional evaluation of GRIA1 missense and truncation variants in individuals with ID: An emerging neurodevelopmental syndrome.

GRIA1 encodes the GluA1 subunit of α-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA) receptors, which are ligand-gated ion channels that act as excitatory receptors for the neurotransmitter L-glutamate (Glu). AMPA receptors (AMPARs) are homo- or heteromeric protein complexes with four subunits, each encoded by different genes, GRIA1 to GRIA4. Although GluA1-containing AMPARs have a crucial role in brain function, the human phenotype associated with deleterious GRIA1 sequence variants has not been established. Subjects with de novo missense and nonsense GRIA1 variants were identified through international collaboration. Detailed phenotypic and genetic assessments of the subjects were carried out and the pathogenicity of the variants was evaluated in vitro to characterize changes in AMPAR function and expression. In addition, two Xenopus gria1 CRISPR-Cas9 F0 models were established to characterize the in vivo consequences. Seven unrelated individuals with rare GRIA1 variants were identified. One individual carried a homozygous nonsense variant (p.Arg377Ter), and six had heterozygous missense variations (p.Arg345Gln, p.Ala636Thr, p.Ile627Thr, and p.Gly745Asp), of which the p.Ala636Thr variant was recurrent in three individuals. The cohort revealed subjects to have a recurrent neurodevelopmental disorder mostly affecting cognition and speech. Functional evaluation of major GluA1-containing AMPAR subtypes carrying the GRIA1 variant mutations showed that three of the four missense variants profoundly perturb receptor function. The homozygous stop-gain variant completely destroys the expression of GluA1-containing AMPARs. The Xenopus gria1 models show transient motor deficits, an intermittent seizure phenotype, and a significant impairment to working memory in mutants. These data support a developmental disorder caused by both heterozygous and homozygous variants in GRIA1 affecting AMPAR function.

18F-FDG PET/CT and radiolabeled leukocyte SPECT/CT imaging for the evaluation of cardiovascular infection in the multimodality context: ASNC Imaging Indications (ASNC I2) Series Expert Consensus Recommendations from ASNC, AATS, ACC, AHA, ASE, EANM, HRS, IDSA, SCCT, SNMMI, and STS.

This document on cardiovascular infection, including infective endocarditis, is the first in the American Society of Nuclear Cardiology Imaging Indications (ASNC I2) series to assess the role of radionuclide imaging in the multimodality context for the evaluation of complex systemic diseases with multi-societal involvement including pertinent disciplines. A rigorous modified Delphi approach was used to determine consensus clinical indications, diagnostic criteria, and an algorithmic approach to diagnosis of cardiovascular infection including infective endocarditis. Cardiovascular infection incidence is increasing and is associated with high morbidity and mortality. Current strategies based on clinical criteria and an initial echocardiographic imaging approach are effective but often insufficient in complicated cardiovascular infection. Radionuclide imaging with 18F-fluorodeoxyglucose (FDG) positron emission tomography/computed tomography (PET/CT) and single photon emission computed tomography/CT leukocyte scintigraphy can enhance the evaluation of suspected cardiovascular infection by increasing diagnostic accuracy, identifying extracardiac involvement, and assessing cardiac implanted device pockets, leads, and all portions of ventricular assist devices. This advanced imaging can aid in key medical and surgical considerations. Consensus diagnostic features include focal/multi-focal or diffuse heterogenous intense 18F-FDG uptake on valvular and prosthetic material, perivalvular areas, device pockets and leads, and ventricular assist device hardware persisting on non-attenuation corrected images. There are numerous clinical indications with a larger role in prosthetic valves, and cardiac devices particularly with possible infective endocarditis or in the setting of prior equivocal or non-diagnostic imaging. Illustrative cases incorporating these consensus recommendations provide additional clarification. Future research is necessary to refine application of these advanced imaging tools for surgical planning, to identify treatment response, and more.

Subgenomic RNA identification in SARS-CoV-2 genomic sequencing data.

We have developed periscope, a tool for the detection and quantification of subgenomic RNA (sgRNA) in SARS-CoV-2 genomic sequence data. The translation of the SARS-CoV-2 RNA genome for most open reading frames (ORFs) occurs via RNA intermediates termed "subgenomic RNAs." sgRNAs are produced through discontinuous transcription, which relies on homology between transcription regulatory sequences (TRS-B) upstream of the ORF start codons and that of the TRS-L, which is located in the 5' UTR. TRS-L is immediately preceded by a leader sequence. This leader sequence is therefore found at the 5' end of all sgRNA. We applied periscope to 1155 SARS-CoV-2 genomes from Sheffield, United Kingdom, and validated our findings using orthogonal data sets and in vitro cell systems. By using a simple local alignment to detect reads that contain the leader sequence, we were able to identify and quantify reads arising from canonical and noncanonical sgRNA. We were able to detect all canonical sgRNAs at the expected abundances, with the exception of ORF10. A number of recurrent noncanonical sgRNAs are detected. We show that the results are reproducible using technical replicates and determine the optimum number of reads for sgRNA analysis. In VeroE6 ACE2+/- cell lines, periscope can detect the changes in the kinetics of sgRNA in orthogonal sequencing data sets. Finally, variants found in genomic RNA are transmitted to sgRNAs with high fidelity in most cases. This tool can be applied to all sequenced COVID-19 samples worldwide to provide comprehensive analysis of SARS-CoV-2 sgRNA.

Protein intrinsically disordered regions have a non-random, modular architecture.

MOTIVATION: Protein sequences can be broadly categorized into two classes: those which adopt stable secondary structure and fold into a domain (i.e. globular proteins), and those that do not. The sequences belonging to this latter class are conformationally heterogeneous and are described as being intrinsically disordered. Decades of investigation into the structure and function of globular proteins has resulted in a suite of computational tools that enable their sub-classification by domain type, an approach that has revolutionized how we understand and predict protein functionality. Conversely, it is unknown if sequences of disordered protein regions are subject to broadly generalizable organizational principles that would enable their sub-classification. RESULTS: Here, we report the development of a statistical approach that quantifies linear variance in amino acid composition across a sequence. With multiple examples, we provide evidence that intrinsically disordered regions are organized into statistically non-random modules of unique compositional bias. Modularity is observed for both low and high-complexity sequences and, in some cases, we find that modules are organized in repetitive patterns. These data demonstrate that disordered sequences are non-randomly organized into modular architectures and motivate future experiments to comprehensively classify module types and to determine the degree to which modules constitute functionally separable units analogous to the domains of globular proteins. AVAILABILITY AND IMPLEMENTATION: The source code, documentation, and data to reproduce all figures are freely available at https://github.com/MWPlabUTSW/Chi-Score-Analysis.git. The analysis is also available as a Google Colab Notebook (https://colab.research.google.com/github/MWPlabUTSW/Chi-Score-Analysis/blob/main/ChiScore_Analysis.ipynb).

Multiple serum biomarkers associate with mortality and interstitial lung disease progression in systemic sclerosis.

OBJECTIVES: To investigate the prognostic utility of 28 serum biomarkers in systemic sclerosis (SSc), SSc-associated interstitial lung disease (SSc-ILD) and clinically relevant disease subgroups. METHODS: Participants with sera, high-resolution CT and lung function within 12 months of baseline were identified from the Australian Scleroderma Cohort Study. Baseline was the time of serum collection. Twenty-seven of the prespecified 28 serum biomarkers were analysed and biomarker associations with mortality and ILD progression were investigated in univariable and multivariable analyses, including within disease subgroups and combined with established risk factors for poorer prognosis in SSc. RESULTS: A total of 407 participants were identified, 252 (61.9%) with SSc-ILD. The median (interquartile range) follow-up after biomarker measurement was 6.31 (3.11-9.22) years. Sixteen biomarkers were associated with increased mortality. High levels of VCAM-1 were most strongly associated with mortality [hazard ratio (HR) 3.55; 95% CI 2.37-5.33; P < 0.001]. Five additional biomarkers had an HR >2: SP-D (2.28, 1.57-3.31; P < 0.001), E-selectin (2.19, 1.53-3.14; P < 0.001), IL-6 (2.15, 1.50-3.09; P < 0.001), MMP-3 (2.05, 1.42-2.95; P < 0.001) and ET-1 (2.03, 1.40-2.92; P < 0.001). Eleven biomarkers were independently associated with mortality following adjustment for sex, age and baseline forced vital capacity (FVC%predicted). Three biomarkers were associated with ILD progression at 1-year follow-up: CXCL4 (odds ratio 2.67, 1.46-4.88; P = 0.001), MMP-1 (2.56, 1.43-4.59; P = 0.002) and ET-1 (2.18, 1.24-3.83; P = 0.007). CONCLUSION: Multiple biomarkers, especially VCAM-1, E-selectin, SP-D and CXCL4, provide prognostic utility beyond that of established risk factors for patients with SSc.

Bioorthogonal Radiolabeling of Azide-Modified Bacteria Using [18F]FB-sulfo-DBCO.

Purpose: This study was motivated by the need for better positron emission tomography (PET)-compatible tools to image bacterial infection. Our previous efforts have targeted bacteria-specific metabolism via assimilation of carbon-11 labeled d-amino acids into the bacterial cell wall. Since the chemical determinants of this incorporation are not fully understood, we sought a high-throughput method to label d-amino acid derived structures with fluorine-18. Our strategy employed a chemical biology approach, whereby an azide (-N3) bearing d-amino acid is incorporated into peptidoglycan muropeptides, with subsequent "click" cycloaddition with an 18F-labeled strained cyclooctyne partner. Procedures: A water-soluble, 18F-labeled and dibenzocyclooctyne (DBCO)-derived radiotracer ([18F]FB-sulfo-DBCO) was synthesized. This tracer was incubated with pathogenic bacteria treated with azide-bearing d-amino acids, and incorporated 18F was determined via gamma counting. In vitro uptake in bacteria previously treated with azide-modified d-amino acids was compared to that in cultures treated with amino acid controls. The biodistribution of [18F]FB-sulfo-DBCO was studied in a cohort of healthy mice with implications for future in vivo imaging. Results: The new strain-promoted azide-alkyne cycloaddition (SPAAC) radiotracer [18F]FB-sulfo-DBCO was synthesized with high radiochemical yield and purity via N-succinimidyl 4-[18F]fluorobenzoate ([18F]SFB). Accumulation of [18F]FB-sulfo-DBCO was significantly higher in several bacteria treated with azide-modified d-amino acids than in controls; for example, we observed 7 times greater [18F]FB-sulfo-DBCO ligation in Staphylococcus aureus cultures incubated with 3-azido-d-alanine versus those incubated with d-alanine. Conclusions: The SPAAC radiotracer [18F]FB-sulfo-DBCO was validated in vitro via metabolic labeling of azide-bearing peptidoglycan muropeptides. d-Amino acid-derived PET radiotracers may be more efficiently screened via [18F]FB-sulfo-DBCO modification.

Evaluation of the Catheter Clamp over Hydrophilic Guide Wire Central Venous Catheter Exchange Technique for Air Embolism Prophylaxis in an In Vitro Model.

PURPOSE: To develop a reproducible in vitro model simulating central venous catheter (CVC) exchange with high potential for air embolization and test the hypothesis that a closed catheter clamp over hydrophilic guide wire exchange technique will significantly reduce the volume of air introduced during CVC exchange. MATERIALS AND METHODS: The model consisted of a 16-F valved sheath, 240-mL container, and pressure transducer submerged in water in a 1,200-mL suction canister system. Continuous wall suction was applied to the canister to maintain negative pressure at -7 mm Hg or -11 mm Hg. Each trial consisted of 0.035-inch hydrophilic guide wire introduction, over-the-wire catheter exchange, and wire removal following clinical protocol. A total of 256 trials were performed, 128 trials at each pressure with the catheter clamp open (n = 64) or closed (n = 64) around the hydrophilic guide wire. RESULTS: There was a statistically significant lower volume of air introduced with closed clamp over-the-wire exchanges than with open clamp exchanges at both pressures (2-tailed t-test, P < .001). At -7 mm Hg, a mean of 48.0 mL (SD ± 9.3) of air was introduced with open clamp and 20.6 mL (SD ± 4.7) of air was introduced with closed clamp. At -11 mm Hg, 97.8 mL (SD ± 11.9) of air was introduced with open clamp and 37.8 mL (SD ± 6.3) of air was introduced with closed clamp. CONCLUSIONS: This study demonstrated the use of a reproducible in vitro model mimicking conditions causing air embolism during CVC exchange. Results showed that CVC exchange using closed catheter clamp over hydrophilic guide wire exchange technique significantly reduced the volume of air introduced per exchange.

18F-FDG PET/CT and radiolabeled leukocyte SPECT/CT imaging for the evaluation of cardiovascular infection in the multimodality context: ASNC Imaging Indications (ASNC I2) Series Expert Consensus Recommendations from ASNC, AATS, ACC, AHA, ASE, EANM, HRS, IDSA, SCCT, SNMMI, and STS.

This document on cardiovascular infection, including infective endocarditis, is the first in the American Society of Nuclear Cardiology Imaging Indications (ASNC I2) series to assess the role of radionuclide imaging in the multimodality context for the evaluation of complex systemic diseases with multi-societal involvement including pertinent disciplines. A rigorous modified Delphi approach was used to determine consensus clinical indications, diagnostic criteria, and an algorithmic approach to diagnosis of cardiovascular infection including infective endocarditis. Cardiovascular infection incidence is increasing and is associated with high morbidity and mortality. Current strategies based on clinical criteria and an initial echocardiographic imaging approach are effective but often insufficient in complicated cardiovascular infection. Radionuclide imaging with 18F-fluorodeoxyglucose (FDG) positron emission tomography/computed tomography (CT) and single photon emission computed tomography/CT leukocyte scintigraphy can enhance the evaluation of suspected cardiovascular infection by increasing diagnostic accuracy, identifying extracardiac involvement, and assessing cardiac implanted device pockets, leads, and all portions of ventricular assist devices. This advanced imaging can aid in key medical and surgical considerations. Consensus diagnostic features include focal/multi-focal or diffuse heterogenous intense 18F-FDG uptake on valvular and prosthetic material, perivalvular areas, device pockets and leads, and ventricular assist device hardware persisting on non-attenuation corrected images. There are numerous clinical indications with a larger role in prosthetic valves, and cardiac devices particularly with possible infective endocarditis or in the setting of prior equivocal or non-diagnostic imaging. Illustrative cases incorporating these consensus recommendations provide additional clarification. Future research is necessary to refine application of these advanced imaging tools for surgical planning, to identify treatment response, and more.

Inter- and intraspecific variability of total mercury concentrations in bats of Texas (USA).

Exposure to mercury (Hg) may cause deleterious health effects in wildlife, including bats. Texas produces more Hg pollution than any other state in the United States, yet only one study has examined Hg accumulation in bats. This study measured the concentration of total Hg (THg) in fur (n = 411) collected from ten bat species across 32 sites in eastern and central Texas, USA. Fur THg concentrations were compared among species, and when samples sizes were large enough, between sex and life stage within a species, and the proximity to coal-fired power plants. For all sites combined and species with a sample size ≥8, mean THg concentrations (µg/g dry weight) were greatest in tri-colored bats (Perimyotis subflavus; 6.04), followed by evening bats (Nycticeius humeralis; 5.89), cave myotis (Myotis velifer; 2.11), northern yellow bats (Lasiurus intermedius; 1.85), Brazilian free-tailed bats (Tadarida brasiliensis; 1.03), and red bats (Lasiurus borealis/blossevillii; 0.974), and lowest in hoary bats (Lasiurus cinereus; 0.809). Within a species, fur THg concentrations did not significantly vary between sex for the five examined species (red bat, northern yellow bat, cave myotis, evening bat, Brazilian free-tailed bat) and only between life stage in evening bats. Site variations in fur THg concentrations were observed for evening bats, tri-colored bats, and Brazilian free-tailed bats. Evening bats sampled closer to point sources of Hg pollution had greater fur THg concentrations than individuals sampled further away. Sixteen percent of evening bats and 8.7% of tri-colored bats had a fur THg concentration exceeding the 10 µg/g toxicity threshold level, suggesting that THg exposure may pose a risk to the health of bats in Texas, particularly those residing in east Texas and on the upper Gulf coast. The results of this study can be incorporated into future management and recovery plans for bats in Texas.

Evaluating the Performance of Pathogen-Targeted Positron Emission Tomography Radiotracers in a Rat Model of Vertebral Discitis-Osteomyelitis.

BACKGROUND: Vertebral discitis-osteomyelitis (VDO) is a devastating infection of the spine that is challenging to distinguish from noninfectious mimics using computed tomography and magnetic resonance imaging. We and others have developed novel metabolism-targeted positron emission tomography (PET) radiotracers for detecting living Staphylococcus aureus and other bacteria in vivo, but their head-to-head performance in a well-validated VDO animal model has not been reported. METHODS: We compared the performance of several PET radiotracers in a rat model of VDO. [11C]PABA and [18F]FDS were assessed for their ability to distinguish S aureus, the most common non-tuberculous pathogen VDO, from Escherichia coli. RESULTS: In the rat S aureus VDO model, [11C]PABA could detect as few as 103 bacteria and exhibited the highest signal-to-background ratio, with a 20-fold increased signal in VDO compared to uninfected tissues. In a proof-of-concept experiment, detection of bacterial infection and discrimination between S aureus and E coli was possible using a combination of [11C]PABA and [18F]FDS. CONCLUSIONS: Our work reveals that several bacteria-targeted PET radiotracers had sufficient signal to background in a rat model of S aureus VDO to be potentially clinically useful. [11C]PABA was the most promising tracer investigated and warrants further investigation in human VDO.

The critical impact of sex on preclinical alcohol research - Insights from zebrafish.

Sex is an important biological variable that is widely recognized in studies of alcohol-related effects. Complementing clinical and preclinical rodent research, the zebrafish (Danio rerio) is the second most used laboratory species, and a powerful model organism in biomedicine. Like clinical and rodent models, zebrafish demonstrate overt sex differences in alcohol-related responses. Collectively, this evidence shows that the zebrafish becomes a sensitive model species to further probe in-depth sex differences commonly reported in alcohol research.

Creation of an Ex Vivo Renal Perfusion Model to Investigate Microwave Ablation.

This study hypothesized that an ex vivo renal perfusion model can create smaller microwave ablation (MWA) measurements during perfused states compared with nonperfused states across multiple device settings. Nine bovine kidneys, a fluoroscopic compatible perfusion model, and a commercially-available clinical MWA system were used to perform 72 ablations (36 perfused and 36 nonperfused) at 9 different device settings. Comparing perfused and nonperfused ablations at each device setting, significant differences in volume existed for 6 of 9 settings (P < .05). Collapsed across time settings, the ablation volumes by power were the following (perfused and nonperfused, P value): 60 W, 2.3 cm3 ± 1.0 and 7.2 cm3 ± 2.7, P < .001; 100 W, 5.4 cm3 ± 2.1 and 11.5 cm3 ± 5.6, P < .01; and 140 W, 11.2 cm3 ± 3.7 and 18.7 cm3 ± 6.3, P < .01. Applied power correlated with ablation volume: perfused, 0.021 cm3/W and R = 0.462, P = .004, and nonperfused, 0.029 cm3/W and R = 0.565, P < .001. These results support that an ex vivo perfused organ system can evaluate MWA systems and demonstrate heat sink perfusion effects of decreased ablation size.

Observer studies of image quality of denoising reduced-count cardiac single photon emission computed tomography myocardial perfusion imaging by three-dimensional Gaussian post-reconstruction filtering and deep learning.

BACKGROUND: The aim of this research was to asses perfusion-defect detection-accuracy by human observers as a function of reduced-counts for 3D Gaussian post-reconstruction filtering vs deep learning (DL) denoising to determine if there was improved performance with DL. METHODS: SPECT projection data of 156 normally interpreted patients were used for these studies. Half were altered to include hybrid perfusion defects with defect presence and location known. Ordered-subset expectation-maximization (OSEM) reconstruction was employed with the optional correction of attenuation (AC) and scatter (SC) in addition to distance-dependent resolution (RC). Count levels varied from full-counts (100%) to 6.25% of full-counts. The denoising strategies were previously optimized for defect detection using total perfusion deficit (TPD). Four medical physicist (PhD) and six physician (MD) observers rated the slices using a graphical user interface. Observer ratings were analyzed using the LABMRMC multi-reader, multi-case receiver-operating-characteristic (ROC) software to calculate and compare statistically the area-under-the-ROC-curves (AUCs). RESULTS: For the same count-level no statistically significant increase in AUCs for DL over Gaussian denoising was determined when counts were reduced to either the 25% or 12.5% of full-counts. The average AUC for full-count OSEM with solely RC and Gaussian filtering was lower than for the strategies with AC and SC, except for a reduction to 6.25% of full-counts, thus verifying the utility of employing AC and SC with RC. CONCLUSION: We did not find any indication that at the dose levels investigated and with the DL network employed, that DL denoising was superior in AUC to optimized 3D post-reconstruction Gaussian filtering.

Structural mechanism of laforin function in glycogen dephosphorylation and lafora disease.

Glycogen is the major mammalian glucose storage cache and is critical for energy homeostasis. Glycogen synthesis in neurons must be tightly controlled due to neuronal sensitivity to perturbations in glycogen metabolism. Lafora disease (LD) is a fatal, congenital, neurodegenerative epilepsy. Mutations in the gene encoding the glycogen phosphatase laforin result in hyperphosphorylated glycogen that forms water-insoluble inclusions called Lafora bodies (LBs). LBs induce neuronal apoptosis and are the causative agent of LD. The mechanism of glycogen dephosphorylation by laforin and dysfunction in LD is unknown. We report the crystal structure of laforin bound to phosphoglucan product, revealing its unique integrated tertiary and quaternary structure. Structure-guided mutagenesis combined with biophysical and biochemical analyses reveal the basis for normal function of laforin in glycogen metabolism. Analyses of LD patient mutations define the mechanism by which subsets of mutations disrupt laforin function. These data provide fundamental insights connecting glycogen metabolism to neurodegenerative disease.

Perspectives on Secondary Mitral Regurgitation in Heart Failure.

PURPOSE OF THE REVIEW: This review focuses on broader perspectives of mitral regurgitation (MR) in patients with heart failure. RECENT FINDINGS: The ratio of regurgitant volume to end-diastolic volume appears to help identify patients who may benefit from valve interventions. Secondary MR is not only attributed to geometric changes of the LV but also related to the structural changes in the mitral valve that include fibrosis of the mitral leaflets and changes in the extracellular matrix. The transition from mild to severe secondary MR can occur at different rates, from a slow LV remodeling process to a more abrupt process precipitated by an inciting event such as atrial fibrillation. Septal flash and apical rocking, two new visual markers of LV mechanical dyssynchrony, appear to be predictive of MR reduction following cardiac resynchronization therapy. Optimal guideline-directed medical therapy has been shown to decrease the severity of secondary MR effectively. A theoretical framework to characterize secondary MR as it relates to the onset of MR is proposed. Type A: Early onset of MR contemporaneous with myocardial injury. The maladaptive LV remodeling occurs in parallel with MR. Type B: LV remodeling proceeds without significant MR until the LV is moderately dilated, which coincides with or without inciting factors such as atrial fibrillation. Type C: LV remodeling proceeds after myocardial injury without significant MR until the LV is severely dilated. MR is a late manifestation of LV remodeling.

Nostalgia enhances route learning in a virtual environment.

Salient landmarks enhance route learning. We hypothesised that semantically salient nostalgic landmarks would improve route learning compared to non-nostalgic landmarks. In two experiments, participants learned a route through a computer-generated maze using directional arrows and wall-mounted pictures. On the test trial, the arrows were removed, and participants completed the maze using only the pictures. In the nostalgia condition, pictures were of popular music artists and TV characters from 5 to 10 years ago. In the control condition, they were recent pictures of these same artists and characters. In Experiment 1, in the test trial, participants in the nostalgia condition completed the maze faster than controls. Experiment 2 conceptually replicated these findings and extended them by exploring boundary conditions. Participants had to learn two mazes sequentially. In Maze 1, we placed nostalgic/control landmarks only at non-decision points (whereas we placed them at decision points in Experiment 1). In Maze 2, we placed nostalgic/control landmarks at decision points during acquisition but removed them in the test trial (whereas they were present in the test trial in Experiment 1). In both mazes, participants in the nostalgia (compared to control) condition completed the test trial faster.

Induction of spatial anxiety in a virtual navigation environment.

Spatial anxiety (i.e., feelings of apprehension and fear about navigating everyday environments) can adversely impact people's ability to reach desired locations and explore unfamiliar places. Prior research has either assessed spatial anxiety as an individual-difference variable or measured it as an outcome, but there are currently no experimental inductions to investigate its causal effects. To address this lacuna, we developed a novel protocol for inducing spatial anxiety within a virtual environment. Participants first learnt a route using directional arrows. Next, we removed the directional arrows and randomly assigned participants to navigate either the same route (n = 22; control condition) or a variation of this route in which we surreptitiously introduced unfamiliar paths and landmarks (n = 22; spatial-anxiety condition). The manipulation successfully induced transient (i.e., state-level) spatial anxiety and task stress but did not significantly reduce task enjoyment. Our findings lay the foundation for an experimental paradigm that will facilitate future work on the causal effects of spatial anxiety in navigational contexts. The experimental task is freely available via the Open Science Framework ( https://osf.io/uq4v7/ ).