Synthetic retinal analogues modify the spectral and kinetic characteristics of microbial rhodopsin optogenetic tools

Optogenetic tools have become indispensable in neuroscience to stimulate or inhibit excitable cells by light. Channelrhodopsin-2 (ChR2) variants have been established by mutating the opsin backbone or by mining related algal genomes. As an alternative strategy, we surveyed synthetic retinal analogues combined with microbial rhodopsins for functional and spectral properties, capitalizing on assays in C. elegans, HEK cells and larval Drosophila. Compared with all-trans retinal (ATR), Dimethylamino-retinal (DMAR) shifts the action spectra maxima of ChR2 variants H134R and H134R/T159C from 480 to 520 nm. Moreover, DMAR decelerates the photocycle of ChR2(H134R) and (H134R/T159C), thereby reducing the light intensity required for persistent channel activation. In hyperpolarizing archaerhodopsin-3 and Mac, naphthyl-retinal and thiophene-retinal support activity alike ATR, yet at altered peak wavelengths. Our experiments enable applications of retinal analogues in colour tuning and altering photocycle characteristics of optogenetic tools, thereby increasing the operational light sensitivity of existing cell lines or transgenic animals

Synthetic retinal analogues modify the spectral and kinetic characteristics of microbial rhodopsin optogenetic tools

Optogenetic tools have become indispensable in neuroscience to stimulate or inhibit excitable cells by light. Channelrhodopsin-2 (ChR2) variants have been established by mutating the opsin backbone or by mining related algal genomes. As an alternative strategy, we surveyed synthetic retinal analogues combined with microbial rhodopsins for functional and spectral properties, capitalizing on assays in C. elegans, HEK cells and larval Drosophila. Compared with all-trans retinal (ATR), Dimethylamino-retinal (DMAR) shifts the action spectra maxima of ChR2 variants H134R and H134R/T159C from 480 to 520 nm. Moreover, DMAR decelerates the photocycle of ChR2(H134R) and (H134R/T159C), thereby reducing the light intensity required for persistent channel activation. In hyperpolarizing archaerhodopsin-3 and Mac, naphthyl-retinal and thiophene-retinal support activity alike ATR, yet at altered peak wavelengths. Our experiments enable applications of retinal analogues in colour tuning and altering photocycle characteristics of optogenetic tools, thereby increasing the operational light sensitivity of existing cell lines or transgenic animals

Integration of datasets for individual prediction of DNA methylation-based biomarkers

BACKGROUND: Epigenetic scores (EpiScores) can provide biomarkers of lifestyle and disease risk. Projecting new datasets onto a reference panel is challenging due to separation of technical and biological variation with array data. Normalisation can standardise data distributions but may also remove population-level biological variation.RESULTS: We compare two birth cohorts (Lothian Birth Cohorts of 1921 and 1936 - nLBC1921 = 387 and nLBC1936 = 498) with blood-based DNA methylation assessed at the same chronological age (79 years) and processed in the same lab but in different years and experimental batches. We examine the effect of 16 normalisation methods on a novel BMI EpiScore (trained in an external cohort, n = 18,413), and Horvath's pan-tissue DNA methylation age, when the cohorts are normalised separately and together. The BMI EpiScore explains a maximum variance of R2=24.5% in BMI in LBC1936 (SWAN normalisation). Although there are cross-cohort R2 differences, the normalisation method makes a minimal difference to within-cohort estimates. Conversely, a range of absolute differences are seen for individual-level EpiScore estimates for BMI and age when cohorts are normalised separately versus together. While within-array methods result in identical EpiScores whether a cohort is normalised on its own or together with the second dataset, a range of differences is observed for between-array methods.CONCLUSIONS: Normalisation methods returning similar EpiScores, whether cohorts are analysed separately or together, will minimise technical variation when projecting new data onto a reference panel. These methods are important for cases where raw data is unavailable and joint normalisation of cohorts is computationally expensive.</p

Alcohol consumption and lifetime change in cognitive ability:a gene × environment interaction study

Studies of the effect of alcohol consumption on cognitive ability are often confounded. One approach to avoid confounding is the Mendelian randomization design. Here, we used such a design to test the hypothesis that a genetic score for alcohol processing capacity moderates the association between alcohol consumption and lifetime change in cognitive ability. Members of the Lothian Birth Cohort 1936 completed the same test of intelligence at age 11 and 70 years. They were assessed for recent alcohol consumption in later life and genotyped for a set of four single-nucleotide polymorphisms in three alcohol dehydrogenase genes. These variants were unrelated to late-life cognition or to socioeconomic status. We found a significant gene × alcohol consumption interaction on lifetime cognitive change (p = 0.007). Individuals with higher genetic ability to process alcohol showed relative improvements in cognitive ability with more consumption, whereas those with low processing capacity showed a negative relationship between cognitive change and alcohol consumption with more consumption. The effect of alcohol consumption on cognitive change may thus depend on genetic differences in the ability to metabolize alcohol

Increased ultra-rare variant load in an isolated Scottish population impacts exonic and regulatory regions

Human population isolates provide a snapshot of the impact of historical demographic processes on population genetics. Such data facilitate studies of the functional impact of rare sequence variants on biomedical phenotypes, as strong genetic drift can result in higher frequencies of variants that are otherwise rare. We present the first whole genome sequencing (WGS) study of the VIKING cohort, a representative collection of samples from the isolated Shetland population in northern Scotland, and explore how its genetic characteristics compare to a mainland Scottish population. Our analyses reveal the strong contributions played by the founder effect and genetic drift in shaping genomic variation in the VIKING cohort. About one tenth of all high-quality variants discovered are unique to the VIKING cohort or are seen at frequencies at least ten fold higher than in more cosmopolitan control populations. Multiple lines of evidence also suggest relaxation of purifying selection during the evolutionary history of the Shetland isolate. We demonstrate enrichment of ultra-rare VIKING variants in exonic regions and for the first time we also show that ultra-rare variants are enriched within regulatory regions, particularly promoters, suggesting that gene expression patterns may diverge relatively rapidly in human isolates

Multi-method genome- and epigenome-wide studies of inflammatory protein levels in healthy older adults

The molecular factors which control circulating levels of inflammatory proteins are not well understood. Furthermore, association studies between molecular probes and human traits are often performed by linear model-based methods which may fail to account for complex structure and interrelationships within molecular datasets.In this study, we perform genome- and epigenome-wide association studies (GWAS/EWAS) on the levels of 70 plasma-derived inflammatory protein biomarkers in healthy older adults (Lothian Birth Cohort 1936; n = 876; Olink® inflammation panel). We employ a Bayesian framework (BayesR+) which can account for issues pertaining to data structure and unknown confounding variables (with sensitivity analyses using ordinary least squares- (OLS) and mixed model-based approaches). We identified 13 SNPs associated with 13 proteins (n = 1 SNP each) concordant across OLS and Bayesian methods. We identified 3 CpG sites spread across 3 proteins (n = 1 CpG each) that were concordant across OLS, mixed-model and Bayesian analyses. Tagged genetic variants accounted for up to 45% of variance in protein levels (for MCP2, 36% of variance alone attributable to 1 polymorphism). Methylation data accounted for up to 46% of variation in protein levels (for CXCL10). Up to 66% of variation in protein levels (for VEGFA) was explained using genetic and epigenetic data combined. We demonstrated putative causal relationships between CD6 and IL18R1 with inflammatory bowel disease and between IL12B and Crohn’s disease. Our data may aid understanding of the molecular regulation of the circulating inflammatory proteome as well as causal relationships between inflammatory mediators and disease

A GABAergic and peptidergic sleep neuron as a locomotion stop neuron with compartmentalized Ca2+dynamics.

Animals must slow or halt locomotion to integrate sensory inputs or to change direction. In Caenorhabditis elegans, the GABAergic and peptidergic neuron RIS mediates developmentally timed quiescence. Here, we show RIS functions additionally as a locomotion stop neuron. RIS optogenetic stimulation caused acute and persistent inhibition of locomotion and pharyngeal pumping, phenotypes requiring FLP-11 neuropeptides and GABA. RIS photoactivation allows the animal to maintain its body posture by sustaining muscle tone, yet inactivating motor neuron oscillatory activity. During locomotion, RIS axonal Ca2+ signals revealed functional compartmentalization: Activity in the nerve ring process correlated with locomotion stop, while activity in a branch correlated with induced reversals. GABA was required to induce, and FLP-11 neuropeptides were required to sustain locomotion stop. RIS attenuates neuronal activity and inhibits movement, possibly enabling sensory integration and decision making, and exemplifies dual use of one cell across development in a compact nervous system

Phase-resolved terahertz self-detection near-field microscopy

At terahertz (THz) frequencies, scattering-type scanning near-field optical microscopy (s-SNOM) based on continuous wave sources mostly relies on cryogenic and bulky detectors, which represents a major constraint for its practical application. Here, we devise a THz s-SNOM system that provides both amplitude and phase contrast and achieves nanoscale (60-70nm) in-plane spatial resolution. It features a quantum cascade laser that simultaneously emits THz frequency light and senses the backscattered optical field through a voltage modulation induced inherently through the self-mixing technique. We demonstrate its performance by probing a phonon-polariton-resonant CsBr crystal and doped black phosphorus flakes