The simplicity of protein sequence-function relationships.

How complex are the rules by which a protein's sequence determines its function? High-order epistatic interactions among residues are thought to be pervasive, suggesting an idiosyncratic and unpredictable sequence-function relationship. But many prior studies may have overestimated epistasis, because they analyzed sequence-function relationships relative to a single reference sequence-which causes measurement noise and local idiosyncrasies to snowball into high-order epistasis-or they did not fully account for global nonlinearities. Here we present a reference-free method that jointly infers specific epistatic interactions and global nonlinearity using a bird's-eye view of sequence space. This technique yields the simplest explanation of sequence-function relationships and is more robust than existing methods to measurement noise, missing data, and model misspecification. We reanalyze 20 experimental datasets and find that context-independent amino acid effects and pairwise interactions, along with a simple nonlinearity to account for limited dynamic range, explain a median of 96% of phenotypic variance and over 92% in every case. Only a tiny fraction of genotypes are strongly affected by higher-order epistasis. Sequence-function relationships are also sparse: a miniscule fraction of amino acids and interactions account for 90% of phenotypic variance. Sequence-function causality across these datasets is therefore simple, opening the way for tractable approaches to characterize proteins' genetic architecture.

Psychometric Properties of a Novel Affective Bias Task and its Application in Clinical and Non-Clinical Populations.

To mitigate limitations in self-reported mood assessments, we introduce a novel affective bias task (ABT). The task quantifies instantaneous emotional state by leveraging the phenomenon of affective bias, in which people interpret external emotional stimuli in a manner consistent with their current emotional state. This study establishes task stability in measuring and tracking depressive symptoms in clinical and non-clinical populations. Initial assessment in a large non-clinical sample established normative ratings. Depressive symptoms were tracked relative to task performance in a non-clinical sample, as well as in a clinical cohort undergoing surgical evaluation for severe epilepsy. In both cohorts, a stronger negative affective bias was associated with higher Beck Depression Inventory (BDI-II) scores. The ABT exhibits high stability and interrater reliability, as well as construct validity in predicting depression levels in both cohorts, suggesting the task as a reliable proxy for mood and a diagnostic tool for detecting depressive symptoms.

JWST detection of a supernova associated with GRB 221009A without an r-process signature.

Identifying the sites of r-process nucleosynthesis, a primary mechanism of heavy element production, is a key goal of astrophysics. The discovery of the brightest gamma-ray burst (GRB) to date, GRB 221009A, presented an opportunity to spectroscopically test the idea that r-process elements are produced following the collapse of rapidly rotating massive stars. Here we present James Webb Space Telescope observations of GRB 221009A obtained +168 and +170 rest-frame days after the gamma-ray trigger, and demonstrate that they are well described by a SN 1998bw-like supernova (SN) and power-law afterglow, with no evidence for a component from r-process emission. The SN, with a nickel mass of approximately 0.09 M ⊙, is only slightly fainter than the brightness of SN 1998bw at this phase, which indicates that the SN is not an unusual GRB-SN. This demonstrates that the GRB and SN mechanisms are decoupled and that highly energetic GRBs are not likely to produce significant quantities of r-process material, which leaves open the question of whether explosions of massive stars are key sources of r-process elements. Moreover, the host galaxy of GRB 221009A has a very low metallicity of approximately 0.12 Z ⊙ and strong H2 emission at the explosion site, which is consistent with recent star formation, hinting that environmental factors are responsible for its extreme energetics.

Genetic code robustness and protein evolvability are correlated and protein-specific.

The relationship between genetic code robustness and protein evolvability is unknown. A new study in PLOS Biology using in silico rewiring of genetic codes and functional protein data identified a positive correlation between code robustness and protein evolvability that is protein-specific.

Epistasis facilitates functional evolution in an ancient transcription factor.

A protein's genetic architecture - the set of causal rules by which its sequence produces its functions - also determines its possible evolutionary trajectories. Prior research has proposed that the genetic architecture of proteins is very complex, with pervasive epistatic interactions that constrain evolution and make function difficult to predict from sequence. Most of this work has analyzed only the direct paths between two proteins of interest - excluding the vast majority of possible genotypes and evolutionary trajectories - and has considered only a single protein function, leaving unaddressed the genetic architecture of functional specificity and its impact on the evolution of new functions. Here, we develop a new method based on ordinal logistic regression to directly characterize the global genetic determinants of multiple protein functions from 20-state combinatorial deep mutational scanning (DMS) experiments. We use it to dissect the genetic architecture and evolution of a transcription factor's specificity for DNA, using data from a combinatorial DMS of an ancient steroid hormone receptor's capacity to activate transcription from two biologically relevant DNA elements. We show that the genetic architecture of DNA recognition consists of a dense set of main and pairwise effects that involve virtually every possible amino acid state in the protein-DNA interface, but higher-order epistasis plays only a tiny role. Pairwise interactions enlarge the set of functional sequences and are the primary determinants of specificity for different DNA elements. They also massively expand the number of opportunities for single-residue mutations to switch specificity from one DNA target to another. By bringing variants with different functions close together in sequence space, pairwise epistasis therefore facilitates rather than constrains the evolution of new functions.

Heavy-element production in a compact object merger observed by JWST.

The mergers of binary compact objects such as neutron stars and black holes are of central interest to several areas of astrophysics, including as the progenitors of gamma-ray bursts (GRBs)1, sources of high-frequency gravitational waves (GWs)2 and likely production sites for heavy-element nucleosynthesis by means of rapid neutron capture (the r-process)3. Here we present observations of the exceptionally bright GRB 230307A. We show that GRB 230307A belongs to the class of long-duration GRBs associated with compact object mergers4-6 and contains a kilonova similar to AT2017gfo, associated with the GW merger GW170817 (refs. 7-12). We obtained James Webb Space Telescope (JWST) mid-infrared imaging and spectroscopy 29 and 61 days after the burst. The spectroscopy shows an emission line at 2.15 microns, which we interpret as tellurium (atomic mass A = 130) and a very red source, emitting most of its light in the mid-infrared owing to the production of lanthanides. These observations demonstrate that nucleosynthesis in GRBs can create r-process elements across a broad atomic mass range and play a central role in heavy-element nucleosynthesis across the Universe.

The simplicity of protein sequence-function relationships.

How complicated is the genetic architecture of proteins - the set of causal effects by which sequence determines function? High-order epistatic interactions among residues are thought to be pervasive, making a protein's function difficult to predict or understand from its sequence. Most studies, however, used methods that overestimate epistasis, because they analyze genetic architecture relative to a designated reference sequence - causing measurement noise and small local idiosyncrasies to propagate into pervasive high-order interactions - or have not effectively accounted for global nonlinearity in the sequence-function relationship. Here we present a new reference-free method that jointly estimates global nonlinearity and specific epistatic interactions across a protein's entire genotype-phenotype map. This method yields a maximally efficient explanation of a protein's genetic architecture and is more robust than existing methods to measurement noise, partial sampling, and model misspecification. We reanalyze 20 combinatorial mutagenesis experiments from a diverse set of proteins and find that additive and pairwise effects, along with a simple nonlinearity to account for limited dynamic range, explain a median of 96% of total variance in measured phenotypes (and >92% in every case). Only a tiny fraction of genotypes are strongly affected by third- or higher-order epistasis. Genetic architecture is also sparse: the number of terms required to explain the vast majority of variance is smaller than the number of genotypes by many orders of magnitude. The sequence-function relationship in most proteins is therefore far simpler than previously thought, opening the way for new and tractable approaches to characterize it.

Brain mechanisms underlying the emotion processing bias in treatment-resistant depression.

Depression is associated with a cognitive bias towards negative information and away from positive information. This biased emotion processing may underlie core depression symptoms, including persistent feelings of sadness or low mood and a reduced capacity to experience pleasure. The neural mechanisms responsible for this biased emotion processing remain unknown. Here, we had a unique opportunity to record stereotactic electroencephalography (sEEG) signals in the amygdala and prefrontal cortex (PFC) from 5 treatment-resistant depression (TRD) patients and 12 epilepsy patients (as control) while they participated in an affective bias task in which happy and sad faces were rated. First, compared with the control group, patients with TRD showed increased amygdala responses to sad faces in the early stage (around 300 ms) and decreased amygdala responses to happy faces in the late stage (around 600 ms) following the onset of faces. Further, during the late stage of happy face processing, alpha-band activity in PFC as well as alpha-phase locking between the amygdala and PFC were significantly greater in TRD patients compared to the controls. Second, after deep brain stimulation (DBS) delivered to bilateral subcallosal cingulate (SCC) and ventral capsule/ventral striatum (VC/VS), atypical amygdala and PFC processing of happy faces in TRD patients remitted toward the normative pattern. The increased amygdala activation during the early stage of sad face processing suggests an overactive bottom-up processing system in TRD. Meanwhile, the reduced amygdala response during the late stage of happy face processing could be attributed to inhibition by PFC through alpha-band oscillation, which can be released by DBS in SCC and VC/VS.

Intracranial stimulation and EEG feature analysis reveal affective salience network specialization.

Emotion is represented in limbic and prefrontal brain areas, herein termed the affective salience network (ASN). Within the ASN, there are substantial unknowns about how valence and emotional intensity are processed-specifically, which nodes are associated with affective bias (a phenomenon in which participants interpret emotions in a manner consistent with their own mood). A recently developed feature detection approach ('specparam') was used to select dominant spectral features from human intracranial electrophysiological data, revealing affective specialization within specific nodes of the ASN. Spectral analysis of dominant features at the channel level suggests that dorsal anterior cingulate (dACC), anterior insula and ventral-medial prefrontal cortex (vmPFC) are sensitive to valence and intensity, while the amygdala is primarily sensitive to intensity. Akaike information criterion model comparisons corroborated the spectral analysis findings, suggesting all four nodes are more sensitive to intensity compared to valence. The data also revealed that activity in dACC and vmPFC were predictive of the extent of affective bias in the ratings of facial expressions-a proxy measure of instantaneous mood. To examine causality of the dACC in affective experience, 130 Hz continuous stimulation was applied to dACC while patients viewed and rated emotional faces. Faces were rated significantly happier during stimulation, even after accounting for differences in baseline ratings. Together the data suggest a causal role for dACC during the processing of external affective stimuli.

Decoding Depression Severity From Intracranial Neural Activity.

BACKGROUND: Disorders of mood and cognition are prevalent, disabling, and notoriously difficult to treat. Fueling this challenge in treatment is a significant gap in our understanding of their neurophysiological basis. METHODS: We recorded high-density neural activity from intracranial electrodes implanted in depression-relevant prefrontal cortical regions in 3 human subjects with severe depression. Neural recordings were labeled with depression severity scores across a wide dynamic range using an adaptive assessment that allowed sampling with a temporal frequency greater than that possible with typical rating scales. We modeled these data using regularized regression techniques with region selection to decode depression severity from the prefrontal recordings. RESULTS: Across prefrontal regions, we found that reduced depression severity is associated with decreased low-frequency neural activity and increased high-frequency activity. When constraining our model to decode using a single region, spectral changes in the anterior cingulate cortex best predicted depression severity in all 3 subjects. Relaxing this constraint revealed unique, individual-specific sets of spatiospectral features predictive of symptom severity, reflecting the heterogeneous nature of depression. CONCLUSIONS: The ability to decode depression severity from neural activity increases our fundamental understanding of how depression manifests in the human brain and provides a target neural signature for personalized neuromodulation therapies.

A kilonova following a long-duration gamma-ray burst at 350 Mpc.

Gamma-ray bursts (GRBs) are divided into two populations1,2; long GRBs that derive from the core collapse of massive stars (for example, ref. 3) and short GRBs that form in the merger of two compact objects4,5. Although it is common to divide the two populations at a gamma-ray duration of 2 s, classification based on duration does not always map to the progenitor. Notably, GRBs with short (≲2 s) spikes of prompt gamma-ray emission followed by prolonged, spectrally softer extended emission (EE-SGRBs) have been suggested to arise from compact object mergers6-8. Compact object mergers are of great astrophysical importance as the only confirmed site of rapid neutron capture (r-process) nucleosynthesis, observed in the form of so-called kilonovae9-14. Here we report the discovery of a possible kilonova associated with the nearby (350 Mpc), minute-duration GRB 211211A. The kilonova implies that the progenitor is a compact object merger, suggesting that GRBs with long, complex light curves can be spawned from merger events. The kilonova of GRB 211211A has a similar luminosity, duration and colour to that which accompanied the gravitational wave (GW)-detected binary neutron star (BNS) merger GW170817 (ref. 4). Further searches for GW signals coincident with long GRBs are a promising route for future multi-messenger astronomy.

Spontaneous neuronal oscillations in the human insula are hierarchically organized traveling waves.

The insula plays a fundamental role in a wide range of adaptive human behaviors, but its electrophysiological dynamics are poorly understood. Here, we used human intracranial electroencephalographic recordings to investigate the electrophysiological properties and hierarchical organization of spontaneous neuronal oscillations within the insula. We analyzed the neuronal oscillations of the insula directly and found that rhythms in the theta and beta frequency oscillations are widespread and spontaneously present. These oscillations are largely organized along the anterior-posterior (AP) axis of the insula. Both the left and right insula showed anterior--to-posterior decreasing gradients for the power of oscillations in the beta frequency band. The left insula also showed a posterior-to-anterior decreasing frequency gradient and an anterior-to-posterior decreasing power gradient in the theta frequency band. In addition to measuring the power of these oscillations, we also examined the phase of these signals across simultaneous recording channels and found that the insula oscillations in the theta and beta bands are traveling waves. The strength of the traveling waves in each frequency was positively correlated with the amplitude of each oscillation. However, the theta and beta traveling waves were uncoupled to each other in terms of phase and amplitude, which suggested that insular traveling waves in the theta and beta bands operate independently. Our findings provide new insights into the spatiotemporal dynamics and hierarchical organization of neuronal oscillations within the insula, which, given its rich connectivity with widespread cortical regions, indicates that oscillations and traveling waves have an important role in intrainsular and interinsular communications.

Epistatic drift causes gradual decay of predictability in protein evolution.

Epistatic interactions can make the outcomes of evolution unpredictable, but no comprehensive data are available on the extent and temporal dynamics of changes in the effects of mutations as protein sequences evolve. Here, we use phylogenetic deep mutational scanning to measure the functional effect of every possible amino acid mutation in a series of ancestral and extant steroid receptor DNA binding domains. Across 700 million years of evolution, epistatic interactions caused the effects of most mutations to become decorrelated from their initial effects and their windows of evolutionary accessibility to open and close transiently. Most effects changed gradually and without bias at rates that were largely constant across time, indicating a neutral process caused by many weak epistatic interactions. Our findings show that protein sequences drift inexorably into contingency and unpredictability, but that the process is statistically predictable, given sufficient phylogenetic and experimental data.

Imaging versus electrographic connectivity in human mood-related fronto-temporal networks.

BACKGROUND: The efficacy of psychiatric DBS is thought to be driven by the connectivity of stimulation targets with mood-relevant fronto-temporal networks, which is typically evaluated using diffusion-weighted tractography. OBJECTIVE: Leverage intracranial electrophysiology recordings to better predict the circuit-wide effects of neuromodulation to white matter targets. We hypothesize strong convergence between tractography-predicted structural connectivity and stimulation-induced electrophysiological responses. METHODS: Evoked potentials were elicited by single-pulse stimulation to two common DBS targets for treatment-resistant depression - the subcallosal cingulate (SCC) and ventral capsule/ventral striatum (VCVS) - in two patients undergoing DBS with stereo-electroencephalographic (sEEG) monitoring. Evoked potentials were compared with predicted structural connectivity between DBS leads and sEEG contacts using probabilistic, patient-specific diffusion-weighted tractography. RESULTS: Evoked potentials and tractography showed strong convergence in both patients in orbitofrontal, ventromedial prefrontal, and lateral prefrontal cortices for both SCC and VCVS stimulation targets. Low convergence was found in anterior cingulate (ACC), where tractography predicted structural connectivity from SCC targets but produced no evoked potentials during SCC stimulation. Further, tractography predicted no connectivity to ACC from VCVS targets, but VCVS stimulation produced robust evoked potentials. CONCLUSION: The two connectivity methods showed significant convergence, but important differences emerged with respect to the ability of tractography to predict electrophysiological connectivity between SCC and VCVS to regions of the mood-related network. This multimodal approach raises intriguing implications for the use of tractography in surgical targeting and provides new data to enhance our understanding of the network-wide effects of neuromodulation.

Deep Brain Stimulation for Depression Informed by Intracranial Recordings.

The success of deep brain stimulation (DBS) for treating Parkinson's disease has led to its application to several other disorders, including treatment-resistant depression. Results with DBS for treatment-resistant depression have been heterogeneous, with inconsistencies largely driven by incomplete understanding of the brain networks regulating mood, especially on an individual basis. We report results from the first subject treated with DBS for treatment-resistant depression using an approach that incorporates intracranial recordings to personalize understanding of network behavior and its response to stimulation. These recordings enabled calculation of individually optimized DBS stimulation parameters using a novel inverse solution approach. In the ensuing double-blind, randomized phase incorporating these bespoke parameter sets, DBS led to remission of symptoms and dramatic improvement in quality of life. Results from this initial case demonstrate the feasibility of this personalized platform, which may be used to improve surgical neuromodulation for a vast array of neurologic and psychiatric disorders.

Mutational sources of trans-regulatory variation affecting gene expression in Saccharomyces cerevisiae.

Heritable variation in a gene's expression arises from mutations impacting cis- and trans-acting components of its regulatory network. Here, we investigate how trans-regulatory mutations are distributed within the genome and within a gene regulatory network by identifying and characterizing 69 mutations with trans-regulatory effects on expression of the same focal gene in Saccharomyces cerevisiae. Relative to 1766 mutations without effects on expression of this focal gene, we found that these trans-regulatory mutations were enriched in coding sequences of transcription factors previously predicted to regulate expression of the focal gene. However, over 90% of the trans-regulatory mutations identified mapped to other types of genes involved in diverse biological processes including chromatin state, metabolism, and signal transduction. These data show how genetic changes in diverse types of genes can impact a gene's expression in trans, revealing properties of trans-regulatory mutations that provide the raw material for trans-regulatory variation segregating within natural populations.

Contingency and chance erase necessity in the experimental evolution of ancestral proteins.

The roles of chance, contingency, and necessity in evolution are unresolved because they have never been assessed in a single system or on timescales relevant to historical evolution. We combined ancestral protein reconstruction and a new continuous evolution technology to mutate and select proteins in the B-cell lymphoma-2 (BCL-2) family to acquire protein-protein interaction specificities that occurred during animal evolution. By replicating evolutionary trajectories from multiple ancestral proteins, we found that contingency generated over long historical timescales steadily erased necessity and overwhelmed chance as the primary cause of acquired sequence variation; trajectories launched from phylogenetically distant proteins yielded virtually no common mutations, even under strong and identical selection pressures. Chance arose because many sets of mutations could alter specificity at any timepoint; contingency arose because historical substitutions changed these sets. Our results suggest that patterns of variation in BCL-2 sequences - and likely other proteins, too - are idiosyncratic products of a particular and unpredictable course of historical events.

One of the most fundamental and unresolved questions in evolutionary biology is whether the outcomes of evolution are predictable. Is the diversity of life we see today the expected result of organisms adapting to their environment throughout history (also known as natural selection) or the product of random chance? Or did chance events early in history shape the paths that evolution could take next, determining the biological forms that emerged under natural selection much later? These questions are hard to study because evolution happened only once, long ago. To overcome this barrier, Xie, Pu, Metzger et al. developed an experimental approach that can evolve reconstructed ancestral proteins that existed deep in the past. Using this method, it is possible to replay evolution multiple times, from various historical starting points, under conditions similar to those that existed long ago. The end products of the evolutionary trajectories can then be compared to determine how predictable evolution actually is. Xie, Pu, Metzger et al. studied proteins belonging to the BCL-2 family, which originated some 800 million years ago. These proteins have diversified greatly over time in both their genetic sequences and their ability to bind to specific partner proteins called co-regulators. Xie, Pu, Metzger et al. synthesized BCL-2 proteins that existed at various times in the past. Each ancestral protein was then allowed to evolve repeatedly under natural selection to acquire the same co-regulator binding functions that evolved during history. At the end of each evolutionary trajectory, the genetic sequence of the resulting BCL-2 proteins was recorded. This revealed that the outcomes of evolution were almost completely unpredictable: trajectories initiated from the same ancestral protein produced proteins with very different sequences, and proteins launched from different ancestral starting points were even more dissimilar. Further experiments identified the mutations in each trajectory that caused changes in coregulator binding. When these mutations were introduced into other ancestral proteins, they did not yield the same change in function. This suggests that early chance events influenced each protein's evolution in an unpredictable way by opening and closing the paths available to it in the future. This research expands our understanding of evolution on a molecular level whilst providing a new experimental approach for studying evolutionary drivers in more detail. The results suggest that BCL-2 proteins, in all their various forms, are unique products of a particular, unpredictable course of history set in motion by ancient chance events.

A hydrophobic ratchet entrenches molecular complexes.

Most proteins assemble into multisubunit complexes1. The persistence of these complexes across evolutionary time is usually explained as the result of natural selection for functional properties that depend on multimerization, such as intersubunit allostery or the capacity to do mechanical work2. In many complexes, however, multimerization does not enable any known function3. An alternative explanation is that multimers could become entrenched if substitutions accumulate that are neutral in multimers but deleterious in monomers; purifying selection would then prevent reversion to the unassembled form, even if assembly per se does not enhance biological function3-7. Here we show that a hydrophobic mutational ratchet systematically entrenches molecular complexes. By applying ancestral protein reconstruction and biochemical assays to the evolution of steroid hormone receptors, we show that an ancient hydrophobic interface, conserved for hundreds of millions of years, is entrenched because exposure of this interface to solvent reduces protein stability and causes aggregation, even though the interface makes no detectable contribution to function. Using structural bioinformatics, we show that a universal mutational propensity drives sites that are buried in multimeric interfaces to accumulate hydrophobic substitutions to levels that are not tolerated in monomers. In a database of hundreds of families of multimers, most show signatures of long-term hydrophobic entrenchment. It is therefore likely that many protein complexes persist because a simple ratchet-like mechanism entrenches them across evolutionary time, even when they are functionally gratuitous.

A blue ring nebula from a stellar merger several thousand years ago.

Stellar mergers are a brief but common phase in the evolution of binary star systems1,2. These events have many astrophysical implications; for example, they may lead to the creation of atypical stars (such as magnetic stars3, blue stragglers4 and rapid rotators5), they play an important part in our interpretation of stellar populations6 and they represent formation channels of compact-object mergers7. Although a handful of stellar mergers have been observed directly8,9, the central remnants of these events were shrouded by an opaque shell of dust and molecules10, making it impossible to observe their final state (for example, as a single merged star or a tighter, surviving binary11). Here we report observations of an unusual, ring-shaped ultraviolet ('blue') nebula and the star at its centre, TYC 2597-735-1. The nebula has two opposing fronts, suggesting a bipolar outflow of material from TYC 2597-735-1. The spectrum of TYC 2597-735-1 and its proximity to the Galactic plane suggest that it is an old star, yet it has abnormally low surface gravity and a detectable long-term luminosity decay, which is uncharacteristic for its evolutionary stage. TYC 2597-735-1 also exhibits Hα emission, radial-velocity variations, enhanced ultraviolet radiation and excess infrared emission-signatures of dusty circumstellar disks12, stellar activity13 and accretion14. Combined with stellar evolution models, the observations suggest that TYC 2597-735-1 merged with a lower-mass companion several thousand years ago. TYC 2597-735-1 provides a look at an unobstructed stellar merger at an evolutionary stage between its dynamic onset and the theorized final equilibrium state, enabling the direct study of the merging process.