An atlas of protein homo-oligomerization across domains of life.

Protein structures are essential to understanding cellular processes in molecular detail. While advances in artificial intelligence revealed the tertiary structure of proteins at scale, their quaternary structure remains mostly unknown. We devise a scalable strategy based on AlphaFold2 to predict homo-oligomeric assemblies across four proteomes spanning the tree of life. Our results suggest that approximately 45% of an archaeal proteome and a bacterial proteome and 20% of two eukaryotic proteomes form homomers. Our predictions accurately capture protein homo-oligomerization, recapitulate megadalton complexes, and unveil hundreds of homo-oligomer types, including three confirmed experimentally by structure determination. Integrating these datasets with omics information suggests that a majority of known protein complexes are symmetric. Finally, these datasets provide a structural context for interpreting disease mutations and reveal coiled-coil regions as major enablers of quaternary structure evolution in human. Our strategy is applicable to any organism and provides a comprehensive view of homo-oligomerization in proteomes.

Convergent evolution in the supercoiling of prokaryotic flagellar filaments.

The supercoiling of bacterial and archaeal flagellar filaments is required for motility. Archaeal flagellar filaments have no homology to their bacterial counterparts and are instead homologs of bacterial type IV pili. How these prokaryotic flagellar filaments, each composed of thousands of copies of identical subunits, can form stable supercoils under torsional stress is a fascinating puzzle for which structural insights have been elusive. Advances in cryoelectron microscopy (cryo-EM) make it now possible to directly visualize the basis for supercoiling, and here, we show the atomic structures of supercoiled bacterial and archaeal flagellar filaments. For the bacterial flagellar filament, we identify 11 distinct protofilament conformations with three broad classes of inter-protomer interface. For the archaeal flagellar filament, 10 protofilaments form a supercoil geometry supported by 10 distinct conformations, with one inter-protomer discontinuity creating a seam inside of the curve. Our results suggest that convergent evolution has yielded stable superhelical geometries that enable microbial locomotion.

Cellular bases of olfactory circuit assembly revealed by systematic time-lapse imaging.

Neural circuit assembly features simultaneous targeting of numerous neuronal processes from constituent neuron types, yet the dynamics is poorly understood. Here, we use the Drosophila olfactory circuit to investigate dynamic cellular processes by which olfactory receptor neurons (ORNs) target axons precisely to specific glomeruli in the ipsi- and contralateral antennal lobes. Time-lapse imaging of individual axons from 30 ORN types revealed a rich diversity in extension speed, innervation timing, and ipsilateral branch locations and identified that ipsilateral targeting occurs via stabilization of transient interstitial branches. Fast imaging using adaptive optics-corrected lattice light-sheet microscopy showed that upon approaching target, many ORN types exhibiting "exploring branches" consisted of parallel microtubule-based terminal branches emanating from an F-actin-rich hub. Antennal nerve ablations uncovered essential roles for bilateral axons in contralateral target selection and for ORN axons to facilitate dendritic refinement of postsynaptic partner neurons. Altogether, these observations provide cellular bases for wiring specificity establishment.

Structural and Proteomic Characterization of the Initiation of Giant Virus Infection.

Since their discovery, giant viruses have expanded our understanding of the principles of virology. Due to their gargantuan size and complexity, little is known about the life cycles of these viruses. To answer outstanding questions regarding giant virus infection mechanisms, we set out to determine biomolecular conditions that promote giant virus genome release. We generated four infection intermediates in Samba virus (Mimivirus genus, lineage A) as visualized by cryoelectron microscopy (cryo-EM), cryoelectron tomography (cryo-ET), and scanning electron microscopy (SEM). Each of these four intermediates reflects similar morphology to a stage that occurs in vivo. We show that these genome release stages are conserved in other mimiviruses. Finally, we identified proteins that are released from Samba and newly discovered Tupanvirus through differential mass spectrometry. Our work revealed the molecular forces that trigger infection are conserved among disparate giant viruses. This study is also the first to identify specific proteins released during the initial stages of giant virus infection.

A Polarizing Issue: Diversity in the Mechanisms Underlying Apico-Basolateral Polarization In Vivo.

Polarization along an apico-basolateral axis is a hallmark of epithelial cells and is essential for their selective barrier and transporter functions, as well as for their ability to provide mechanical resiliency to organs. Loss of polarity along this axis perturbs development and is associated with a wide number of diseases. We describe three steps involved in polarization: symmetry breaking, polarity establishment, and polarity maintenance. While the proteins involved in these processes are highly conserved among epithelial tissues and species, the execution of these steps varies widely and is context dependent. We review both theoretical principles underlying these steps and recent work demonstrating how apico-basolateral polarity is established in vivo in different tissues, highlighting how developmental and physiological contexts play major roles in the execution of the epithelial polarity program.

Structural Studies of Amyloid Proteins at the Molecular Level.

Dozens of proteins are known to convert to the aggregated amyloid state. These include fibrils associated with systemic and neurodegenerative diseases and cancer, functional amyloid fibrils in microorganisms and animals, and many denatured proteins. Amyloid fibrils can be much more stable than other protein assemblies. In contrast to globular proteins, a single protein sequence can aggregate into several distinctly different amyloid structures, termed polymorphs, and a given polymorph can reproduce itself by seeding. Amyloid polymorphs may be the molecular basis of prion strains. Whereas the Protein Data Bank contains some 100,000 globular protein and 3,000 membrane protein structures, only a few dozen amyloid protein structures have been determined, and most of these are short segments of full amyloid-forming proteins. Regardless, these amyloid structures illuminate the architecture of the amyloid state, including its stability and its capacity for formation of polymorphs.

Symmetry Breaking in the Mammalian Embryo.

We present an overview of symmetry breaking in early mammalian development as a continuous process from compaction to specification of the body axes. While earlier studies have focused on individual symmetry-breaking events, recent advances enable us to explore progressive symmetry breaking during early mammalian development. Although we primarily discuss embryonic development of the mouse, as it is the best-studied mammalian model system to date, we also highlight the shared and distinct aspects between different mammalian species. Finally, we discuss how insights gained from studying mammalian development can be generalized in light of self-organization principles. With this review, we hope to highlight new perspectives in studying symmetry breaking and self-organization in multicellular systems.

Structure of the human ATP synthase.

Biological energy currency ATP is produced by F1Fo-ATP synthase. However, the molecular mechanism for human ATP synthase action remains unknown. Here, we present snapshot images for three main rotational states and one substate of human ATP synthase using cryoelectron microscopy. These structures reveal that the release of ADP occurs when the ß subunit of F1Fo-ATP synthase is in the open conformation, showing how ADP binding is coordinated during synthesis. The accommodation of the symmetry mismatch between F1 and Fo motors is resolved by the torsional flexing of the entire complex, especially the γ subunit, and the rotational substep of the c subunit. Water molecules are identified in the inlet and outlet half-channels, suggesting that the proton transfer in these two half-channels proceed via a Grotthus mechanism. Clinically relevant mutations are mapped to the structure, showing that they are mainly located at the subunit-subunit interfaces, thus causing instability of the complex.

Activation and closed-state inactivation mechanisms of the human voltage-gated KV4 channel complexes.

The voltage-gated ion channel activity depends on both activation (transition from the resting state to the open state) and inactivation. Inactivation is a self-restraint mechanism to limit ion conduction and is as crucial to membrane excitability as activation. Inactivation can occur when the channel is open or closed. Although open-state inactivation is well understood, the molecular basis of closed-state inactivation has remained elusive. We report cryo-EM structures of human KV4.2 channel complexes in inactivated, open, and closed states. Closed-state inactivation of KV4 involves an unprecedented symmetry breakdown for pore closure by only two of the four S4-S5 linkers, distinct from known mechanisms of open-state inactivation. We further capture KV4 in a putative resting state, revealing how voltage sensor movements control the pore. Moreover, our structures provide insights regarding channel modulation by KChIP2 and DPP6 auxiliary subunits. Our findings elucidate mechanisms of closed-state inactivation and voltage-dependent activation of the KV4 channel.

Structure and desensitization of AMPA receptor complexes with type II TARP γ5 and GSG1L.

AMPA receptors (AMPARs) mediate the majority of excitatory neurotransmission. Their surface expression, trafficking, gating, and pharmacology are regulated by auxiliary subunits. Of the two types of TARP auxiliary subunits, type I TARPs assume activating roles, while type II TARPs serve suppressive functions. We present cryo-EM structures of GluA2 AMPAR in complex with type II TARP γ5, which reduces steady-state currents, increases single-channel conductance, and slows recovery from desensitization. Regulation of AMPAR function depends on its ligand-binding domain (LBD) interaction with the γ5 head domain. GluA2-γ5 complex shows maximum stoichiometry of two TARPs per AMPAR tetramer, being different from type I TARPs but reminiscent of the auxiliary subunit GSG1L. Desensitization of both GluA2-GSG1L and GluA2-γ5 complexes is accompanied by rupture of LBD dimer interface, while GluA2-γ5 but not GluA2-GSG1L LBD dimers remain two-fold symmetric. Different structural architectures and desensitization mechanisms of complexes with auxiliary subunits endow AMPARs with broad functional capabilities.

Establishment and maintenance of random monoallelic expression.

This Review elucidates the regulatory principles of random monoallelic expression by focusing on two well-studied examples: the X-chromosome inactivation regulator Xist and the olfactory receptor gene family. Although the choice of a single X chromosome or olfactory receptor occurs in different developmental contexts, common gene regulatory principles guide monoallelic expression in both systems. In both cases, an event breaks the symmetry between genetically and epigenetically identical copies of the gene, leading to the expression of one single random allele, stabilized through negative feedback control. Although many regulatory steps that govern the establishment and maintenance of monoallelic expression have been identified, key pieces of the puzzle are still missing. We provide an overview of the current knowledge and models for the monoallelic expression of Xist and olfactory receptors. We discuss their similarities and differences, and highlight open questions and approaches that could guide the study of other monoallelically expressed genes.

Gate-tunable subband degeneracy in semiconductor nanowires.

Degeneracy and symmetry have a profound relation in quantum systems. Here, we report gate-tunable subband degeneracy in PbTe nanowires with a nearly symmetric cross-sectional shape. The degeneracy is revealed in electron transport by the absence of a quantized plateau. Utilizing a dual gate design, we can apply an electric field to lift the degeneracy, reflected as emergence of the plateau. This degeneracy and its tunable lifting were challenging to observe in previous nanowire experiments, possibly due to disorder. Numerical simulations can qualitatively capture our observation, shedding light on device parameters for future applications.

Breaking the symmetry of cell contractility drives tubulogenesis via CXCL1 polarization.

The intricate interplay between biomechanical and biochemical pathways in modulating morphogenesis is an interesting research topic. How biomechanical force regulates epithelial cell tubulogenesis remains poorly understood. Here, we established a model of tubulogenesis by culturing renal proximal tubular epithelial cells on a collagen gel while manipulating contractile force. Epithelial cells were dynamically self-organized into tubule-like structures by augmentation of cell protrusions and cell-cell association. Reduction and asymmetric distribution of phosphorylated myosin light chain 2, the actomyosin contractility, in cells grown on soft matrix preceded tube connection. Notably, reducing matrix stiffness via sonication of collagen fibrils and inhibiting actomyosin contractility with blebbistatin promoted tubulogenesis, whereas inhibition of cytoskeleton polymerization suppressed it. CXC chemokine ligand 1 (CXCL1) expression was transcriptionally upregulated in cells undergoing tubulogenesis. Additionally, inhibiting actomyosin contractility facilitated CXCL1 polarization and cell protrusions preceding tube formation. Conversely, inhibiting the CXCL1-CXC receptor 1 pathway hindered cell protrusions and tubulogenesis. Mechanical property asymmetry with cell-collagen fibril interaction patterns at cell protrusions and along the tube structure supported the association of anisotropic contraction with tube formation. Furthermore, suppressing the mechanosensing machinery of integrin subunit beta 1 reduced CXCL1 expression, collagen remodeling, and impaired tubulogenesis. In summary, symmetry breaking of cell contractility on a soft collagen gel promotes CXCL1 polarization at cell protrusions which in turn facilitates cell-cell association and thus tubule connection.

Band gap opening of metallic single-walled carbon nanotubes via noncovalent symmetry breaking.

Covalent bonding interactions determine the energy-momentum (E-k) dispersion (band structure) of solid-state materials. Here, we show that noncovalent interactions can modulate the E-k dispersion near the Fermi level of a low-dimensional nanoscale conductor. We demonstrate that low energy band gaps may be opened in metallic carbon nanotubes through polymer wrapping of the nanotube surface at fixed helical periodicity. Electronic spectral, chiro-optic, potentiometric, electronic device, and work function data corroborate that the magnitude of band gap opening depends on the nature of the polymer electronic structure. Polymer dewrapping reverses the conducting-to-semiconducting phase transition, restoring the native metallic carbon nanotube electronic structure. These results address a long-standing challenge to develop carbon nanotube electronic structures that are not realized through disruption of π conjugation, and establish a roadmap for designing and tuning specialized semiconductors that feature band gaps on the order of a few hundred meV.

Democratizing protein language models with parameter-efficient fine-tuning.

Proteomics has been revolutionized by large protein language models (PLMs), which learn unsupervised representations from large corpora of sequences. These models are typically fine-tuned in a supervised setting to adapt the model to specific downstream tasks. However, the computational and memory footprint of fine-tuning (FT) large PLMs presents a barrier for many research groups with limited computational resources. Natural language processing has seen a similar explosion in the size of models, where these challenges have been addressed by methods for parameter-efficient fine-tuning (PEFT). In this work, we introduce this paradigm to proteomics through leveraging the parameter-efficient method LoRA and training new models for two important tasks: predicting protein-protein interactions (PPIs) and predicting the symmetry of homooligomer quaternary structures. We show that these approaches are competitive with traditional FT while requiring reduced memory and substantially fewer parameters. We additionally show that for the PPI prediction task, training only the classification head also remains competitive with full FT, using five orders of magnitude fewer parameters, and that each of these methods outperform state-of-the-art PPI prediction methods with substantially reduced compute. We further perform a comprehensive evaluation of the hyperparameter space, demonstrate that PEFT of PLMs is robust to variations in these hyperparameters, and elucidate where best practices for PEFT in proteomics differ from those in natural language processing. All our model adaptation and evaluation code is available open-source at https://github.com/microsoft/peft_proteomics. Thus, we provide a blueprint to democratize the power of PLM adaptation to groups with limited computational resources.

Quantitative videomicroscopy reveals latent control of cell-pair rotations in vivo.

Collective cell rotations are widely used during animal organogenesis. Theoretical and in vitro studies have conceptualized rotating cells as identical rigid-point objects that stochastically break symmetry to move monotonously and perpetually within an inert environment. However, it is unclear whether this notion can be extrapolated to a natural context, where rotations are ephemeral and heterogeneous cellular cohorts interact with an active epithelium. In zebrafish neuromasts, nascent sibling hair cells invert positions by rotating ≤180° around their geometric center after acquiring different identities via Notch1a-mediated asymmetric repression of Emx2. Here, we show that this multicellular rotation is a three-phasic movement that progresses via coherent homotypic coupling and heterotypic junction remodeling. We found no correlation between rotations and epithelium-wide cellular flow or anisotropic resistive forces. Moreover, the Notch/Emx2 status of the cell dyad does not determine asymmetric interactions with the surrounding epithelium. Aided by computer modeling, we suggest that initial stochastic inhomogeneities generate a metastable state that poises cells to move and spontaneous intercellular coordination of the resulting instabilities enables persistently directional rotations, whereas Notch1a-determined symmetry breaking buffers rotational noise.

An acytokinetic cell division creates PIP2-enriched membrane asymmetries leading to slit diaphragm assembly in Drosophila nephrocytes.

Vertebrate podocytes and Drosophila nephrocytes display slit diaphragms, specialised cell junctions that are essential for the execution of the basic excretory function of ultrafiltration. To elucidate the mechanisms of slit diaphragm assembly we have studied their formation in Drosophila embryonic garland nephrocytes. These cells of mesenchymal origin lack overt apical-basal polarity. We find that their initial membrane symmetry is broken by an acytokinetic cell division that generates PIP2-enriched domains at their equator. The PIP2-enriched equatorial cortex becomes a favourable domain for hosting slit diaphragm proteins and the assembly of the first slit diaphragms. Indeed, when this division is either prevented or forced to complete cytokinesis, the formation of diaphragms is delayed to larval stages. Furthermore, although apical polarity determinants also accumulate at the equatorial cortex, they do not appear to participate in the recruitment of slit diaphragm proteins. The mechanisms we describe allow the acquisition of functional nephrocytes in embryos, which may confer on them a biological advantage similar to the formation of the first vertebrate kidney, the pronephros.

Persistent corotation of the large-scale flow of thermal convection and an immersed free body.

Inspired by the superrotation of the Earth's solid core, we investigate the dynamics of a free-rotating body as it interacts with the large-scale circulation (LSC) of the Rayleigh-Bénard thermal convection in a cylindrical container. A surprising and persistent corotation of both the free body and the LSC emerges, breaking the axial symmetry of the system. The corotational speed increases monotonically with the intensity of thermal convection, measured by the Rayleigh number Ra, which is proportional to the temperature difference between the heated bottom and cooled top. The rotational direction occasionally and spontaneously reverses, occurring more frequently at higher Ra. The reversal events follow a Poisson process; it is feasible that flow fluctuations randomly interrupt and reestablish the rotation-sustaining mechanism. This corotation is powered by thermal convection alone and promoted by the addition of a free body, enriching the classical dynamical system.

The mechano-chemical circuit drives skin organoid self-organization.

Stem cells in organoids self-organize into tissue patterns with unknown mechanisms. Here, we use skin organoids to analyze this process. Cell behavior videos show that the morphological transformation from multiple spheroidal units with morphogenesis competence (CMU) to planar skin is characterized by two abrupt cell motility-increasing events before calming down. The self-organizing processes are controlled by a morphogenetic module composed of molecular sensors, modulators, and executers. Increasing dermal stiffness provides the initial driving force (driver) which activates Yap1 (sensor) in epidermal cysts. Notch signaling (modulator 1) in epidermal cyst tunes the threshold of Yap1 activation. Activated Yap1 induces Wnts and MMPs (epidermal executers) in basal cells to facilitate cellular flows, allowing epidermal cells to protrude out from the CMU. Dermal cell-expressed Rock (dermal executer) generates a stiff force bridge between two CMU and accelerates tissue mixing via activating Laminin and ß1-integrin. Thus, this self-organizing coalescence process is controlled by a mechano-chemical circuit. Beyond skin, self-organization in organoids may use similar mechano-chemical circuit structures.

Quantum reality with negative-mass particles.

Physical interpretations of the time-symmetric formulation of quantum mechanics, due to Aharonov, Bergmann, and Lebowitz are discussed in terms of weak values. The most direct, yet somewhat naive, interpretation uses the time-symmetric formulation to assign eigenvalues to unmeasured observables of a system, which results in logical paradoxes, and no clear physical picture. A top-down ontological model is introduced that treats the weak values of observables as physically real during the time between pre- and post-selection (PPS), which avoids these paradoxes. The generally delocalized rank-1 projectors of a quantum system describe its fundamental ontological elements, and the highest-rank projectors corresponding to individual localized objects describe an emergent particle model, with unusual particles, whose masses and energies may be negative or imaginary. This retrocausal top-down model leads to an intuitive particle-based ontological picture, wherein weak measurements directly probe the properties of these exotic particles, which exist whether or not they are actually measured.