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Spinal cord injury (SCI) disrupts the structural and functional connectivity between the higher center and the spinal cord, resulting in severe motor, sensory, and autonomic dysfunction with a variety of complications. The pathophysiology of SCI is complicated and multifaceted, and thus individual treatments acting on a specific aspect or process are inadequate to elicit neuronal regeneration and functional recovery after SCI. Combinatory strategies targeting multiple aspects of SCI pathology have achieved greater beneficial effects than individual therapy alone. Although many problems and challenges remain, the encouraging outcomes that have been achieved in preclinical models offer a promising foothold for the development of novel clinical strategies to treat SCI. In this review, we characterize the mechanisms underlying axon regeneration of adult neurons and summarize recent advances in facilitating functional recovery following SCI at both the acute and chronic stages. In addition, we analyze the current status, remaining problems, and realistic challenges towards clinical translation. Finally, we consider the future of SCI treatment and provide insights into how to narrow the translational gap that currently exists between preclinical studies and clinical practice. Going forward, clinical trials should emphasize multidisciplinary conversation and cooperation to identify optimal combinatorial approaches to maximize therapeutic benefit in humans with SCI.

Mammalian respiratory complex I (CI) is a 45-subunit, redox-driven proton pump that generates an electrochemical gradient across the mitochondrial inner membrane to power ATP synthesis in mitochondria. In the present study, we report cryo-electron microscopy structures of CI from Sus scrofa in six treatment conditions at a resolution of 2.4–3.5 Å, in which CI structures of each condition can be classified into two biochemical classes (active or deactive), with a notably higher proportion of active CI particles. These structures illuminate how hydrophobic ubiquinone-10 (Q10) with its long isoprenoid tail is bound and reduced in a narrow Q chamber comprising four different Q10-binding sites. Structural comparisons of active CI structures from our decylubiquinone–NADH and rotenone–NADH datasets reveal that Q10 reduction at site 1 is not coupled to proton pumping in the membrane arm, which might instead be coupled to Q10 oxidation at site 2. Our data overturn the widely accepted previous proposal about the coupling mechanism of CI. Cryo-EM structures of the porcine respiratory complex I reveal how Q10 is bound and reduced in the Q chamber comprising four different Q10-binding sites. A ‘two-Q’ model is proposed for the coupling mechanism of complex I.

The respiratory chain complexes I, III and IV (CI, CIII and CIV) are present in the bacterial membrane or the inner mitochondrial membrane and have a role of transferring electrons and establishing the proton gradient for ATP synthesis by complex V. The respiratory chain complexes can assemble into supercomplexes (SCs), but their precise arrangement is unknown. Here we report a 5.4 Å cryo-electron microscopy structure of the major 1.7 megadalton SCI 1 III 2 IV 1 respirasome purified from porcine heart. The CIII dimer and CIV bind at the same side of the L-shaped CI, with their transmembrane domains essentially aligned to form a transmembrane disk. Compared to free CI, the CI in the respirasome is more compact because of interactions with CIII and CIV. The NDUFA11 and NDUFB9 supernumerary subunits of CI contribute to the oligomerization of CI and CIII. The structure of the respirasome provides information on the precise arrangements of the respiratory chain complexes in mitochondria. Respirasomes are supercomplexes of mitochondrial electron transport chain complexes that are responsible for cellular respiration and energy production; a cryo-electron microscopy structural study of the respirasome is presented. Inside the mammalian respirasome supercomplex Mitochondrial electron transport chain complexes are responsible for cellular respiration and energy production. They are organized in supercomplexes called respirasomes. Two studies in this issue of Nature report cryo-electron microscopy structures of the supercomplex consisting of complex I, the dimer of complex III and complex IV at resolutions ranging from 5.4 Å to 7.8 Å. Maojun Yang and colleagues study the respirasome isolated from porcine heart, whereas Leonid Sazanov and colleagues obtain it from ovine heart. The structures provide insights into the organization of subunits within complexes and the interactions between the complexes.

Respiration is one of the most basic features of living organisms, and the electron transport chain complexes are probably the most complicated protein system in mitochondria. Complex-IV is the terminal enzyme of the electron transport chain, existing either as randomly scattered complexes or as a component of supercomplexes. NDUFA4 was previously assumed as a subunit of complex-I, but recent biochemical data suggested it may be a subunit of complex-IV. However, no structural evidence supporting this notion was available till now. Here we obtained the 3.3 Å resolution structure of complex-IV derived from the human supercomplex I 1 III 2 IV 1 and assigned the NDUFA4 subunit into complex-IV. Intriguingly, NDUFA4 lies exactly at the dimeric interface observed in previously reported crystal structures of complex-IV homodimer which would preclude complex-IV dimerization. Combining previous structural and biochemical data shown by us and other groups, we propose that the intact complex-IV is a monomer containing 14 subunits.

The mitochondrial adenosine triphosphate (ATP) synthase produces most of the ATP required by mammalian cells. We isolated porcine tetrameric ATP synthase and solved its structure at 6.2-angstrom resolution using a single-particle cryo–electron microscopy method. Two classical V-shaped ATP synthase dimers lie antiparallel to each other to form an H-shaped ATP synthase tetramer, as viewed from the matrix. ATP synthase inhibitory factor subunit 1 (IF1) is a well-known in vivo inhibitor of mammalian ATP synthase at low pH. Two IF1 dimers link two ATP synthase dimers, which is consistent with the ATP synthase tetramer adopting an inhibited state. Within the tetramer, we refined structures of intact ATP synthase in two different rotational conformations at 3.34- and 3.45-Å resolution.

Piezo proteins are evolutionarily conserved and functionally diverse mechanosensitive cation channels. However, the overall structural architecture and gating mechanisms of Piezo channels have remained unknown. Here we determine the cryo-electron microscopy structure of the full-length (2,547 amino acids) mouse Piezo1 (Piezo1) at a resolution of 4.8 Å. Piezo1 forms a trimeric propeller-like structure (about 900 kilodalton), with the extracellular domains resembling three distal blades and a central cap. The transmembrane region has 14 apparently resolved segments per subunit. These segments form three peripheral wings and a central pore module that encloses a potential ion-conducting pore. The rather flexible extracellular blade domains are connected to the central intracellular domain by three long beam-like structures. This trimeric architecture suggests that Piezo1 may use its peripheral regions as force sensors to gate the central ion-conducting pore. Piezo1, a mechanosensitive cation channel, senses shear stress of blood flow for proper blood vessel development, regulates red blood cell function and controls cell migration and differentiation; here a trimeric architecture of this novel class of ion channel is reported, suggesting that Piezo1 may use its peripheral propeller-like ‘blades’ as force sensors to gate the central ion-conducting pore. Piezo1 channel architecture Mechanosensitive cation channels couple mechanical stimuli to various biological activities, including touch, hearing and blood pressure regulation, through a process termed 'mechanotransduction'. Piezo proteins have recently been identified as pore-forming subunits of mechanosensitive cation channels in metazoans, and Piezo1 senses shear stress of blood flow for proper blood vessel development, regulates red blood cell function, and controls cell migration and differentiation. These authors report a high-resolution cryo-electron microscopy structure of the full-length mouse Piezo1, which reveals the trimeric three-bladed propeller-like architecture of this membrane protein. They propose that Piezo1 may use its peripheral propeller-like 'blades' as force sensors to gate the central ion-conducting pore.

Respiration is one of the most vital and basic features of living organisms. In mammals, respiration is accomplished by respiratory chain complexes located on the mitochondrial inner membrane. In the past century, scientists put tremendous efforts in understanding these complexes, but failed to solve the high resolution structure until recently. In 2016, three research groups reported the structure of respiratory chain supercomplex from different species, and fortunately the structure solved by our group has the highest resolution. In this review, we will compare the recently solved structures of respirasome, probe into the relationship between cristae shape and respiratory chain organization, and discuss the highly disputed issues afterwards. Besides, our group reported the first high resolution structure of respirasome and medium resolution structure of megacomplex from cultured human cells this year. Definitely, these supercomplex structures will provide precious information for conquering the mitochondrial malfunction diseases.

Mutations in epidermal growth factor receptor (EGFR) play critical roles in the pathogenesis of non-small cell lung cancer (NSCLC), and they are highly associated with sensitivity to tyrosine kinase inhibitors (TKIs). While the pathogenic and pharmacological characteristics of common mutations in EGFR have been thoroughly investigated, those of uncommon mutations remain to be elucidated. Traditional approaches to study common mutations by randomized controlled trials are not feasible for uncommon mutations owing to their rarity. Therefore, by systematically reviewing laboratory and clinical studies of the G719X mutation, one of the uncommon mutations, we concluded that the G719X mutation was intermediately sensitive to TKIs, with an average response rate of 35.1% (47/134). Moreover, accordingly, we proposed a comprehensive model to investigate uncommon mutations in EGFR. The model involves both basic and clinical components, composed of structural analyses, functional alterations, cell viabilities and animal models with various types of clinical studies. In this review, we systematically reviewed studies of the G719X mutation and put forward a research model that could be generalized to explore uncommon mutations in diseases associated with gene mutations.

DNA replication in eukaryotes is strictly regulated by several mechanisms. A central step in this replication is the assembly of the heterohexameric minichromosome maintenance (MCM2–7) helicase complex at replication origins during G1 phase as an inactive double hexamer. Here, using cryo-electron microscopy, we report a near-atomic structure of the MCM2–7 double hexamer purified from yeast G1 chromatin. Our structure shows that two single hexamers, arranged in a tilted and twisted fashion through interdigitated amino-terminal domain interactions, form a kinked central channel. Four constricted rings consisting of conserved interior β-hairpins from the two single hexamers create a narrow passageway that tightly fits duplex DNA. This narrow passageway, reinforced by the offset of the two single hexamers at the double hexamer interface, is flanked by two pairs of gate-forming subunits, MCM2 and MCM5. These unusual features of the twisted and tilted single hexamers suggest a concerted mechanism for the melting of origin DNA that requires structural deformation of the intervening DNA. Cryo-electron microscopy is used to visualize the double hexamer of the eukaryotic minichromosome maintenance complex (MCM), which is assembled during the G1 phase of DNA replication; two interdigitated hexamers have a central channel that tightly fits a DNA duplex, and the orientation of the tilted single hexamers sheds light on many functional aspects, particularly in the initial origin DNA melting. Replication-ready MCM complex In eukaryotes, DNA replication begins with the binding of a hexameric ring of minichromosome maintenance (MCM) proteins at regions known as replication origins during the G1 phase of the cell cycle. The resulting complex is dormant until the cell enters S phase, when replication occurs. This entails conversion of an MCM double hexamer into an active species, but the structure of this complex was unknown. Ning Gao and colleagues have used cryo-electron microscopy to visualize the double hexamer complex. They observe two interdigitated hexamers that have a central channel that tightly fits a DNA duplex. The orientation of the single rings suggests models in which relative movements between the two hexamers would deform the origin DNA so that other replication proteins can bind to the melted DNA double helix.

CRISPR-Cas systems are prokaryotic adaptive immune systems and phages use anti-CRISPR proteins (Acrs) to counteract these systems. Here, we report the structures of AcrIF24 and its complex with the crRNA-guided surveillance (Csy) complex. The HTH motif of AcrIF24 can bind the Acr promoter region and repress its transcription, suggesting its role as an Aca gene in self-regulation. AcrIF24 forms a homodimer and further induces dimerization of the Csy complex. Apart from blocking the hybridization of target DNA to the crRNA, AcrIF24 also induces the binding of non-sequence-specific dsDNA to the Csy complex, similar to AcrIF9, although this binding seems to play a minor role in AcrIF24 inhibitory capacity. Further structural and biochemical studies of the Csy-AcrIF24-dsDNA complexes and of AcrIF24 mutants reveal that the HTH motif of AcrIF24 and the PAM recognition loop of the Csy complex are structural elements essential for this non-specific dsDNA binding. Moreover, AcrIF24 and AcrIF9 display distinct characteristics in inducing non-specific DNA binding. Together, our findings highlight a multifunctional Acr and suggest potential wide distribution of Acr-induced non-specific DNA binding. Phages use anti-CRISPR proteins (Acrs) to counteract the bacterial CRISPR-Cas systems. Here, the authors characterize AcrIF24, which functions as an Aca (Acr-associated) to repress and regulate its own transcription, dimerizes the Csy complex, blocks the hybridization of target DNA, and tethers non-sequence-specific DNA to the Csy complex.