A genome assembly of ginger (Zingiber officinale Roscoe) provides insights into genome evolution and 6-gingerol biosynthesis.

Ginger is cultivated in tropical and subtropical regions and is one of the most crucial spices worldwide owing to its special taste and scent. Here, we present a high-quality genome assembly for 'Small Laiwu Ginger', a famous cultivated ginger in northern China. The ginger genome was phased into two haplotypes, haplotype A (1.55Gb), and haplotype B (1.44Gb). Analysis of Ty1/Copia and Ty3/Gypsy LTR retrotransposon families revealed that both have undergone multiple retrotransposon bursts about 0-1 million years ago. In addition to a recent whole-genome duplication event, there has been a lineage-specific expansion of genes involved in stilbenoid, diarylheptanoid, and gingerol biosynthesis, thereby enhancing 6-gingerol biosynthesis. Furthermore, we focused on the biosynthesis of 6-gingerol, the most important gingerol, and screened key transcription factors ZoMYB106 and ZobHLH148 that regulate 6-gingerol synthesis by transcriptomic and metabolomic analysis in the ginger rhizome at four growth stages. The results of yeast one-hybrid, electrophoretic mobility shift, and dual-luciferase reporter gene assays showed that both ZoMYB106 and ZobHLH148 bind to the promoters of the key rate-limiting enzyme genes ZoCCOMT1 and ZoCCOMT2 in the 6-gingerol synthesis pathway and promote their transcriptional activities. The reference genome, transcriptome, and metabolome data pave the way for further research on the molecular mechanism underlying the biosynthesis of 6-gingerol. Furthermore, it provides precious new resources for the study on the biology and molecular breeding of ginger.

Ionic Bonding Without Directionality Facilitates Efficient Interfacial Bridging for Perovskite Solar Cells.

Interface passivation through Lewis acid-base coordinate chemistry in perovskite solar cells (PSCs) is a universal strategy to reduce interface defects and hinder ion migration. However, the formation of coordinate covalent bonding demands strict directional alignment of coordinating atoms. Undoubtedly, this limits the selected range of the interface passivation molecules, because a successful molecular bridge between charge transport layer and perovskite bottom interface needs a well-placed molecular orientation. In this study, it is discovered that potassium ions can migrate to the hollow sites of multiple iodine ions from perovskite to form K-Ix ionic bonding, and the ionic bonds without directionality can support molecular backbone rotation to facilitate polar sites (carboxyl groups) chelating Pb at the bottom perovskite interface, finally forming a closed-loop bonding structure. The synergy of coordinate and ionic bonding significantly reduces interface defects, changes electric field distribution, and immobilizes iodine at the perovskite bottom interface, resulting in eliminating the hysteresis effect and enhancing the performance of PSCs. As a result, the corresponding devices achieve a high efficiency exceeding 24.5% (0.09 cm2), and a mini-module with 21% efficiency (12.4 cm2). These findings provide guidelines for designing molecular bridging strategies at the buried interface of PSCs.

Light-Mediated Enhancement of Glucose-Stimulated Insulin Release of Optogenetically Engineered Human Pancreatic Beta-Cells.

Enhancement of glucose-stimulated insulin secretion (GSIS) in exogenously delivered pancreatic ß-cells is desirable, for example, to overcome the insulin resistance manifested in type 2 diabetes or to reduce the number of ß-cells for supporting homeostasis of blood sugar in type 1 diabetes. Optogenetically engineered cells can potentiate their function with exposure to light. Given that cyclic adenosine monophosphate (cAMP) mediates GSIS, we surmised that optoamplification of GSIS is feasible in human ß-cells carrying a photoactivatable adenylyl cyclase (PAC). To this end, human EndoC-ßH3 cells were engineered to express a blue-light-activated PAC, and a workflow was established combining the scalable manufacturing of pseudoislets (PIs) with efficient adenoviral transduction, resulting in over 80% of cells carrying PAC. Changes in intracellular cAMP and GSIS were determined with the photoactivation of PAC in vitro as well as after encapsulation and implantation in mice with streptozotocin-induced diabetes. cAMP rapidly rose in ß-cells expressing PAC with illumination and quickly declined upon its termination. Light-induced amplification in cAMP was concomitant with a greater than 2-fold GSIS vs ß-cells without PAC in elevated glucose. The enhanced GSIS retained its biphasic pattern, and the rate of oxygen consumption remained unchanged. Diabetic mice receiving the engineered ß-cell PIs exhibited improved glucose tolerance upon illumination compared to those kept in the dark or not receiving cells. The findings support the use of optogenetics for molecular customization of the ß-cells toward better treatments for diabetes without the adverse effects of pharmacological approaches.

Precise CRISPR/Cpf1 genome editing system in the Deinococcus radiodurans with superior DNA repair mechanisms.

Deinococcus radiodurans, with its high homologous recombination (HR) efficiency of double-stranded DNA breaks (DSBs), is a model organism for studying genome stability maintenance and an attractive microbe for industrial applications. Here, we developed an efficient CRISPR/Cpf1 genome editing system in D. radiodurans by evaluating and optimizing double-plasmid strategies and four Cas effector proteins from various organisms, which can precisely introduce different types of template-dependent mutagenesis without off-target toxicity. Furthermore, the role of DNA repair genes in determining editing efficiency in D. radiodurans was evaluated by introducing the CRISPR/Cpf1 system into 13 mutant strains lacking various DNA damage response and repair factors. In addition to the crucial role of RecA-dependent HR required for CRISPR/Cpf1 editing, D. radiodurans showed higher editing efficiency when lacking DdrB, the single-stranded DNA annealing (SSA) protein involved in the RecA-independent DSB repair pathway. This suggests a possible competition between HR and SSA pathways in the CRISPR editing of D. radiodurans. Moreover, off-target effects were observed during the genome editing of the pprI knockout strain, a master DNA damage response gene in Deinococcus species, which suggested that precise regulation of DNA damage response is critical for a high-fidelity genome editing system.

The Deinococcus protease PprI senses DNA damage by directly interacting with single-stranded DNA.

Bacteria have evolved various response systems to adapt to environmental stress. A protease-based derepression mechanism in response to DNA damage was characterized in Deinococcus, which is controlled by the specific cleavage of repressor DdrO by metallopeptidase PprI (also called IrrE). Despite the efforts to document the biochemical, physiological, and downstream regulation of PprI-DdrO, the upstream regulatory signal activating this system remains unclear. Here, we show that single-stranded DNA physically interacts with PprI protease, which enhances the PprI-DdrO interactions as well as the DdrO cleavage in a length-dependent manner both in vivo and in vitro. Structures of PprI, in its apo and complexed forms with single-stranded DNA, reveal two DNA-binding interfaces shaping the cleavage site. Moreover, we show that the dynamic monomer-dimer equilibrium of PprI is also important for its cleavage activity. Our data provide evidence that single-stranded DNA could serve as the signal for DNA damage sensing in the metalloprotease/repressor system in bacteria. These results also shed light on the survival and acquired drug resistance of certain bacteria under antimicrobial stress through a SOS-independent pathway.

Femtosecond Electron Diffraction Reveals Local Disorder and Local Anharmonicity in Thermoelectric SnSe.

In addition to long-range periodicity, local disorder, with local structures deviating from the average lattice structure, dominates the physical properties of phonons, electrons, and spin subsystems in crystalline functional materials. Experimentally characterizing the 3D atomic configuration of such a local disorder and correlating it with advanced functions remains challenging. Using a combination of femtosecond electron diffraction, structure factor calculations, and time-dependent density functional theory molecular dynamics simulations, the static local disorder and its local anharmonicity in thermoelectric SnSe are identified exclusively. The ultrafast structural dynamics reveal that the crystalline SnSe is composed of multiple locally correlated configurations dominated by the static off-symmetry displacements of Sn (≈0.4 Å) and such a set of locally correlated structures is termed local disorder. Moreover, the anharmonicity of this local disorder induces an ultrafast atomic displacement within 100 fs, indicating the signature of probable THz Einstein oscillators. The identified local disorder and local anharmonicity suggest a glass-like thermal transport channel, which updates the fundamental insight into the long-debated ultralow thermal conductivity of SnSe. The method of revealing the 3D local disorder and the locally correlated interactions by ultrafast structural dynamics will inspire broad interest in the construction of structure-property relationships in material science.

Biomarkers of Inflammation and Longitudinal Evaluation of Lung Function, Physical Activity, and Grip Strength: A Secondary Analysis in the CASCADE Study.

Rationale: Physical activity, lung function, and grip strength are associated with exacerbations, hospitalizations, and mortality in people with chronic obstructive pulmonary disease (COPD). We tested whether baseline inflammatory biomarkers were associated with longitudinal outcomes of these physiologic measurements. Methods: The COPD Activity: Serotonin Transporter, Cytokines, and Depression (CASCADE) study was a prospective observational study of individuals with COPD. A total of 14 inflammatory biomarkers were measured at baseline. Participants were followed for 2 years. We analyzed associations between baseline biomarkers and forced expiratory volume in 1 second (FEV1), physical activity, and grip strength. We used a hierarchical hypothesis testing procedure to reduce type I error. We used Pearson correlations to test associations between baseline biomarkers and longitudinal changes in the outcomes of interest. We used Fisher's linear discriminant analysis to test if linear combinations of baseline biomarkers predict rapid FEV1 decline. Finally, we used linear mixed modeling to test associations between baseline biomarkers and outcomes of interest at baseline, year 1, and year 2; models were adjusted for age, smoking status, baseline biomarkers, and FEV1. Results: A total of 302 participants (age 67.5 ± 8.5 years, 19.5% female, 28.5% currently smoking) were included. Baseline biomarkers were not associated with longitudinal changes in grip strength, physical activity, or rapid FEV1 decline. Higher interleukin-6 and C-reactive protein were associated with lower physical activity at baseline and these relationships persisted at year 1 and year 2. Conclusion: Baseline inflammatory biomarkers did not predict changes in lung function or physical activity, but higher inflammatory biomarkers were associated with persistently low levels of physical activity.

Origin of the multi-phasic quenching dynamics in the BLUF domains across the species.

Blue light using flavin (BLUF) photoreceptors respond to light via one of nature's smallest photo-switching domains. Upon photo-activation, the flavin cofactor in the BLUF domain exhibits multi-phasic dynamics, quenched by a proton-coupled electron transfer reaction involving the conserved Tyr and Gln. The dynamic behavior varies drastically across different species, the origin of which remains controversial. Here, we incorporate site-specific fluorinated Trp into three BLUF proteins, i.e., AppA, OaPAC and SyPixD, and characterize the percentages for the Wout, WinNHin and WinNHout conformations using 19F nuclear magnetic resonance spectroscopy. Using femtosecond spectroscopy, we identify that one key WinNHin conformation can introduce a branching one-step proton transfer in AppA and a two-step proton transfer in OaPAC and SyPixD. Correlating the flavin quenching dynamics with the active-site structural heterogeneity, we conclude that the quenching rate is determined by the percentage of WinNHin, which encodes a Tyr-Gln configuration that is not conducive to proton transfer.

A Practical Approach for Targeting Structural Variants Genome-wide in Plasma Cell-free DNA.

Interrogating plasma cell-free DNA (cfDNA) to detect cancer offers promise; however, no current tests scan structural variants (SVs) throughout the genome. Here, we report a simple molecular workflow to enrich a tumorigenic SV (DNA palindromes/fold-back inversions) that often demarcates genomic amplification and its feasibility for cancer detection by combining low-throughput next-generation sequencing with automated machine learning (Genome-wide Analysis of Palindrome Formation, GAPF-seq). Tumor DNA signal manifested as skewed chromosomal distributions of high-coverage 1-kb bins (HCBs), differentiating 39 matched breast tumor DNA from normal DNA with an average AUC of 0.9819. In a proof-of-concept liquid biopsy study, cfDNA from 0.5 mL plasma from prostate cancer patients was sufficient for binary classification against matched buffy coat DNA with an average AUC of 0.965. HCBs on the X chromosome emerged as a determinant feature and were associated with AR amplification. GAPF-seq could generate unique cancer-specific SV profiles in an agnostic liquid biopsy setting.

The Zn2+ transporter ZIP7 enhances endoplasmic-reticulum-associated protein degradation and prevents neurodegeneration in Drosophila.

Proteotoxic stress drives numerous degenerative diseases. Cells initially adapt to misfolded proteins by activating the unfolded protein response (UPR), including endoplasmic-reticulum-associated protein degradation (ERAD). However, persistent stress triggers apoptosis. Enhancing ERAD is a promising therapeutic approach for protein misfolding diseases. The ER-localized Zn2+ transporter ZIP7 is conserved from plants to humans and required for intestinal self-renewal, Notch signaling, cell motility, and survival. However, a unifying mechanism underlying these diverse phenotypes was unknown. In studying Drosophila border cell migration, we discovered that ZIP7-mediated Zn2+ transport enhances the obligatory deubiquitination of proteins by the Rpn11 Zn2+ metalloproteinase in the proteasome lid. In human cells, ZIP7 and Zn2+ are limiting for deubiquitination. In a Drosophila model of neurodegeneration caused by misfolded rhodopsin (Rh1), ZIP7 overexpression degrades misfolded Rh1 and rescues photoreceptor viability and fly vision. Thus, ZIP7-mediated Zn2+ transport is a previously unknown, rate-limiting step for ERAD in vivo with therapeutic potential in protein misfolding diseases.

Global profiling of functional histidines in live cells using small-molecule photosensitizer and chemical probe relay labelling.

Recent advances in chemical proteomics have focused on developing chemical probes that react with nucleophilic amino acid residues. Although histidine is an attractive candidate due to its importance in enzymatic catalysis, metal binding and protein-protein interaction, its moderate nucleophilicity poses challenges. Its modification is frequently influenced by cysteine and lysine, which results in poor selectivity and narrow proteome coverage. Here we report a singlet oxygen and chemical probe relay labelling method that achieves high selectivity towards histidine. Libraries of small-molecule photosensitizers and chemical probes were screened to optimize histidine labelling, enabling histidine profiling in live cells with around 7,200 unique sites. Using NMR spectroscopy and X-ray crystallography, we characterized the reaction mechanism and the structures of the resulting products. We then applied this method to discover unannotated histidine sites key to enzymatic activity and metal binding in select metalloproteins. This method also revealed the accessibility change of histidine mediated by protein-protein interaction that influences select protein subcellular localization, underscoring its capability in discovering functional histidines.

A Functional InDel in the WRKY10 Promoter Controls the Degree of Flesh Red Pigmentation in Apple.

MYB transcription factors have been linked to anthocyanin synthesis and various color phenotypes in plants. In apple, MYB10 confers a red-flesh phenotype due to a minisatellite insertion in its R6 promoter, but R6:MYB10 genotypes exhibit various degrees of red pigmentation in the flesh, suggesting the involvement of other genetic factors. Here, it is shown that MdWRKY10, a transcription factor identified via DNA pull-down trapping, binds to the promoter of MdMYB10 and activates its transcription. MdWRKY10 specifically interacts with the WDR protein MdTTG1 to join the apple MYB-bHLH-WDR (MBW) complex, which significantly enhances its transcriptional activation activity. A 163-bp InDel detected in the promoter region of the alleles of MdWRKY10 in a hybrid population of identical heterozygous genotypes regarding R6 by structural variation analysis, contains a typical W-box element that MdWRKY10 binds to for transactivation. This leads to increased transcript levels of MdWRKY10 and MdMYB10 and enhanced anthocyanin synthesis in the flesh, largely accounting for the various degrees of flesh red pigmentation in the R6 background. These findings reveal a novel regulatory role of the WRKY-containing protein complex in the formation of red flesh apple phenotypes and provide broader insights into the molecular mechanism governing anthocyanin synthesis in plants.

An Integrated Optogenetic and Bioelectronic Platform for Regulating Cardiomyocyte Function.

Bioelectronic medicine is emerging as a powerful approach for restoring lost endogenous functions and addressing life-altering maladies such as cardiac disorders. Systems that incorporate both modulation of cellular function and recording capabilities can enhance the utility of these approaches and their customization to the needs of each patient. Here is report an integrated optogenetic and bioelectronic platform for stable and long-term stimulation and monitoring of cardiomyocyte function in vitro. Optical inputs are achieved through the expression of a photoactivatable adenylyl cyclase, that when irradiated with blue light causes a dose-dependent and time-limited increase in the secondary messenger cyclic adenosine monophosphate with subsequent rise in autonomous cardiomyocyte beating rate. Bioelectronic readouts are obtained through a multi-electrode array that measures real-time electrophysiological responses at 32 spatially-distinct locations. Irradiation at 27 µW mm-2 results in a 14% elevation of the beating rate within 20-25 min, which remains stable for at least 2 h. The beating rate can be cycled through "on" and "off" light states, and its magnitude is a monotonic function of irradiation intensity. The integrated platform can be extended to stretchable and flexible substrates, and can open new avenues in bioelectronic medicine, including closed-loop systems for cardiac regulation and intervention, for example, in the context of arrythmias.

An Integrated Optogenetic and Bioelectronic Platform for Regulating Cardiomyocyte Function.

We report an integrated optogenetic and bioelectronic platform for stable and long-term modulation and monitoring of cardiomyocyte function in vitro. Optogenetic inputs were achieved through expression of a photoactivatable adenylyl cyclase (bPAC), that when activated by blue light caused a dose-dependent and time-limited increase in autonomous cardiomyocyte beat rate. Bioelectronic readouts were achieved through an integrated planar multi-electrode array (MEA) that provided real-time readouts of electrophysiological activity from 32 spatially-distinct locations. Irradiation at 27 µW/mm2 resulted in a ca. 14% increase in beat rate within 20-25 minutes, which remained stable for at least 2 hours. The beating rate could be cycled through repeated "on" and "off' states, and its magnitude was a monotonic function of irradiation intensity. Our integrated platform opens new avenues in bioelectronic medicine, including closed-loop feedback systems, with potential applications for cardiac regulation including arrhythmia diagnosis and intervention.