Sensory error drives fine motor adjustment.

Fine audiovocal control is a hallmark of human speech production and depends on precisely coordinated muscle activity guided by sensory feedback. Little is known about shared audiovocal mechanisms between humans and other mammals. We hypothesized that real-time audiovocal control in bat echolocation uses the same computational principles as human speech. To test the prediction of this hypothesis, we applied state feedback control (SFC) theory to the analysis of call frequency adjustments in the echolocating bat, Hipposideros armiger. This model organism exhibits well-developed audiovocal control to sense its surroundings via echolocation. Our experimental paradigm was analogous to one implemented in human subjects. We measured the bats' vocal responses to spectrally altered echolocation calls. Individual bats exhibited highly distinct patterns of vocal compensation to these altered calls. Our findings mirror typical observations of speech control in humans listening to spectrally altered speech. Using mathematical modeling, we determined that the same computational principles of SFC apply to bat echolocation and human speech, confirming the prediction of our hypothesis.

Rice OsPUB16 modulates the 'SAPK9-OsMADS23-OsAOC' pathway to reduce plant water-deficit tolerance by repressing ABA and JA biosynthesis.

Ubiquitin-mediated proteolysis plays crucial roles in plant responses to environmental stress. However, the mechanism by which E3 ubiquitin ligases modulate plant stress response still needs to be elucidated. In this study, we found that rice PLANT U-BOX PROTEIN 16 (OsPUB16), a U-box E3 ubiquitin ligase, negatively regulates rice drought response. Loss-of-function mutants of OsPUB16 generated through CRISPR/Cas9 system exhibited the markedly enhanced water-deficit tolerance, while OsPUB16 overexpression lines were hypersensitive to water deficit stress. Moreover, OsPUB16 negatively regulated ABA and JA response, and ospub16 mutants produced more endogenous ABA and JA than wild type when exposed to water deficit. Mechanistic investigations revealed that OsPUB16 mediated the ubiquitination and degradation of OsMADS23, which is the substrate of OSMOTIC STRESS/ABA-ACTIVATED PROTEIN KINASE 9 (SAPK9) and increases rice drought tolerance by promoting ABA biosynthesis. Further, the ChIP-qPCR analysis and transient transactivation activity assays demonstrated that OsMADS23 activated the expression of JA-biosynthetic gene OsAOC by binding to its promoter. Interestingly, SAPK9-mediated phosphorylation on OsMADS23 reduced its ubiquitination level by interfering with the OsPUB16-OsMADS23 interaction, which thus enhanced OsMADS23 stability and promoted OsAOC expression. Collectively, our findings establish that OsPUB16 reduces plant water-deficit tolerance by modulating the 'SAPK9-OsMADS23-OsAOC' pathway to repress ABA and JA biosynthesis.

A Route to Two-Dimensional Room-Temperature Organometallic Multiferroics: The Marriage of d-p Spin Coupling and Structural Inversion Symmetry Breaking.

Two-dimensional (2D) room-temperature multiferroic materials are highly desirable but still very limited. Herein, we propose a potential strategy to obtain such materials in 2D metal-organic frameworks (MOFs) by utilizing the d-p direct spin coupling in conjunction with center-symmetry-breaking six-membered heterocyclic rings. Based on this strategy, a screening of 128 2D MOFs results in the identification of three multiferroics, that is, Cr(1,2-oxazine)2, Cr(1,2,4-triazine)2, and Cr(1,2,3,4-trazine)2, simultaneously exhibiting room-temperature ferrimagnetism and ferroelectricity/antiferroelectricity. The room-temperature ferrimagnetic order (306-495 K) in these MOFs originates from the strong d-p direct magnetic exchange interaction between Cr cations and ligand anions. Specifically, Cr(1,2-oxazine)2 exhibits ferroelectric behavior with an out-of-plane polarization of 4.24 pC/m, whereas the other two manifest antiferroelectric characteristics. Notably, all three materials present suitable polarization switching barriers (0.18-0.31 eV). Furthermore, these MOFs are all bipolar magnetic semiconductors with moderate band gaps, in which the spin direction of carriers can be manipulated by electrical gating.

Vertical Dipole Dominates Charge Carrier Lifetime in Monolayer Janus MoSSe.

Two-dimensional semiconductor materials with vertical dipoles are promising photocatalysts as vertical dipoles not only promote the electron-hole separation but also enhance the carrier redox ability. However, the influence of vertical dipoles on carrier recombination in such materials, especially the competing relationship between vertical dipoles and band gaps, is not yet clear. Herein, first-principles calculations and nonadiabatic molecular dynamics simulations were combined to clarify the influence of band gap and vertical dipole on the carrier lifetime in Janus MoSSe monolayer. By comparing with the results of MoS2 and MoSe2 as well as exploring the carrier lifetime of MoSSe under strain regulation, it has been demonstrated that the vertical dipole, rather than the band gap, is the dominant factor affecting the carrier lifetime. Strikingly, a linear relationship between the carrier lifetime and vertical dipole is revealed. These findings have important implications for the design of high-performance photocatalysts and optoelectronic devices.

OsNF-YA3 regulates plant growth and osmotic stress tolerance by interacting with SLR1 and SAPK9 in rice.

The antagonism between gibberellin (GA) and abscisic acid (ABA) signaling pathways is vital to balance plant growth and stress response. Nevertheless, the mechanism by which plants determine the balance remains to be elucidated. Here, we report that rice NUCLEAR FACTOR-Y A3 (OsNF-YA3) modulates GA- and ABA-mediated balance between plant growth and osmotic stress tolerance. OsNF-YA3 loss-of-function mutants exhibit stunted growth, compromised GA biosynthetic gene expression, and decreased GA levels, while its overexpression lines have promoted growth and enhanced GA content. Chromatin immunoprecipitation-quantitative polymerase chain reaction analysis and transient transcriptional regulation assays demonstrate that OsNF-YA3 activates GA biosynthetic gene OsGA20ox1 expression. Furthermore, the DELLA protein SLENDER RICE1 (SLR1) physically interacts with OsNF-YA3 and thus inhibits its transcriptional activity. On the other side, OsNF-YA3 negatively regulates plant osmotic stress tolerance by repressing ABA response. OsNF-YA3 reduces ABA levels by transcriptionally regulating ABA catabolic genes OsABA8ox1 and OsABA8ox3 by binding to their promoters. Furthermore, OSMOTIC STRESS/ABA-ACTIVATED PROTEIN KINASE 9 (SAPK9), the positive component in ABA signaling, interacts with OsNF-YA3 and mediates OsNF-YA3 phosphorylation, resulting in its degradation in plants. Collectively, our findings establish OsNF-YA3 as an important transcription factor that positively modulates GA-regulated plant growth and negatively controls ABA-mediated water-deficit and salt tolerance. These findings shed light on the molecular mechanism underlying the balance between the growth and stress response of the plant.

Cognitive enhancing effect of rTMS combined with tDCS in patients with major depressive disorder: a double-blind, randomized, sham-controlled study.

BACKGROUND: Cognitive dysfunction is one of the common symptoms in patients with major depressive disorder (MDD). Repetitive transcranial magnetic stimulation (rTMS) and transcranial direct current stimulation (tDCS) have been studied separately in the treatment of cognitive dysfunction in MDD patients. We aimed to investigate the effectiveness and safety of rTMS combined with tDCS as a new therapy to improve neurocognitive impairment in MDD patients. METHODS: In this brief 2-week, double-blind, randomized, and sham-controlled trial, a total of 550 patients were screened, and 240 MDD inpatients were randomized into four groups (active rTMS + active tDCS, active rTMS + sham tDCS, sham rTMS + active tDCS, sham rTMS + sham tDCS). Finally, 203 patients completed the study and received 10 treatment sessions over a 2-week period. The Repeatable Battery for the Assessment of Neuropsychological Status (RBANS) was performed to assess patients' cognitive function at baseline and week 2. Also, we applied the 24-item Hamilton Depression Rating Scale (HDRS-24) to assess patients' depressive symptoms at baseline and week 2. RESULTS: After 10 sessions of treatment, the rTMS combined with the tDCS group showed more significant improvements in the RBANS total score, immediate memory, and visuospatial/constructional index score (all p < 0.05). Moreover, post hoc tests revealed a significant increase in the RBANS total score and Visuospatial/Constructional in the combined treatment group compared to the other three groups but in the immediate memory, the combined treatment group only showed a better improvement than the sham group. The results also showed the RBANS total score increased significantly higher in the active rTMS group compared with the sham group. However, rTMS or tDCS alone was not superior to the sham group in terms of other cognitive performance. In addition, the rTMS combined with the tDCS group showed a greater reduction in HDRS-24 total score and a better depression response rate than the other three groups. CONCLUSIONS: rTMS combined with tDCS treatment is more effective than any single intervention in treating cognitive dysfunction and depressive symptoms in MDD patients. TRIAL REGISTRATION: Chinese Clinical Trial Registry (ChiCTR2100052122).

A superior heterologous prime-boost vaccination strategy against COVID-19: A bivalent vaccine based on yeast-derived RBD proteins followed by a heterologous vaccine.

Various vaccines have been challenged by SARS-CoV-2 variants. Here, we reported a yeast-derived recombinant bivalent vaccine (Bivalent wild-type [Wt]+De) based on the wt and Delta receptor-binding domain (RBD). Yeast derived RBD proteins based on the wt and Delta mutant were used as the prime vaccine. It was found that, in the presence of aluminium hydroxide (Alum) and unmethylated CpG-oligodeoxynucleotides (CpG) adjuvants, more cross-protective immunity against SARS-CoV-2 prototype and variants were elicited by bivalent vaccine than monovalent wtRBD or Delta RBD. Furthermore, a heterologous boosting strategy consisting of two doses of bivalent vaccines followed by one dose adenovirus vectored vaccine exhibited cross-neutralization capacity and specific T cell responses against Delta and Omicron (BA.1 and BA.4/5) variants in mice, superior to a homologous vaccination strategy. This study suggested that heterologous prime-boost vaccination with yeast-derived bivalent protein vaccine could be a potential approach to address the challenge of emerging variants.

Nuclear factor OsNF-YC5 modulates rice seed germination by regulating synergistic hormone signaling.

Regulation of seed dormancy/germination is of great importance for seedling establishment and crop production. Nuclear factor-Y (NF-Y) transcription factors regulate plant growth and development, as well as stress responses; however, their roles in seed germination remain largely unknown. In this study, we reported that NF-Y gene OsNF-YC5 knockout increased, while its overexpression reduced, the seed germination in rice (Oryza sativa L.). ABA-induced seed germination inhibition assays showed that the osnf-yc5 mutant was less sensitive but OsNF-YC5-overexpressing lines were more sensitive to exogenous ABA than the wild type. Meanwhile, MeJA treatment substantially enhanced the ABA sensitivity of OsNF-YC5-overexpressing lines during seed germination. Mechanistic investigations revealed that the interaction of OSMOTIC STRESS/ABA-ACTIVATED PROTEIN KINASE 9 (SAPK9) with OsNF-YC5 enhanced the stability of OsNF-YC5 by protein phosphorylation, while the interaction between JASMONATE ZIM-domain protein 9 (OsJAZ9) and OsNF-YC5 repressed OsNF-YC5 transcriptional activity and promoted its degradation. Furthermore, OsNF-YC5 transcriptionally activated ABA catabolic gene OsABA8ox3, reducing ABA levels in germinating seeds. However, the transcriptional regulation of OsABA8ox3 by OsNF-YC5 was repressed by addition of OsJAZ9. Notably, OsNF-YC5 improved seed germination under salinity conditions. Further investigation showed that OsNF-YC5 activated the high-affinity K+ transporter gene (OsHAK21) expression, and addition of SAPK9 could increase the transcriptional regulation of OsHAK21 by OsNF-YC5, thus substantially reducing the ROS levels to enhance seed germination under salt stress. Our findings establish that OsNF-YC5 integrates ABA and JA signaling during rice seed germination, shedding light on the molecular networks of ABA-JA synergistic interaction.

Correction: Single-cell in vivo imaging of cellular circadian oscillators in zebrafish.

[This corrects the article DOI: 10.1371/journal.pbio.3000435.].

Engineering Streptomyces sp. CPCC 204095 for the targeted high-level production of isatropolone A by elucidating its pathway-specific regulatory mechanism.

BACKGROUND: Isatropolone A and C, produced by Streptomyces sp. CPCC 204095, belong to an unusual class of non-benzenoid aromatic compounds and contain a rare seven-membered ring structure. Isatropolone A exhibits potent activity against Leishmania donovani, comparable to the only oral drug miltefosine. However, its variably low productivity represents a limitation for this lead compound in the future development of new anti-leishmaniasis drugs to meet unmet clinical needs. RESULTS: Here we first elucidated the regulatory cascade of biosynthesis of isatropolones, which consists of two SARP family regulators, IsaF and IsaJ. Through a series of in vivo and in vitro experiments, IsaF was identified as a pathway-specific activator that orchestrates the transcription of the gene cluster essential for isatropolone biosynthesis. Interestingly, IsaJ was found to only upregulate the expression of the cytochrome P450 monooxygenase IsaS, which is crucial for the yield and proportion of isatropolone A and C. Through targeted gene deletions of isaJ or isaS, we effectively impeded the conversion of isatropolone A to C. Concurrently, the facilitation of isaF overexpression governed by selected promoters, prompted the comprehensive activation of the production of isatropolone A. Furthermore, meticulous optimization of the fermentation parameters was conducted. These strategies culminated in the attainment of an unprecedented maximum yield-980.8 mg/L of isatropolone A-achieved in small-scale solid-state fermentation utilizing the genetically modified strains, thereby establishing the highest reported titer to date. CONCLUSION: In Streptomyces sp. CPCC 204095, the production of isatropolone A and C is modulated by the SARP regulators IsaF and IsaJ. IsaF serves as a master pathway-specific regulator for the production of isatropolones. IsaJ, on the other hand, only dictates the transcription of IsaS, the enzyme responsible for the conversion of isatropolone A and C. By engineering the expression of these pivotal genes, we have devised a strategy for genetic modification aimed at the selective and high-yield biosynthesis of isatropolone A. This study not only unveils the unique regulatory mechanisms governing isatropolone biosynthesis for the first time, but also establishes an essential engineering framework for the targeted high-level production of isatropolone A.

Tailoring the ion storage of MXene by aramid nanofibers towards self-standing electrodes for flexible solid-state supercapacitors.

Two-dimensional (2D) transition metal carbides and nitrides (MXenes) are a class of 2D nanomaterials that can offer excellent properties for high-performance supercapacitors. Nevertheless, irreversible restacking of MXene sheets decreases the interlayer spacing, which inhibits the ion intercalation between the MXene nanosheets and finally deteriorates the electrochemical performance of supercapacitors. Herein, aramid nanofibers (ANFs) are mixed with Ti3C2TxMXene to prepare MXene/ANFs composite films. The restacking of MXene sheets is inhibited by the electrostatic repulsion between ANFs and MXene. The ANFs act as intercalation agents to increase the interlayer spacing of the composite films, which can improve the ion storage ability of supercapacitors. Furthermore, the ANFs enhance the mechanical strength of the composite films due to the strong hydrogen bonding interaction and nanomechanical interlocking between ANFs and MXene, endowing the composite films with self-standing property. The resultant composite films are used as electrodes for flexible solid-state supercapacitors to achieve high specific capacitance (996.5 mF cm-2at 5 mV s-1) and outstanding cycling stability. Thus, this work provides a potential strategy to regulate the properties of 2D nanomaterials, which may expand the application of them in energy storage, ionic separation, osmotic energy conversion and beyond.

Synthesis and Application of Specific N2H4 Fluorescent Probes with AIE Effect Based on Pyrazole Structure.

Two fluorescent probes, Y1-2 were synthesized from 2-acetonaphthone, 4-acetylbiphenyl, and phenyl hydrazine by Vilsmeier-Haack reaction and Knoevenagel condensation. Their recognition efficacies for N2H4 were tested by UV-visible absorption spectroscopy and fluorescence emission spectroscopy. The recognition mechanism were studies by density-functional theory calculations, and the effect of pH on N2H4 recognition was also studied. The results showed that the probe Y1-2 has high selectivity and a low detection limit for N2H4, and the recognition of N2H4 can be accomplished at physiological pH. The probes have had obvious aggregation-induced luminescence effect, large Stokes shift, high sensitivity, and can be successfully applied to live cell imaging.

OsMADS23 phosphorylated by SAPK9 confers drought and salt tolerance by regulating ABA biosynthesis in rice.

Some of MADS-box transcription factors (TFs) have been shown to play essential roles in the adaptation of plant to abiotic stress. Still, the mechanisms that MADS-box proteins regulate plant stress response are not fully understood. Here, a stress-responsive MADS-box TF OsMADS23 from rice conferring the osmotic stress tolerance in plants is reported. Overexpression of OsMADS23 remarkably enhanced, but knockout of the gene greatly reduced the drought and salt tolerance in rice plants. Further, OsMADS23 was shown to promote the biosynthesis of endogenous ABA and proline by activating the transcription of target genes OsNCED2, OsNCED3, OsNCED4 and OsP5CR that are key components for ABA and proline biosynthesis, respectively. Then, the convincing evidence showed that the OsNCED2-knockout mutants had lower ABA levels and exhibited higher sensitivity to drought and oxidative stress than wild type, which is similar to osmads23 mutant. Interestingly, the SnRK2-type protein kinase SAPK9 was found to physically interact with and phosphorylate OsMADS23, and thus increase its stability and transcriptional activity. Furthermore, the activation of OsMADS23 by SAPK9-mediated phosphorylation is dependent on ABA in plants. Collectively, these findings establish a mechanism that OsMADS23 functions as a positive regulator in response to osmotic stress by regulating ABA biosynthesis, and provide a new strategy for improving drought and salt tolerance in rice.

Chemically Controlled Reversible Magnetic Phase Transition in Two-Dimensional Organometallic Lattices.

Developing an efficient method to reversibly control materials' spin order is urgently needed but challenging in spintronics. Though various physical field control methods have been advancing, the chemical control of spin is little exploited. Here, we propose a chemical means for such spin manipulation, i.e., utilizing the well-known lactim-lactam tautomerization to reversibly modulate the magnetic phase transition in two-dimensional (2D) organometallic lattices. The proposal is verified by theoretically designing several 2D organometallic frameworks with antiferromagnetic to ferrimagnetic spin order transformation modulated by lactim-lactam tautomerization on organic linkers. The transition originates from the change in spin states of organic linkers (from singlet to doublet) via tautomerization. Such a transition further switches materials' electronic structures from normal semiconductors with zero spin polarization to bipolar magnetic semiconductors with valence and conduction band edges 100% spin polarized in opposite spin channels. Moreover, the magnitude of magnetic anisotropy energy also enhances by 5- to 9-fold.

Gate-Tunable Spin Seebeck Effect and Pure Spin Current Generation in Molecular Junctions Based on Bipolar Magnetic Molecules.

Generating pure spin currents is very desirable in spintronics, as it provides a promising way to substantially reduce Joule heating and achieve ultrahigh integration density. However, to date, most spintronic devices exhibit spin currents that are accompanied by charge currents. The generation of pure spin currents on the nanoscale, particularly at the single-molecule level, remains challenging. Here, we propose that by exploiting our recently reported bipolar magnetic molecules (BMMs) as the core component of single-molecule devices, where the highest occupied molecular orbital (HOMO) and lowest unoccupied molecular orbital (LUMO) come from different spin channels, the generation of pure spin currents can be easily realized via the spin Seebeck effect (SSE) with applied temperature gradient. Moreover, the spin Seebeck coefficient can be modulated over a wide range by applying an external gate voltage. The proposal is verified through first-principles calculations on two BMM-based molecular junctions.

Quintuple Function Integration in Two-Dimensional Cr(II) Five-Membered Heterocyclic Metal Organic Frameworks via Tuning Ligand Spin and Lattice Symmetry.

Two-dimensional (2D) semiconductors (SCs) integrated with two or more functions are the cornerstone for constructing multifunctional nanodevices but remain largely limited. Here, by tuning the spin state of organic linkers and the symmetry/topology of crystal lattices, we predict a class of unprecedented multifunctional SCs in 2D Cr(II) five-membered heterocyclic metal organic frameworks that simultaneously possess auxetic effect, room-temperature ferrimagnetism, chiral ferroelectricity (FE), electrically reversible spin polarization, and topological nodal lines/points. Taking 2D Cr(TDZ)2 (TDZ = 1.2.5-thiadiazole) as an exemplification, the auxetic effect is produced by the antitetra-chiral lattice structure. The high temperature ferrimagnetism originates from the strong d-p direct magnetic exchange interaction between Cr cations and TDZ doublet radical anions. Meanwhile, the clockwise-counterclockwise alignment of TDZ's dipoles results in unique 2D chiral FE with atomic-scale vortex-antivortex states. 2D Cr(TDZ)2 is an intrinsic bipolar magnetic SC where half-metallic conduction with switchable spin-polarization direction can be induced by applying a gate voltage. In addition, the symmetry of the little group C4 of the lattice structure endows 2D Cr(TDZ)2 with topological nodal lines and a quadratic nodal point in the Brillouin zone near the Fermi level.

Frizzled-7-targeting antibody (SHH002-hu1) potently suppresses non-small-cell lung cancer via Wnt/ß-catenin signaling.

Non-small-cell lung cancer (NSCLC) is one of the deadliest cancers worldwide, and metastasis is considered one of the leading causes of treatment failure in NSCLC. Wnt/ß-catenin signaling is crucially involved in epithelial-mesenchymal transition (EMT), a crucial factor in promoting metastasis, and also contributes to resistance developed by NSCLC to targeted agents. Frizzled-7 (Fzd7), a critical receptor of Wnt/ß-catenin signaling, is aberrantly expressed in NSCLC and has been confirmed to be positively correlated with poor clinical outcomes. SHH002-hu1, a humanized antibody targeting Fzd7, was previously successfully generated by our group. Here, we studied the anti-tumor effects of SHH002-hu1 against NSCLC and revealed the underlying mechanism. First, immunofluorescence (IF) and near-infrared (NIR) imaging assays showed that SHH002-hu1 specifically binds Fzd7+ NSCLC cells and targets NSCLC tissues. Wound healing and transwell invasion assays indicated that SHH002-hu1 significantly inhibits the migration and invasion of NSCLC cells. Subsequently, TOP-FLASH/FOP-FLASH luciferase reporter, IF, and western blot assays validated that SHH002-hu1 effectively suppresses the activation of Wnt/ß-catenin signaling, and further attenuates the EMT of NSCLC cells. Finally, the subcutaneous xenotransplanted tumor model of A549/H1975, as well as the popliteal lymph node (LN) metastasis model, was established, and SHH002-hu1 was demonstrated to inhibit the growth of NSCLC xenografts and suppress LN metastasis of NSCLC. Above all, SHH002-hu1 with selectivity toward Fzd7+ NSCLC and the potential of inhibiting invasion and metastasis of NSCLC via disrupting Wnt/ß-catenin signaling, is indicated as a good candidate for the targeted therapy of NSCLC.

Structural and Interfacial Manipulation of Multifunctional Skeletons Enabled Shuttling-Free and Dendrite-Free Li-S Full Batteries.

Li-S batteries are regarded as promising devices for energy storage systems owing to high energy density, low cost, and environmental friendliness. However, challenges of polysulfides shuttling in sulfur cathode and dendrite growth of lithium anode severely hinder the practical application. Developing advanced skeletons simultaneously regulating the cathode and anode is significant and challenging. Hence, a novel and scalable strategy combining spray drying and topological nitriding is proposed, and hierarchically assembled rGO hollow microspheres encapsulated highly porous nanospheres consisted of ultrafine Nb4 N5 -Nb2 O5 or Nb4 N5 nanoparticles as multifunctional skeletons for S and Li are designed. In such unique architecture, a 3D highly porous structure provides abundant void space for loading of S and Li, and accommodates volume change during cycling. Moreover, Nb4 N5 -Nb2 O5 heterostructured interface promotes adsorption-conversion process of polysulfides, while strong lithophilic Nb4 N5 induces the selective infiltration of Li into the void of the skeleton and regulates the uniform deposition and growth. More interestingly, in situ generated Li3 N@Nb ion/electron conducting interfaces induced by the reaction of Nb4 N5 and Li reduce the nucleation overpotential and induce selective deposition of Li into the cavity. Consequently, the Li-S full cell exhibits superior cycling stability and impressive rate performance with a low capacity ratio of negative/positive.

Cellulose synthesis complexes are homo-oligomeric and hetero-oligomeric in Physcomitrium patens.

The common ancestor of seed plants and mosses contained homo-oligomeric cellulose synthesis complexes (CSCs) composed of identical subunits encoded by a single CELLULOSE SYNTHASE (CESA) gene. Seed plants use different CESA isoforms for primary and secondary cell wall deposition. Both primary and secondary CESAs form hetero-oligomeric CSCs that assemble and function in planta only when all the required isoforms are present. The moss Physcomitrium (Physcomitrella) patens has seven CESA genes that can be grouped into two functionally and phylogenetically distinct classes. Previously, we showed that PpCESA3 and/or PpCESA8 (class A) together with PpCESA6 and/or PpCESA7 (class B) form obligate hetero-oligomeric complexes required for normal secondary cell wall deposition. Here, we show that gametophore morphogenesis requires a member of class A, PpCESA5, and is sustained in the absence of other PpCESA isoforms. PpCESA5 also differs from the other class A PpCESAs as it is able to self-interact and does not co-immunoprecipitate with other PpCESA isoforms. These results are consistent with the hypothesis that homo-oligomeric CSCs containing only PpCESA5 subunits synthesize cellulose required for gametophore morphogenesis. Analysis of mutant phenotypes also revealed that, like secondary cell wall deposition, normal protonemal tip growth requires class B isoforms (PpCESA4 or PpCESA10), along with a class A partner (PpCESA3, PpCESA5, or PpCESA8). Thus, P. patens contains both homo-oligomeric and hetero-oligomeric CSCs.

Single-cell in vivo imaging of cellular circadian oscillators in zebrafish.

The circadian clock is a cell-autonomous time-keeping mechanism established gradually during embryonic development. Here, we generated a transgenic zebrafish line carrying a destabilized fluorescent protein driven by the promoter of a core clock gene, nr1d1, to report in vivo circadian rhythm at the single-cell level. By time-lapse imaging of this fish line and 3D reconstruction, we observed the sequential initiation of the reporter expression starting at photoreceptors in the pineal gland, then spreading to the cells in other brain regions at the single-cell level. Even within the pineal gland, we found heterogeneous onset of nr1d1 expression, in which each cell undergoes circadian oscillation superimposed over a cell type-specific developmental trajectory. Furthermore, we found that single-cell expression of nr1d1 showed synchronous circadian oscillation under a light-dark (LD) cycle. Remarkably, single-cell oscillations were dramatically dampened rather than desynchronized in animals raised under constant darkness, while the developmental trend still persists. It suggests that light exposure in early zebrafish embryos has significant effect on cellular circadian oscillations.