Organisms respond to tissue damage through the upregulation of protective responses which restore tissue structure and metabolic function. Mitochondria are key sources of intracellular oxidative ...metabolic signals that maintain cellular homeostasis. Here we report that tissue and cellular wounding triggers rapid and reversible mitochondrial fragmentation. Elevated mitochondrial fragmentation either in fzo-1 fusion-defective mutants or after acute drug treatment accelerates actin-based wound closure. Wounding triggered mitochondrial fragmentation is independent of the GTPase DRP-1 but acts via the mitochondrial Rho GTPase MIRO-1 and cytosolic Ca
. The fragmented mitochondria and accelerated wound closure of fzo-1 mutants are dependent on MIRO-1 function. Genetic and transcriptomic analyzes show that enhanced mitochondrial fragmentation accelerates wound closure via the upregulation of mtROS and Cytochrome P450. Our results reveal how mitochondrial dynamics respond to cellular and tissue injury and promote tissue repair.
Membrane repair is essential for cell and organism survival. Exocytosis and endocytosis facilitate membrane repair in small wounds within a single cell; however, it remains unclear how large wounds ...in the plasma membrane are repaired in metazoans. Here, we show that wounding triggers rapid transcriptional upregulation and dynamic recruitment of the fusogen EFF-1 to the wound site in C. elegans epidermal cells. EFF-1 recruitment at the wounded membrane depends on the actin cytoskeleton and is important for membrane repair. We identified syntaxin-2 (SYX-2) as an essential regulator of EFF-1 recruitment. SYX-2 interacts with the C terminus of EFF-1 to promote its recruitment, facilitating both endoplasmic and exoplasmic membrane repair. Furthermore, we show that SYX-2-EFF-1 repair machinery acts downstream of the ESCRT III signal. Together, our findings identify a key pathway underlying membrane repair and provide insights into tissue repair and regenerative medicine after injury.
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•The fusogen EFF-1 is required for membrane repair and animal survival•EFF-1 recruitment to the wounded membrane depends on actin polymerization•Wounding-induced SYX-2 promotes EFF-1 recruitment•SYX-2 interacts with EFF-1 and acts downstream of ESCRT III during membrane repair
The ability to repair plasma membrane damage is critical to all eukaryotic cells. Meng et al. demonstrate that damage-induced F-actin and ESCRT III signals trigger sequential recruitment of the fusogens syntaxin-2 and EFF-1 to facilitate repair of the plasma membrane in C. elegans skin cells.
Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is currently a global pandemic. CoVs are known to generate negative subgenomes (subgenomic ...RNAs sgRNAs) through transcription-regulating sequence (TRS)-dependent template switching, but the global dynamic landscapes of coronaviral subgenomes and regulatory rules remain unclear. Here, using next-generation sequencing (NGS) short-read and Nanopore long-read poly(A) RNA sequencing in two cell types at multiple time points after infection with SARS-CoV-2, we identified hundreds of template switches and constructed the dynamic landscapes of SARS-CoV-2 subgenomes. Interestingly, template switching could occur in a bidirectional manner, with diverse SARS-CoV-2 subgenomes generated from successive template-switching events. The majority of template switches result from RNA-RNA interactions, including seed and compensatory modes, with terminal pairing status as a key determinant. Two TRS-independent template switch modes are also responsible for subgenome biogenesis. Our findings reveal the subgenome landscape of SARS-CoV-2 and its regulatory features, providing a molecular basis for understanding subgenome biogenesis and developing novel anti-viral strategies.
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•Dynamic subgenome landscapes of SARS-CoV-2 in two host cells are constructed•Bidirectional and successive template switching diversify sgRNA biogenesis•Several key determinants governing template switching efficacy are discovered•Canonical TRS-independent RNA-RNA interaction mediates template switches
Wang et al. construct dynamic landscapes of SARS-CoV-2 subgenomic RNAs (sgRNAs) by using integrated poly(A) RNA sequencing at various time points after infection. Computational analyses reveal novel modes of viral sgRNA biogenesis and decode regulatory features, including several key determinants governing the efficacy of template switching in coronaviral RNA transcription.
Postmeiotic spermatids use a unique strategy to coordinate gene expression with morphological transformation, in which transcription and translation take place at separate developmental stages, but ...how mRNAs stored as translationally inert messenger ribonucleoproteins in developing spermatids become activated remains largely unknown. Here, we report that the RNA binding protein FXR1, a member of the fragile X–related (FXR) family, is highly expressed in late spermatids and undergoes liquid-liquid phase separation (LLPS) to merge messenger ribonucleoprotein granules with the translation machinery to convert stored mRNAs into a translationally activated state. Germline-specific
Fxr1
ablation in mice impaired the translation of target mRNAs and caused defective spermatid development and male infertility, and a phase separation–deficient FXR1
L351P
mutation in
Fxr1
knock-in mice produced the same developmental defect. These findings uncover a mechanism for translational reprogramming with LLPS as a key driver in spermiogenesis.
Driving translation in spermatids
Translation control becomes a key mechanism for gene regulation during postmeiotic male germ cell differentiation because of the uncoupling between transcription and translation. Kang
et al
. identified a member of the fragile X-related (FXR) protein family, FXR1, as a translation activator for stored mRNAs to instruct spermiogenesis in mice (see the Perspective by Ramat and Simonelig). Mechanistically, highly elevated FXR1 in late spermatids forms condensates through phase separation to organize the stored mRNAs into FXR1 granules, and then recruits the translational machinery to activate translation. Either germline-specific depletion of Fxr1 or modeling a phase separation–deficient mutation in Fxr1 similarly causes spermatogenic failure and male infertility in mice. —DJ
Phase separation of the RNA-binding protein FXR1 links translation activation to spermatid development in mice.
INTRODUCTION
In mammals, spermiogenesis (postmeiotic male germ cell differentiation) is a highly orchestrated developmental process controlled by a group of genes collectively referred to as spermiogenic genes. Because nuclear condensation during spermiogenesis gradually halts transcription, spermiogenic genes are transcribed in advance during the earlier stages of male germ development and stored as translationally inert messenger ribonucleoproteins (mRNPs) in developing spermatids until they are needed for translation. Such inert mRNPs are usually organized into mRNP granules called germ granules, which serve as storage facilities for nontranslating mRNAs in various types of germ cells. However, little is known about how those mRNAs stored in inert mRNPs are activated during late spermiogenesis.
RATIONALE
To understand how translationally inert mRNAs are activated during spermiogenesis, we screened potential translational regulators by proteomic analysis of polysomes from mouse testes. FXR1, a member of the fragile X–related (FXR) protein family, stood out from the screen as a translational regulator in late spermatids. By performing eCLIP and polysome profiling, in combination with generating a germline-specific
Fxr1
knockout (
Fxr1
cko
) mouse model, we investigated whether FXR1 is required for translation activation in late spermatids. To decipher the mechanism underlying FXR1-mediated translation regulation, we identified the potential cofactor(s) of FXR1 in mouse testes using immunoprecipitation coupled with mass spectrometry. We observed the formation of FXR1 granules through liquid-liquid phase separation (LLPS), which recruits translation factors in late spermatids, and used the TRICK (translating RNA imaging by coat protein knock-off) reporter system to determine whether FXR1 LLPS is required for target translation in cultured cells. To further investigate whether FXR1 LLPS is critical for target translation in mouse spermatids, we ectopically expressed wild-type FXR1, LLPS-deficient FXR1
L351P
mutants, or LLPS-restored FXR1
L351P
-IDR
FUS
mutants in
Fxr1
cko
testes using lentiviral testis transduction. Finally, by generating germline-specific
Fxr1
L351P
knock-in mice, we determined whether FXR1 LLPS is indispensable to translation activation in late spermatids, spermiogenesis, and male fertility in mice.
RESULTS
We found that FXR1 was much more enriched in polysomes from 35-day postpartum (dpp) testes relative to 25-dpp testes, suggesting a role for FXR1 in translation activation in late spermatids. We identified a group of 770 mRNAs as being likely direct FXR1-activated targets, and demonstrated that germline-specific
Fxr1
deletion in mice markedly reduced target translation in late spermatids. Consistent with FXR1 functioning in translation activation in late spermatids,
Fxr1
cko
male mice were infertile and displayed spermatogenic failure at late spermiogenesis. Interestingly, we observed a pronounced up-regulation of FXR1 and the formation of abundant, distinct condensates in late spermatids, suggesting concentration-dependent LLPS. Mechanistic studies revealed that FXR1 undergoes LLPS to form condensates that assemble target mRNAs as mRNP granules and then recruit translational machinery to activate the stored mRNAs. Consistently, ectopic expression of wild-type FXR1 or FXR1
L351P
-IDR
FUS
, but not FXR1
L351P
, activated target translation in cultured cells and successfully rescued target translation in late spermatids and spermiogenesis in
Fxr1
cko
mice. Furthermore,
Fxr1
L351P
knock-in mutant mice highly phenocopy
Fxr1
cko
mice, directly supporting the indispensability of FXR1 LLPS to target translation in late spermatids, spermiogenesis, and male fertility in mice.
CONCLUSION
Our findings demonstrate that FXR1 is an essential translation activator that instructs spermiogenesis in mice and unveil a key contribution of FXR1 LLPS to the translation activation of stored mRNAs in mouse spermatid and male fertility in mice. In addition, our study pinpoints the importance of LLPS in a developmental process in vivo.
FXR1-containing granules mediate translation activation in late spermatids.
During late spermiogenesis, elevated FXR1 undergoes LLPS to assemble target mRNAs as FXR1 mRNP granules that recruit translational machinery by interacting with the eukaryotic translation initiation factor 4 gamma 3 (EIF4G3) to activate the stored mRNAs in late spermatids. These phase-separated FXR1 granules drive a large translation program to instruct spermatid development and sperm production in mice.
In this investigation, a novel water-insoluble slow-release fertilizer, biuret polyphosphoramide (BPAM), was formulated and synthesized from urea, phosphoric acid (H3PO4), and ferric oxide (Fe2O3). ...The structure of BPAM was characterized by Fourier transform infrared (FTIR) spectroscopy. Subsequently, a coated slow-release BPAM fertilizer with superabsorbent was prepared by ionic cross-linked carboxymethylchitosan (the core), acrylic acid, acrylamide, and active carbon (the coating). The variable influences on the water absorbency were investigated and optimized. Component analysis results showed that the coated slow-release BPAM contained 5.66% nitrogen and 11.7% phosphorus. The property of water retention, the behavior of slow release of phosphorus, and the capacity of adsorption of cations were evaluated, and the results revealed that the product not only had good slow-release property and excellent water retention capacity but also higher adsorption capacities of cations in saline soil.
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•A novel electrical-sensitive poly (NVP-co-AA) hydrogel was prepared from linear copolymer.•Gelation mechanism was studied by FTIR and solid-state CP/MAS NMR techniques.•Swelling and ...bending phenomenon of the hydrogel were investigated under an electric field.•The bending mechanism was reinterpreted.
On the basis of the existing knowledge of reactions and properties of poly (N-vinylpyrrolidone) (PVP) and poly (acrylic acid) (PAA), a novel pH- and electrical-sensitive poly (N-vinylpyrrolidone-co-acrylic acid) poly (NVP-co-AA) hydrogel was formulated and prepared from linear poly (NVP-co-AA). Gelation reaction was initiated by potassium persulfate (KPS) used as a radical initiator, and the mechanism of crosslinking reaction was investigated by FTIR and solid-state CP/MAS NMR techniques. The morphology of hydrogel with flexible chain nature and large free volume was verified by measurement of scanning electron microscope. The swelling behavior and bending phenomenon of the hydrogel were investigated. In the electric field, the gel bent toward cathode independently of the pH of buffer solution, applied voltage, crosslinking extent and polyion content in the backbone of gel. But the deflection and speed of the bending depended on them. Bending of poly (NVP-co-AA) gel was reinterpreted by bending theory of polyelectrolyte gel based upon the change of osmotic pressure, which is due to the difference of ion concentration between the sides facing anode and cathode.
Wounding initiates intricate responses crucial for tissue repair and regeneration. Yet, the gene regulatory networks governing wound healing remain poorly understood. Here, employing single-worm RNA ...sequencing (swRNA-seq) across 12 time-points, we delineated a three-stage wound repair process in C. elegans: response, repair, and remodeling. Integrating diverse datasets, we constructed a dynamic regulatory network comprising 241 transcription regulators and their inferred targets. We identified potentially seven autoregulatory TFs and five cross-autoregulatory loops involving pqm-1 and jun-1. We revealed that TFs might interact with chromatin factors and form TF-TF combinatory modules via intrinsically disordered regions to enhance response robustness. We experimentally validated six regulators functioning in transcriptional and translocation-dependent manners. Notably, nhr-76, daf-16, nhr-84, and oef-1 are potentially required for efficient repair, while elt-2 may act as an inhibitor. These findings elucidate transcriptional responses and hierarchical regulatory networks during C. elegans wound repair, shedding light on mechanisms underlying tissue repair and regeneration.Time-course swRNA-seq in C. elegans reveals a three-stage process in wound repair potentially controlled by a complex gene regulatory network containing autoregulatory loops and more than 200 TFs, such as crucial factors nhr-76 and daf-16.
Polysome profiling is widely used to isolate and analyze polysome fractions, which consist of actively translating mRNAs and ribosomes. Compared to ribosome profiling and translating ribosome ...affinity purification, polysome profiling is simpler and less time consuming in sample preparation and library constructions. Spermiogenesis, i.e., the post-meiotic phase of male germ cell development, is a highly coordinated developmental process in which transcription and translation are decoupled because of nuclear condensation, resulting in translation regulation as the major mode for the regulation of gene expression in post-meiotic spermatids. To understand the translation regulation during spermiogenesis, an overview of translational state of spermiogenic mRNAs is required. Here, we describe a protocol to identify translating mRNAs using polysome profiling. Briefly, mouse testes are gently homogenized to release polysomes containing translating mRNAs, following polysome-bound mRNAs isolated by sucrose density gradient purification and characterized by RNA-seq. This protocol allows to quickly isolate translating mRNAs from testes and analyze the discrepancy of translational efficiency in mouse testes from different mouse lines. Key features Quickly obtain polysome RNAs from testes. Omit RNase digestion and RNA recovery from gel. High efficiency and robustness compared to ribo-seq. Graphical overview
Mouse testes are homogenized and lysed in
, and polysome RNAs are enriched by sucrose gradient centrifugation and used to calculate translation efficiency in
.
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•Responsive composite materials of aligned carbon nanotubes are summarized.•The composite materials exhibit rapid and reversible deformation.•The composite materials exhibit rapid and ...reversible chromatic.
Responsive materials have been investigated for a lot of applications such as smart devices and sensors, which change shape, color, transparency, or functionality when stimulated by external stimuli. However, the practical applications of responsive materials have been largely hampered by their irreversible transitions under limited stimuli such as temperature, pH, and chemical. As a result, a lot of research has been paid to improve the reversibility of chromatism and deformation. In this review, the recent research of responsive materials based on aligned carbon nanotubes is described, with the focus on the responsive materials in response to light and electricity.
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