The miR159a-DUO1 module regulates pollen development by modulating auxin biosynthesis and starch metabolism in citrus.

Achieving seedlessness in citrus varieties is one of the important objectives of citrus breeding. Male sterility associated with abnormal pollen development is an important factor in seedlessness. However, our understanding of the regulatory mechanism underlying the seedlessness phenotype in citrus is still limited. Here, we determined that the miR159a-DUO1 module played an important role in regulating pollen development in citrus, which further indirectly modulated seed development and fruit size. Both the overexpression of csi-miR159a and the knocking out of DUO1 in Hong Kong kumquat (Fortunella hindsii) resulted in small and seedless fruit phenotypes. Moreover, pollen was severely aborted in both transgenic lines, with arrested pollen mitotic I and abnormal pollen starch metabolism. Through additional cross-pollination experiments, DUO1 was proven to be the key target gene for miR159a to regulate male sterility in citrus. Based on DNA affinity purification sequencing (DAP-seq), RNA-seq, and verified interaction assays, YUC2/YUC6, SS4 and STP8 were identified as downstream target genes of DUO1, those were all positively regulated by DUO1. In transgenic F. hindsii lines, the miR159a-DUO1 module down-regulated the expression of YUC2/YUC6, which decreased indoleacetic acid (IAA) levels and modulated auxin signaling to repress pollen mitotic I. The miR159a-DUO1 module reduced the expression of the starch synthesis gene SS4 and sugar transport gene STP8 to disrupt starch metabolism in pollen. Overall, this work reveals a new mechanism by which the miR159a-DUO1 module regulates pollen development and elucidates the molecular regulatory network underlying male sterility in citrus.

Nuclear KRT19 is a transcriptional corepressor promoting histone deacetylation and liver tumorigenesis.

BACKGROUND AND AIMS: Epigenetic reprogramming and escape from terminal differentiation are poorly understood enabling characteristics of liver cancer. Keratin 19 (KRT19), classically known to form the intermediate filament cytoskeleton, is a marker of stemness and worse prognosis in liver cancer. This study aimed to address the functional roles of KRT19 in liver tumorigenesis and to elucidate the underlying mechanisms. APPROACH AND RESULTS: Using multiplexed genome editing of hepatocytes in vivo, we demonstrated that KRT19 promoted liver tumorigenesis in mice. Cell fractionation revealed a previously unrecognized nuclear fraction of KRT19. Tandem affinity purification identified histone deacetylase 1 and REST corepressor 1, components of the corepressor of RE-1 silencing transcription factor (CoREST) complex as KRT19-interacting proteins. KRT19 knockout markedly enhanced histone acetylation levels. Mechanistically, KRT19 promotes CoREST complex formation by enhancing histone deacetylase 1 and REST corepressor 1 interaction, thus increasing the deacetylase activity. ChIP-seq revealed hepatocyte-specific genes, such as hepatocyte nuclear factor 4 alpha ( HNF4A ), as direct targets of KRT19-CoREST. In addition, we identified forkhead box P4 as a direct activator of aberrant KRT19 expression in liver cancer. Furthermore, treatment of primary liver tumors and patient-derived xenografts in mice suggest that KRT19 expression has the potential to predict response to histone deacetylase 1 inhibitors especially in combination with lenvatinib. CONCLUSIONS: Our data show that nuclear KRT19 acts as a transcriptional corepressor through promoting the deacetylase activity of the CoREST complex, resulting in dedifferentiation of liver cancer. These findings reveal a previously unrecognized function of KRT19 in directly shaping the epigenetic landscape in cancer.

Structural insights into histone exchange by human SRCAP complex.

Histone variant H2A.Z is found at promoters and regulates transcription. The ATP-dependent chromatin remodeler SRCAP complex (SRCAP-C) promotes the replacement of canonical histone H2A-H2B dimer with H2A.Z-H2B dimer. Here, we determined structures of human SRCAP-C bound to H2A-containing nucleosome at near-atomic resolution. The SRCAP subunit integrates a 6-subunit actin-related protein (ARP) module and an ATPase-containing motor module. The ATPase-associated ARP module encircles half of the nucleosome along the DNA and may restrain net DNA translocation, a unique feature of SRCAP-C. The motor module adopts distinct nucleosome binding modes in the apo (nucleotide-free), ADP-bound, and ADP-BeFx-bound states, suggesting that ATPase-driven movement destabilizes H2A-H2B by unwrapping the entry DNA and pulls H2A-H2B out of nucleosome through the ZNHIT1 subunit. Structure-guided chromatin immunoprecipitation sequencing analysis confirmed the requirement of H2A-contacting ZNHIT1 in maintaining H2A.Z occupancy on the genome. Our study provides structural insights into the mechanism of H2A-H2A.Z exchange mediated by SRCAP-C.

Association between NTCP hepatic expression and inflammation/fibrosis as well as gender-specific differences in chronic HBV-infected patients.

To investigate the relationship between the expression of hepatitis B virus (HBV) functional receptor sodium taurocholate cotransporting polypeptide (NTCP) with disease progression and gender-specific differences in chronic HBV-infected patients. Liver samples were collected from chronic HBV-infected patients who underwent percutaneous liver biopsy or liver surgery. HBV DNA levels and the mRNA and protein expression levels of NTCP in liver tissues were determined. The relationship between NTCP expression and HBV DNA levels, inflammatory activity, fibrosis, and gender-specific differences were analyzed. A total of 94 chronic HBV-infected patients were included. Compared with patients with a METAVIR score of A0-1 or F0-1, patients with score of A2 or F2/F3 had a relatively higher level of NTCP expression. NTCP levels were positively correlated with HBV DNA levels. The inflammatory activity scores and fibrosis scores of women <50 years were significantly lower than those of women ≥50 years and age-matched males. In patients with score A0-2 or F0-3, women <50 years have lower NTCP expression level compared to women ≥50 years and age-matched males. NTCP can promote the disease progression by affecting the viral load of HBV. The NTCP expression difference may be why male and postmenopausal women are more prone to disease progression than reproductive women.

Interplay between the transcription preinitiation complex and the +1 nucleosome.

Eukaryotic transcription starts with the assembly of a preinitiation complex (PIC) on core promoters. Flanking this region is the +1 nucleosome, the first nucleosome downstream of the core promoter. While this nucleosome is rich in epigenetic marks and plays a key role in transcription regulation, how the +1 nucleosome interacts with the transcription machinery has been a long-standing question. Here, we summarize recent structural and functional studies of the +1 nucleosome in complex with the PIC. We specifically focus on how differently organized promoter-nucleosome templates affect the assembly of the PIC and PIC-Mediator on chromatin and result in distinct transcription initiation.

Ascorbic Acid Reprograms Epigenome and Epitranscriptome by Reducing Fe(III) in the Catalytic Cycle of Dioxygenases.

Ascorbic acid (ASC) has been reported to stimulate DNA iterative oxidase ten-eleven translocation (TET) enzymes, Jumonji C-domain-containing histone demethylases, and potentially RNA m6A demethylases FTO and ALKBH5 as a cofactor. Although ascorbic acid has been widely investigated in reprogramming DNA and histone methylation status in vitro, in cultured cells and mouse models, its specific role in the catalytic cycle of dioxygenases remains enigmatic. Here, we systematically investigated the stimulation of ASC toward TET2, ALKBH3, histone demethylases, and FTO. We find that ASC reprograms epitranscriptome by erasing the hypermethylated m6A sites in mRNA. Biochemistry and electron spin resonance assays demonstrate that ASC enters the active pocket of dioxygenases and reduces Fe(III), either incorporated upon protein synthesis or generated upon rebounding the hydroxyl radical during oxidation, into Fe(II). Finally, we propose a remedied model for the catalytic cycle of dioxygenases by adding in the essential cofactor, ASC, which refreshes and regenerates inactive dioxygenase through recycling Fe(III) into Fe(II) in a dynamic "hit-and-run" manner.

Effect of vane angle on turbulence-chemistry interactions, temperature, flow field, burnout, and NOx emissions in swirling flames fueled by semicoke mixtures.

To enhance the combustion efficiency and reduce NOx emissions in large-scale semicoke and bituminous coal blends, an extensive numerical study was conducted. The focus of this study was to optimize the quaternary air vane angle (αv) through detailed analysis of the temperature and flow fields, turbulence-chemistry interactions, char burnout, and NOx formation in a carefully scaled 1:5 dual-swirl burner. The results showed that with increasing αv, the high-temperature flame region was narrowed and the peak temperature was reduced along with the broadened inner recirculation zone and the shrunken external recirculation zone due to better pulverized fuel-oxidant blending and reinforced convective heat transfer. The peak turbulent Damköhler number Dat evidently increased from 197.5 to 496 with increasing αv, which implied a strengthened homogeneous combustion. Additionally, the corresponding mixing time scales increased while the chemical kinetics time scales decreased, which denoted that an intense diffusing flame was generated with a strong turbulent intensity. The peak heterogeneous Damköhler number Das-O2 showed a reduction from 2.54 to 2.27, while the peak values of Das-CO2 and Das-H2O decreased from 0.1 to 0.077 and from 0.02 to 0.015, respectively. The char-O2 reaction was controlled by diffusion/kinetics; both char-CO2 and char-H2O reactions were determined by kinetics, and all gas‒solid reactions showed a kinetically controlled regime. With increasing αv, the enlarged inner recirculation region increased the residence time, and a higher dilution level lessened the peak temperature, which led to reductions in fuel-NOx and the thermal-NOx. The αv range of 30-45° (or swirl number Sn = 0.55-0.95) was suggested by taking the high burnout and low-NOx formation into account.

Perioperative outcomes of bi-pigtail catheter drainage strategy versus conventional chest tube after uniportal video-assisted thoracic lung surgery.

OBJECTIVES: Chest tube (CT) drainage is a main cause of postoperative pain in lung surgery. Here, we introduced a novel drainage strategy with bi-pigtail catheters (PCs) and conducted a randomized controlled trial to compare with conventional CT drainage after uniportal video-assisted thoracic surgery lung surgery. METHODS: A single-centre, prospective, open-labelled, randomized controlled trial (ChiCTR2000035337) was conducted with a preplanned sample size of 396. The primary outcome was the numerical pain rating scale (NPRS) on the first postoperative day. Secondary outcomes included other indicators of postoperative pain, drainage volume, duration of drainage, postoperative hospital stay, incidence of postoperative complications, CT reinsertion and medical costs. RESULTS: A total number of 396 patients were randomized between August 2020 and January 2021, 387 of whom were included in the final analysis. The baseline and clinical characteristics of the patients were well balanced between 2 groups. The NPRS on the first postoperative day was significantly lower in the PC group than in the CT group (2.40 ± 1.27 vs 3.02 ± 1.39, p < 0.001), as well as the second/third-day NPRS, the incidence of sudden severe pain (9/192, 4.7% vs 34/195, 17.4%, P < 0.001) and pain requiring intervention (19/192, 9.9% vs 46/195, 23.6%, P < 0.001). In addition, the medical cost in the PC group was lower (US$7809 ± 1646 vs US$8205 ± 1815, P = 0.025). Univariable and multivariable analyses revealed that the drainage strategy was the only factor influencing the incidence of pain requiring intervention. CONCLUSIONS: The drainage strategy with bi-PCs in patients undergoing uniportal video-assisted thoracic surgery lung surgery alleviates postoperative pain with adequate safety and efficacy.

Structural visualization of transcription initiation in action.

Transcription initiation is a complex process, and its mechanism is incompletely understood. We determined the structures of de novo transcribing complexes TC2 to TC17 with RNA polymerase II halted on G-less promoters when nascent RNAs reach 2 to 17 nucleotides in length, respectively. Connecting these structures generated a movie and a working model. As initially synthesized RNA grows, general transcription factors (GTFs) remain bound to the promoter and the transcription bubble expands. Nucleoside triphosphate (NTP)-driven RNA-DNA translocation and template-strand accumulation in a nearly sealed channel may promote the transition from initially transcribing complexes (ITCs) (TC2 to TC9) to early elongation complexes (EECs) (TC10 to TC17). Our study shows dynamic processes of transcription initiation and reveals why ITCs require GTFs and bubble expansion for initial RNA synthesis, whereas EECs need GTF dissociation from the promoter and bubble collapse for promoter escape.

A COF template-derived mesoporous CeO2-supported Au nanoparticles catalyst for the oxidative esterification of benzaldehydes and benzyl alcohols.

The direct oxidative esterification of benzaldehydes and benzyl alcohols to high value-added aromatic esters under mild and green reaction conditions is significant in the fine chemical industry. The accurate design of catalysts with high catalytic performance is crucial for this process. Herein, 2,4,6-trimethylpyridine, benzoic anhydride, and terephthalaldehyde were used to prepare a covalent organic framework (COF) material, which was then used as a template to construct a mesoporous CeO2-supported Au nanoparticles catalyst. The obtained Au@CeO2 catalyst was thoroughly characterized, and it possessed a mesoporous structure with a high surface area. Meanwhile, the as-prepared Au@CeO2 exhibited excellent catalytic performance in the oxidative esterification of benzaldehydes and benzyl alcohols with methanol, affording the corresponding aromatic esters under mild and green reaction conditions. Furthermore, the Au@CeO2 catalyst could also be recycled. Therefore, this study provides a green and sustainable pathway for the synthesis of high-value-added esters through a direct oxidative esterification strategy.

Two genetic variants in the SRD5A2 gene are found to be associated with sex differences in the disease characteristics of patients with chronic hepatitis B virus infection.

BACKGROUND: To examine the expression characteristics of single nucleotide polymorphisms (SNPs) in the SRD5A2 gene and investigate their potential association with differences in the clinical characteristics between sexes in patients with chronic hepatitis B virus (HBV) infection. METHODS: A total of 30 loci in six genes primarily involved in the metabolism and signaling of sex hormones/sex hormone receptors, namely AKR1C2, AKR1C3, HSD17B6, SRD5A1, SRD5A2, and ESR1, were genotyped in 1007 patients from eight counties (cities) in Northeastern China with chronic HBV infection and 1040 healthy controls, and their association with viral replication characteristics and the differences in disease severity between sexes was assessed. Western blotting was conducted to determine the hepatic SRD5A2 protein level and its relationship with the inflammatory activity and fibrosis degree in male and female patients. RESULTS: Two SNP loci in the SRD5A2 gene (rs12470143 and rs7594951) exhibited significant differences in genotype and allele frequencies between sexes, with the proportion of T alleles significantly higher in males than in females. It was found that the incidence and severity of HBV-related liver fibrosis were significantly higher in patients with the T/T genotype in SRD5A2 rs12470143 and rs7594951 than those with the non-T/T genotype. Additionally, serum HBV DNA levels were significantly elevated in T/T patients compared to non-T/T patients. Female patients exhibited significantly lower serum DNA levels compared to male patients. Western blot analysis indicated that greater hepatic SRD5A2 protein levels were associated with higher METAVIR inflammation and fibrosis scores. Furthermore, multivariate analysis showed that the two genetic variants in the SRD5A2 gene (rs12470143 C > T, r7594951 C > T), together with the male sex, age > 50 years old, HBeAg positive status, elevated serum HBsAg load, high serum HBV DNA load, and HBV genotype C, were independent risk factors for HBV-related liver fibrosis. CONCLUSIONS: This study demonstrated that two genetic variants in the SRD5A2 gene (rs12470143 C > T, r7594951 C > T) are associated with sex differences in the clinical characteristics of patients with chronic HBV infection.

This study genotyped 30 genetic loci in six genes primarily involved in the metabolism and signaling pathways of sex hormones/sex hormone receptors, including AKR1C2, AKR1C3, HSD17B6, SRD5A1, SRD5A2, and ESR1, in 1007 patients with chronic hepatitis B virus (HBV) infection and 1040 healthy controls. It was found that two single nucleotide polymorphism (SNP) loci in the SRD5A2 gene (rs12470143 and rs7594951) showed significant differences in genotype and allele frequencies between male and female patients. Further, the proportion of the T alleles was significantly higher in males than in females. The study also found that patients with the T/T genotype had a higher incidence and severity of HBV-related liver fibrosis compared to those with other genotypes. Additionally, serum HBV DNA levels were significantly higher in T/T patients compared to non-T/T patients. Female patients had lower serum DNA levels compared to male patients. Further analysis showed that higher levels of the SRD5A2 protein were associated with increased inflammation and fibrosis scores in the liver. Multivariate analysis revealed that the two genetic variants in the SRD5A2 gene, together with male sex, age over 50, HBeAg positive status, elevated serum HBsAg load, high serum HBV DNA load, and HBV genotype C, were independent risk factors for HBV-related liver fibrosis. In summary, this study demonstrated that genetic variations in the SRD5A2 gene are associated with differences in the clinical characteristics of male and female patients with chronic HBV infection.

Metabolic disorders induced by PNPLA3 and TM6SF2 gene variants affect chronic kidney disease in patients infected with non-genotype 3 hepatitis C virus.

BACKGROUND: Patients with chronic hepatitis C virus (HCV) infections differ in their risk for metabolic disorders and chronic kidney disease (CKD). The aim of this study was to investigate the effect of metabolic disorders induced by genetic factors on CKD in HCV-infected patients. METHODS: Patients with chronic non-genotype 3 HCV infection with or without CKD were examined. PNPLA3 and TM6SF2 variants were determined using high-throughput sequencing. The relationships of variants and different combinations with metabolic disorders were analyzed in CKD patients. Univariate and multivariate analyses were used to identify factors associated with CKD. RESULTS: There were 1022 patients with chronic HCV infection, 226 with CKD and 796 without CKD. The CKD group had more severe metabolic disorders, and also had higher prevalences of liver steatosis, the PNPLA3 rs738409 non-CC genotype, and the TM6SF2 rs58542926 CC genotype (all P < 0.05). Relative to patients with the PNPLA3 rs738409 CC genotype, patients with the non-CC genotype had a significantly decreased eGFR and a greater prevalence of advanced CKD (CKD G4-5). Patients with the TM6SF2 rs58542926 CC genotype had a lower eGFR and a higher prevalence of CKD G4-5 than those with the non-CC genotype. Multivariable analysis indicated that multiple metabolic abnormalities, including liver steatosis and the PNPLA3 rs738409 C > G variant, increased the risk of CKD, but the TM6SF2 rs58542926 C > T variant decreased the risk of CKD. CONCLUSION: Specific PNPLA3 rs738409 and TM6SF2 rs58542926 variants are independent risk factors for CKD in patients with chronic HCV infections and are associated with the severity of renal injury.

Laser capture microdissection-based spatiotemporal transcriptomes uncover regulatory networks during seed abortion in seedless Ponkan (Citrus reticulata).

Seed abortion is an important process in the formation of seedless characteristics in citrus fruits. However, the molecular regulatory mechanism underlying citrus seed abortion is poorly understood. Laser capture microdissection-based RNA-seq combined with Pacbio-seq was used to profile seed development in the Ponkan cultivars 'Huagan No. 4' (seedless Ponkan) (Citrus reticulata) and 'E'gan No. 1' (seeded Ponkan) (C. reticulata) in two types of seed tissue across three developmental stages. Through comparative transcriptome and dynamic phytohormone analyses, plant hormone signal, cell division and nutrient metabolism-related processes were revealed to play critical roles in the seed abortion of 'Huagan No. 4'. Moreover, several genes may play indispensable roles in seed abortion of 'Huagan No. 4', such as CrWRKY74, CrWRKY48 and CrMYB3R4. Overexpression of CrWRKY74 in Arabidopsis resulted in severe seed abortion. By analyzing the downstream regulatory network, we further determined that CrWRKY74 participated in seed abortion regulation by inducing abnormal programmed cell death. Of particular importance is that a preliminary model was proposed to depict the regulatory networks underlying seed abortion in citrus. The results of this study provide novel insights into the molecular mechanism across citrus seed development, and reveal the master role of CrWRKY74 in seed abortion of 'Huagan No. 4'.

Intracellular common gardens reveal niche differentiation in transposable element community during bacterial adaptive evolution.

The distribution and abundance of transposable elements across the tree of life have significantly shaped the evolution of cellular organisms, but the underlying mechanisms shaping these ecological patterns remain elusive. Here we establish a "common garden" approach to study causal ecological interactions between a xenogeneic conditional lethal sacB gene and the community of transposable insertion sequences (ISs) in a multipartite prokaryote genome. Xenogeneic sacB of low, medium, or high GC content was individually inserted into three replicons of a model bacterium Sinorhizobium fredii, and exhibited replicon- and GC-dependent variation in genetic stability. This variation was largely attributable to multidimensional niche differentiation for IS community members. The transposition efficiency of major active ISs depended on the nucleoid-associated xenogeneic silencer MucR. Experimentally eliminating insertion activity of specific ISs by deleting MucR strongly demonstrated a dominant role of niche differentiation among ISs. This intracellular common garden approach in the experimental evolution context allows not only for evaluating genetic stability of natural and synthetic xenogeneic genes of different sequence signatures in host cells but also for tracking and testing causal relationships in unifying ecological principles in genome ecology.

Renewable-to-ammonia: Configuration strategy and technoeconomic analysis.

The increasing demand for chemical raw materials has provided opportunities for the ammonia (NH3) industry. However, little attention has been devoted to the economic feasibility of renewable-to-ammonia (RE2A). Therefore, this paper proposes a technoeconomic model to research the optimal capacity configuration and quantify the levelized cost of ammonia (LCOA) for RE2A, which is a retrofitted plant based on coal-to-ammonia (C2A). A cost model of C2A is established as a benchmark to evaluate the economic feasibility of RE2A. A case study in Inner Mongolia is adopted, which shows that the monthly NH3 output is 7-11×103t, which satisfies actual industrial production. The LCOA of RE2A is 469$/t, with investment in wind turbines accounting for 58%, which is lower than the NH3 market price (605$-650$/t). The LCOA of RE2A will equal that of C2A with a carbon tax of 47.1$/t CO2, which confirms the economic advantages of RE2A in the future.

Structure of nucleosome-bound human PBAF complex.

BAF and PBAF are mammalian SWI/SNF family chromatin remodeling complexes that possess multiple histone/DNA-binding subunits and create nucleosome-depleted/free regions for transcription activation. Despite previous structural studies and recent advance of SWI/SNF family complexes, it remains incompletely understood how PBAF-nucleosome complex is organized. Here we determined structure of 13-subunit human PBAF in complex with acetylated nucleosome in ADP-BeF3-bound state. Four PBAF-specific subunits work together with nine BAF/PBAF-shared subunits to generate PBAF-specific modular organization, distinct from that of BAF at various regions. PBAF-nucleosome structure reveals six histone-binding domains and four DNA-binding domains/modules, the majority of which directly bind histone/DNA. This multivalent nucleosome-binding pattern, not observed in previous studies, suggests that PBAF may integrate comprehensive chromatin information to target genomic loci for function. Our study reveals molecular organization of subunits and histone/DNA-binding domains/modules in PBAF-nucleosome complex and provides structural insights into PBAF-mediated nucleosome association complimentary to the recently reported PBAF-nucleosome structure.