A plant-specific SWR1 chromatin-remodeling complex couples histone H2A.Z deposition with nucleosome sliding.

Deposition of H2A.Z in chromatin is known to be mediated by a conserved SWR1 chromatin-remodeling complex in eukaryotes. However, little is known about whether and how the SWR1 complex cooperates with other chromatin regulators. Using immunoprecipitation followed by mass spectrometry, we found all known components of the Arabidopsis thaliana SWR1 complex and additionally identified the following three classes of previously uncharacterized plant-specific SWR1 components: MBD9, a methyl-CpG-binding domain-containing protein; CHR11 and CHR17 (CHR11/17), ISWI chromatin remodelers responsible for nucleosome sliding; and TRA1a and TRA1b, accessory subunits of the conserved NuA4 histone acetyltransferase complex. MBD9 directly interacts with CHR11/17 and the SWR1 catalytic subunit PIE1, and is responsible for the association of CHR11/17 with the SWR1 complex. MBD9, TRA1a, and TRA1b function as canonical components of the SWR1 complex to mediate H2A.Z deposition. CHR11/17 are not only responsible for nucleosome sliding but also involved in H2A.Z deposition. These results indicate that the association of the SWR1 complex with CHR11/17 may facilitate the coupling of H2A.Z deposition with nucleosome sliding, thereby co-regulating gene expression, development, and flowering time.

Dual Recognition of H3K4me3 and DNA by the ISWI Component ARID5 Regulates the Floral Transition in Arabidopsis.

Chromatin remodeling and histone modifications are important for development and floral transition in plants. However, it is largely unknown whether and how these two epigenetic regulators coordinately regulate the important biological processes. Here, we identified three types of Imitation Switch (ISWI) chromatin-remodeling complexes in Arabidopsis (Arabidopsis thaliana). We found that AT-RICH INTERACTING DOMAIN5 (ARID5), a subunit of a plant-specific ISWI complex, can regulate development and floral transition. The ARID-PHD dual domain cassette of ARID5 recognizes both the H3K4me3 histone mark and AT-rich DNA. We determined the ternary complex structure of the ARID5 ARID-PHD cassette with an H3K4me3 peptide and an AT-containing DNA. The H3K4me3 peptide is combinatorially recognized by the PHD and ARID domains, while the DNA is specifically recognized by the ARID domain. Both PHD and ARID domains are necessary for the association of ARID5 with chromatin. The results suggest that the dual recognition of AT-rich DNA and H3K4me3 by the ARID5 ARID-PHD cassette may facilitate the association of the ISWI complex with specific chromatin regions to regulate development and floral transition.

The CCR4-NOT complex component NOT1 regulates RNA-directed DNA methylation and transcriptional silencing by facilitating Pol IV-dependent siRNA production.

Small interfering RNAs (siRNAs) are responsible for establishing and maintaining DNA methylation through the RNA-directed DNA methylation (RdDM) pathway in plants. Although siRNA biogenesis is well known, it is relatively unclear about how the process is regulated. By a forward genetic screen in Arabidopsis thaliana, we identified a mutant defective in NOT1 and demonstrated that NOT1 is required for transcriptional silencing at RdDM target genomic loci. We demonstrated that NOT1 is required for Pol IV-dependent siRNA accumulation and DNA methylation at a subset of RdDM target genomic loci. Furthermore, we revealed that NOT1 is a constituent of a multi-subunit CCR4-NOT deadenylase complex by immunoprecipitation combined with mass spectrometry and demonstrated that the CCR4-NOT components can function as a whole to mediate chromatin silencing. Therefore, our work establishes that the CCR4-NOT complex regulates the biogenesis of Pol IV-dependent siRNAs, and hence facilitates DNA methylation and transcriptional silencing in Arabidopsis.

The HDA19 histone deacetylase complex is involved in the regulation of flowering time in a photoperiod-dependent manner.

Chromatin modifications are known to affect flowering time in plants, but little is known about how these modifications regulate flowering time in response to environmental signals like photoperiod. In Arabidopsis thaliana, HDC1, a conserved subunit of the RPD3-like histone deacetylase (HDAC) complex, was previously reported to regulate flowering time via the same mechanism as does the HDAC HDA6. Here, we demonstrate that HDC1, SNLs and MSI1 are shared subunits of the HDA6 and HDA19 HDAC complexes. While the late-flowering phenotype of the hda6 mutant is independent of photoperiod, the hda19, hdc1 and snl2/3/4 mutants flower later than or at a similar time to the wild-type in long-day conditions but flower earlier than the wild-type in short-day conditions. Our genome-wide analyses indicate that the effect of hdc1 on histone acetylation and transcription is comparable with that of hda19 but is different from that of hda6. Especially, we demonstrate that the HDA19 complex directly regulates the expression of two flowering repressor genes related to the gibberellin signaling pathway. Thus, the study reveals a photoperiod-dependent role of the HDA19 HDAC complex in the regulation of flowering time.

Characterization of Myostain (MSTN) and Myogenic Differentiation Antigen (MyoD) and the Effect of Dexamethasone on Their Expression in Large-Scale Loach Paramisgurnus dabryanus.

Myostatin (MSTN) and myogenic differentiation antigen (MyoD) play an essential role in specification and differentiation of skeletal muscle. However, the role of stress in the regulation of MyoD and MSTN has not been fully revealed and more evidence should be provided. Here, we reported the cloning and expressional analyses of MSTN and MyoD in Large-scale Loach Paramisgurnus dabryanus (hereafter PdMSTN and PdMyoD). Injecting individuals with 0, 60, 600, and 1,200 µg/kg dexamethasone (DXM) for five consecutive days resulted in a dose-dependent change of PdMSTN and PdMyoD expression. The expression of PdMSTN was upregulated with increasing DXM concentrations, while PdMyoD expression was downregulated. The changes in the expression of these genes at different time points for 10 consecutive days were studied after individuals were treated with 600 µg/kg DXM. Compared with the control group, PdMSTN expression decreased and PdMyoD expression increased before 12 h, and both PdMSTN and PdMyoD expression levels increased at 24 h, which was significantly higher than those in control group. At a prolonged treatment of 5-10 d, expression levels of PdMSTN and PdMyoD had significantly reduced. The results indicate that both PdMyoD and PdMSTN are involved in DXM-induced stress in Large-scale Loach.

A methylated-DNA-binding complex required for plant development mediates transcriptional activation of promoter methylated genes.

Although the mechanism of DNA methylation-mediated gene silencing is extensively studied, relatively little is known about how promoter methylated genes are protected from transcriptional silencing. SUVH1, an Arabidopsis Su(var)3-9 homolog, was previously shown to be required for the expression of a few promoter methylated genes. By chromatin immunoprecipitation combined with sequencing, we demonstrate that SUVH1 binds to methylated genomic loci targeted by RNA-directed DNA methylation. SUVH1 and its homolog SUVH3 function partially redundantly and interact with three DNAJ domain-containing homologs, SDJ1, SDJ2, and SDJ3, thus forming a complex which we named SUVH-SDJ. The SUVH-SDJ complex components are co-localized in a large number of methylated promoters and are required for the expression of a subset of promoter methylated genes. We demonstrate that the SUVH-SDJ complex components have transcriptional activation activity. SUVH1 and SUVH3 function synergistically with SDJ1, SDJ2, and SDJ3 and are required for plant viability. This study reveals how the SUVH-SDJ complex protects promoter methylated genes from transcriptional silencing and suggests that the transcriptional activation of promoter methylated genes mediated by the SUVH-SDJ complex may play a critical role in plant growth and development.

Clinical Value of Metagenomics Next-Generation Sequencing in Antibiotic Resistance of a Patient with Severe Refractory Mycoplasma pneumoniae Pneumonia: A Case Report.

Background: Mycoplasma pneumoniae is an important infectious pathogen of lower respiratory tract infection in children and adolescents. Macrolide resistant M. pneumoniae (MRMP) has become increasingly prevalent, and identifying pathogen resistance genes is crucial for treatment. Case Presentation: We report a patient with severe refractory M. pneumoniae pneumonia (MPP). The failure of initial clinical treatment prompted the re-analysis of metagenomic next-generation sequencing (mNGS) data for macrolide-resistant gene. Macrolide-resistance 23S ribosomal RNA gene was confirmed with read depth of 64 X for the A2063G mutation, which can decrease the affinity of macrolide with M. pneumoniae ribosome resulting in macrolide resistance. Furthermore, antimicrobial susceptibility testing demonstrated that M. pneumoniae was resistant to macrolide. PCR confirmatory test about M. pneumoniae resistance A2063G mutation, clinical treatment course and prognosis with altered treatment strategy, and M. pneumoniae antimicrobial susceptibility confirmed that the severe refractory MPP was due to macrolide resistant M. pneumoniae. Conclusion: As a new molecular level detection, mNGS is an effective method for detecting M. pneumoniae resistance genes. Early recognition of macrolide resistance and suitable antibiotics strategy is of vital importance for the prognosis of severe refractory MPP.

16S-FASAS: an integrated pipeline for synthetic full-length 16S rRNA gene sequencing data analysis.

Background: The full-length 16S rRNA sequencing can better improve the taxonomic and phylogenetic resolution compared to the partial 16S rRNA gene sequencing. The 16S-FAS-NGS (16S rRNA full-length amplicon sequencing based on a next-generation sequencing platform) technology can generate high-quality, full-length 16S rRNA gene sequences using short-read sequencers, together with assembly procedures. However there is a lack of a data analysis suite that can help process and analyze the synthetic long read data. Results: Herein, we developed software named 16S-FASAS (16S full-length amplicon sequencing data analysis software) for 16S-FAS-NGS data analysis, which provided high-fidelity species-level microbiome data. 16S-FASAS consists of data quality control, de novo assembly, annotation, and visualization modules. We verified the performance of 16S-FASAS on both mock and fecal samples. In mock communities, we proved that taxonomy assignment by MegaBLAST had fewer misclassifications and tended to find more low abundance species than the USEARCH-UNOISE3-based classifier, resulting in species-level classification of 85.71% (6/7), 85.71% (6/7), 72.72% (8/11), and 70% (7/10) of the target bacteria. When applied to fecal samples, we found that the 16S-FAS-NGS datasets generated contigs grouped into 60 and 56 species, from which 71.62% (43/60) and 76.79% (43/56) were shared with the Pacbio datasets. Conclusions: 16S-FASAS is a valuable tool that helps researchers process and interpret the results of full-length 16S rRNA gene sequencing. Depending on the full-length amplicon sequencing technology, the 16S-FASAS pipeline enables a more accurate report on the bacterial complexity of microbiome samples. 16S-FASAS is freely available for use at https://github.com/capitalbio-bioinfo/FASAS.

Complete mitochondrial genome of Schizopygopsis chengi chengi (Cyprinidae, Schizothoracinae, Schizopygopsis).

The complete mitochondrial DNA sequence of Schizopygopsis chengi chengi collected from the Dadu River was determined in this study. The complete mitochondrial genome of S. chengi chengi was a circular molecule of 16 688 bp in size, and all genes showed the typical gene arrangement conforming to the vertebrate consensus. Twelve protein-coding genes (except ND6) of S. chengi chengi and other 22 Schizothoracinae species from two genera were used for phylogenetic analysis by Bayesian inference and maximum likelihood methods. The topology confirmed the close genetic relationship between S. chengi chengi and S. chengi baoxinggensis.