Single Cell Analysis Linking Ribosomal (r)DNA and rRNA Copy Numbers to Cell Size and Growth Rate Provides Insights into Molecular Protistan Ecology

Ribosomal (r)RNA and rDNA have been golden molecular markers in microbial ecology. However, it remains poorly understood how ribotype copy number (CN)‐based characteristics are linked with diversity, abundance, and activity of protist populations and communities observed at organismal levels. Here, we applied a single‐cell approach to quantify ribotype CNs in two ciliate species reared at different temperatures. We found that in actively growing cells, the per‐cell rDNA and rRNA CNs scaled with cell volume (CV) to 0.44 and 0.58 powers, respectively. The modeled rDNA and rRNA concentrations thus appear to be much higher in smaller than in larger cells. The observed rRNA:rDNA ratio scaled with CV^0.14. The maximum growth rate could be well predicted by a combination of per‐cell ribotype CN and temperature. Our empirical data and modeling on single‐cell ribotype scaling are in agreement with both the metabolic theory of ecology and the growth rate hypothesis, providing a quantitative framework for linking cellular rDNA and rRNA CNs with body size, growth (activity), and biomass stoichiometry. This study also demonstrates that the expression rate of rRNA genes is constrained by cell size, and favors biomass rather than abundance‐based interpretation of quantitative ribotype data in population and community ecology of protists.     

Genome-wide identification of novel microRNAs from genome sequences using computational approach in the mudskipper (<Emphasis Type="Italic">Boleophthalmus pectinirostris</Emphasis>)

MicroRNAs (miRNAs), approximately 22 nucleotides (nt) long, are small, non-coding RNA molecules with important regulatory functions in gene expression. They are mostly conserved among the organisms and this conservation makes them a good source for the identification of novel miRNAs by computational genomic homology. The miRNA repertoire of a major aquaculture species, Boleophthalmus pectinirostris, has been unknown until recently. Currently, the B. pectinirostris whole-genome sequences have been completed, making it more convenient for us to focus on computational prediction for novel miRNA homologs. Following a range of strict filtering criteria, a total of 62 potential miRNAs were identified for the first time; they belong to 39 different miRNA families. All these miRNAs were observed in the stem portion of the stable stem–loop structures. The minimum free energy (MFE) of the predicted miRNAs ranged from ?21.6 to ?62.7 kcal/mol with an average of ?39.2 kcal/mol. The A + U ranged from 32.5 to 69.1% with an average value of 52.2%. The phylogenetic analysis of predicted miRNAs revealed that miR-23a-3p, miR-184-3p, miR-214-5p, and miR-338-3p from B. pectinirostris are evolutionary highly conserved showing more similarity with other fish species. To verify the predicted miRNAs, selected miRNAs representing 16 of the 39 families were confirmed by stem–loop RT-PCR, indicating that the computational approach that we used to identify the miRNAs is a highly efficient and affordable alternative method. Taken together, these findings provide a reference point for further research on miRNAs identification in fish species, meanwhile, our study also will be useful for further insight into biological functions of miRNAs and improved understanding of genome in B. pectinirostris.     

Comparative metabolomics reveals the mechanism of avermectin production enhancement by <Emphasis Type="Italic">S</Emphasis>-adenosylmethionine

It was found that S-adenosylmethionine (SAM) could effectively improve avermectin titer with 30–60 μg/mL addition to FH medium. To clearly elucidate the mechanism of SAM on intracellular metabolites of Streptomyces avermitilis, a GC–MS-based comparative metabolomics approach was carried out. First, 230 intracellular metabolites were identified and 14 of them remarkably influenced avermectin biosynthesis were discriminative biomarkers between non-SAM groups and SAM-treated groups by principal components analysis (PCA) and partial least squares (PLS). Based on further key metabolic pathway analyses, these biomarkers, such as glucose, oxaloacetic acid, fatty acids (in soybean oil), threonine, valine, and leucine, were identified as potentially beneficial precursors and added in medium. Compared with single-precursor feeding, the combined feeding of the precursors and SAM markedly increased the avermectin titer. The co-feeding approach not only directly verified our hypothesis on the mechanism of SAM by comparative metabolomics, but also provided a novel strategy to increase avermectin production.     

Long noncoding RNA expression profiles in chondrogenic and hypertrophic differentiation of mouse mesenchymal stem cells

Long noncoding RNAs (lncRNAs) are important regulators for a variety of biological processes. Chondrogenic differentiation of mesenchymal stem cells (MSCs) is a crucial stage in chondrogenesis while chondrocyte hypertrophy is related to endochondral ossification and osteoarthritis. However, the effects of lncRNAs on chondrogenic and hypertrophic differentiation of mouse MSCs are unclear. To explore the potential mechanisms of lncRNAs during chondrogenesis and chondrocyte hypertrophy, microarray was performed to investigate the expression profiles of lncRNA and mRNA in MSCs, pre-chondrocytes, and hypertrophic chondrocytes. Then, we validated microarray data by RT-PCR and screened three lncRNAs from upregulating groups during chondrogenesis and chondrocyte hypertrophy respectively. After downregulating any of the above lncRNAs, we found that the expression of chondrogenesis-related genes such as Sox9 and Col2a1 and hypertrophy-related genes including Runx2 and Col10a1 was inhibited, respectively. Furthermore, the target genes of above lncRNAs were predicted by bioinformatics approaches. Gene ontology and Kyoto encyclopedia of genes and genome biological pathway analysis were also made to speculate the functions of above lncRNAs. In conclusion, the study first revealed the expression profile of lncRNAs in chondrogenic and hypertrophic differentiations of mouse MSCs and presented a new prospect for the underlying mechanisms of chondrogenesis and endochondral ossification.     

The characterization of plant species using first‐derivative fluorescence spectra

Plants are one of the most important parts of the ecological system and demand a reliable method for accurate classification. In this study, the first‐derivative fluorescence spectral curves (FDFSCs) based on laser‐induced fluorescence technology were proposed for the characterization of plant species. The measurement system is mainly composed of a spectrometer, an excitation light source (the two excitation wavelengths are 460 and 556 nm, respectively), and an intensified charge‐coupled device camera. FDFSCs were calculated from the deviation between the fluorescence values at each wavelength, plus and minus one band, divided by the wavelength range. Principal component analysis was utilized to analyze the FDFSCs by extracting the main attributes and reducing the dimensionality of variables. A support vector machine was used to evaluate FDFSC performance for the identification of plant species. Plant species that are difficult to distinguished by the naked eye, can be identified effectively using the proposed FDFSCs. For the 556 nm and 460 nm excitation wavelengths, the overall identification rates of the six plant species evaluated were 93.3% and 91.7%, respectively. Experimental results demonstrated that the combination of the FDFSCs with multivariate analysis could provide a simple and reliable method for the characterization of plant species.     

De-dimerization of PTB is catalyzed by PDI and is involved in the regulation of p53 translation

Polypyrimidine tract-binding protein (PTB) is an RNA binding protein existing both as dimer and monomer and shuttling between nucleus and cytoplasm. However, the regulation of PTB dimerization and the relationship between their functions and subcellular localization are unknown. Here we find that PTB presents as dimer and monomer in nucleus and cytoplasm respectively, and a disulfide bond involving Cysteine 23 is critical for the dimerization of PTB. Additionally, protein disulfide isomerase (PDI) is identified to be the enzyme that catalyzes the de-dimerization of PTB, which is dependent on the CGHC active site of the a’ domain of PDI. Furthermore, upon DNA damage induced by topoisomerase inhibitors, PTB is demonstrated to be de-dimerized with cytoplasmic accumulation. Finally, cytoplasmic PTB is found to associate with the ribosome and enhances the translation of p53. Collectively, these findings uncover a previously unrecognized mechanism of PTB dimerization, and shed light on the de-dimerization of PTB functionally linking to cytoplasmic localization and translational regulation.     

Spikes with short inter-spike intervals in frog retinal ganglion cells are more correlated with their adjacent neurons’ activities

Correlated firings among neurons have been extensively investigated; however, previous studies on retinal ganglion cell (RGC) population activities were mainly based on analyzing the correlated activities between the entire spike trains. In the present study, the correlation properties were explored based on burst-like activities and solitary spikes separately. The results indicate that: (1) burst-like activities were more correlated with other neurons’ activities; (2) burst-like spikes correlated with their neighboring neurons represented a smaller receptive field than that of correlated solitary spikes. These results suggest that correlated burst-like spikes should be more efficient in signal transmission, and could encode more detailed spatial information.     

Extracellular vesicles package dsDNA to aggravate Crohn’s disease by activating the STING pathway

Crohn’s disease (CD) is an intestinal immune-dysfunctional disease. Extracellular vesicles (EVs) are membrane-enclosed particles full of functional molecules, e.g., nuclear acids. Recently, EVs have been shown to participate in the development of CD by realizing intercellular communication among intestinal cells. However, the role of EVs carrying double-strand DNA (dsDNA) shed from sites of intestinal inflammation in CD has not been investigated. Here we isolated EVs from the plasma or colon lavage of murine colitis and CD patients. The level of exosomal dsDNA, including mtDNA and nDNA, significantly increased in murine colitis and active human CD, and was positively correlated with the disease activity. Moreover, the activation of the STING pathway was verified in CD. EVs from the plasma of active human CD triggered STING activation in macrophages in vitro. EVs from LPS-damaged colon epithelial cells were also shown to raise inflammation in macrophages via activating the STING pathway, but the effect disappeared after the removal of exosomal dsDNA. These findings were further confirmed in STING-deficient mice and macrophages. STING deficiency significantly ameliorated colitis. Besides, potential therapeutic effects of GW4869, an inhibitor of EVs release were assessed. The application of GW4869 successfully ameliorated murine colitis by inhibiting STING activation. In conclusion, exosomal dsDNA was found to promote intestinal inflammation via activating the STING pathway in macrophages and act as a potential mechanistic biomarker and therapeutic target of CD.Subject terms: Acute inflammation, Monocytes and macrophages, Signal transduction