Somatostatin-14 [somatotropin release-inhibiting factor (SRIF)] reduces hippocampal epileptiform activity but the contribution of its specific receptors (sst1-5) is poorly understood. We have focused on the role of sst1 and sst2 in mediating SRIF modulation of epilepsy using hippocampal slices of wild-type (WT) and sst1 or sst2 knockout (KO) mice. Recordings of epileptiform discharge induced by Mg2+ -free medium with 4-aminopyridine were performed from the CA3 region before and after the application of SRIF compounds. In WT mice, SRIF and the sst1 agonist CH-275 reduce epilepsy whereas sst1 blockade with its antagonist SRA-880 increases the bursting discharge. Activation of sst2 does not affect the bursting frequency unless its agonist octreotide is applied with SRA-880, indicating that sst1 masks sst2-mediated modulation of epilepsy. In sst1 KO mice: (i) the bursting frequency is lower than in WT; (ii) SRIF, CH-275 and SRA-880 are ineffective on epilepsy and (iii) octreotide is also devoid of effects, whereas blockade of sst2 with the antagonist D-Tyr8 Cyn 154806 increases the bursting frequency. In sst2 KO mice, the SRIF ligand effects are similar to those in WT. In the whole hippocampus of sst1 KO mice, sst2 mRNA, protein and binding are higher than in WT and reverse transcription-polymerase chain reaction of the CA3 subarea confirms an increase of the sst2 messenger. We conclude that sst1 mediates inhibitory actions of SRIF and that interactions between sst1 and sst2 may prevent sst2 modulation of epilepsy. We suggest that, in sst1 KO mice, activation of over-expressed sst2 reduces the bursting frequency, indicating that sst2 density represents the rate-limiting factor for ss(2-mediated modulation of epilepsy.

Uncovering the full potential of gas-fermenting Clostridia, attractive autotrophic bacteria capable of using synthesis gases (CO–CO2–H2) to produce a range of chemicals and fuels, for industrial applications relies on having efficient molecular tools for genetic modifications. Although the CRISPR-Cas9-mediated genome editing system has been developed in Clostridia, its use is limited owing to low GC content (approx. 30%) in these anaerobes. Therefore, the effector protein Cas12a, which recognizes T-rich instead of G-rich protospacer-adjacent motifs (PAMs), has evident advantages over Cas9 in CRISPR genome editing in Clostridia. Here, we developed the CRISPR-Cas12a system for efficient gene deletion and regulation in the gas-fermenting Clostridium ljungdahlii species. On the basis of screening for the most suitable Cas12a and significantly improved electrotransformation efficiency that bypassed poor repair efficiency of the Cas12a-caused DNA double-strand break (DSB) in C. ljungdahlii, efficient deletion (80–100%) of four genes (pyrE, pta, adhE1, and ctf) was achieved by using the CRISPR-FnCas12a system. Furthermore, a DNase-deactivated FnCas12a (ddCas12a) was adopted to construct a CRISPRi system to downregulate targeted genes, reaching over 80% repression for most of the chosen binding sites. This CRISPRi system was also used in a butyric acid-producing C. ljungdahlii strain to redirect carbon flux, leading to 20–40% reductions in ethanol titer that were accompanied by increased butyric acid titer. These results demonstrate the high efficiency of the CRISPR-FnCas12a system for genome engineering in C. ljungdahlii, which effectively expands the existing CRISPR-Cas toolbox in gas-fermenting Clostridium species and may play important roles in genetic manipulations where CRISPR-Cas9 is incompetent.

In this study, we used pan RNA-seq analysis to reveal the ubiquitous existence of both 5′ and 3′ end small RNAs (5′ and 3′ sRNAs). 5′ and 3′ sRNAs alone can be used to annotate nuclear non-coding and mitochondrial genes at 1-bp resolution and identify new steady RNAs, which are usually transcribed from functional genes. Then, we provided a simple and cost effective way for the annotation of nuclear non-coding and mitochondrial genes and the identification of new steady RNAs, particularly long non-coding RNAs (lncRNAs). Using 5′ and 3′ sRNAs, the annotation of human mitochondrial was corrected and a novel ncRNA named non-coding mitochondrial RNA 1 (ncMT1) was reported for the first time in this study. We also found that most of human tRNA genes have downstream lncRNA genes as lncTRS-TGA1-1 and corrected the misunderstanding of them in previous studies. Using 5′, 3′, and intronic sRNAs, we reported for the first time that enzymatic double-stranded RNA (dsRNA) cleavage and RNA interference (RNAi) might be involved in the RNA degradation and gene expression regulation of U1 snRNA in human. We provided a different perspective on the regulation of gene expression in U1 snRNA. We also provided a novel view on cancer and virus-induced diseases, leading to find diagnostics or therapy targets from the ribonuclease III (RNase III) family and its related pathways. Our findings pave the way toward a rediscovery of dsRNA cleavage and RNAi, challenging classical theories.

Herein, by using two fluorinated and chlorinated monomers with similar structures in different molar ratios and dithieno­[3′,2′:3,4;2″,3″:5,6]­benzo­[1,2-c]­[1,2,5]­thiadiazole (DTBT) as the third unit, a family of polymer donors D18, D18–20%Cl, D18–40%Cl, and D18–Cl are synthesized for OSCs. With appropriate chlorinated monomer proportion, the terpolymer D18–20%Cl exhibits proper HOMO energy level and higher packing density compared with that of other control polymers. Moreover, the D18–20%Cl:Y6 blend films have favorable morphology with better face-on crystallization and better charge transport. Consequently, the D18–20%Cl:Y6-based OSCs obtain a top-ranked PCE of 18.28% with overall improved device parameters compared to the controlled D18:Y6 or D18-Cl:Y6-based OSCs (17.50% or 17.02%), which represents the highest PCE for the reported terpolymer-based binary OSCs. Notably, the D18–20%Cl:Y6-based OSCs exhibit over 17% efficiency in a wide molecular weight range. These results demonstrate that the ternary copolymerization of DTBT and two similar moieties is an efficient approach for achieving efficient terpolymer donors with well batch-to-batch reproducibility.

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Protein lysine acetylation, a prevalent posttranslational modification, regulates numerous crucial biological processes in cells. Nevertheless, how lysine acetylation interacts with other types of regulation to coordinate metabolism remains largely unknown owing to the complexity of the process. Here, using a representative gas-fermenting bacterium, Clostridium ljungdahlii, we revealed a novel regulatory mechanism that employs both the lysine acetylation and transcriptional regulation systems to interactively control CO2 fixation, a key biological process for utilizing this one-carbon gas. A dominant lysine acetyltransferase/deacetylase system, At2/Dat1, was identified and found to regulate FDH1 (formate dehydrogenase responsible for CO2 fixation) activity via a crucial acetylation site (lysine-29). Notably, the global transcription factor CcpA was also shown to be regulated by At2/Dat1; in turn, CcpA could directly control At2 expression, thus indicating an unreported interaction mode between the acetylation system and transcription factors. Moreover, CcpA was observed to negatively regulate FDH1 expression, which, when combined with At2/Dat1, leads to the collaborative regulation of this enzyme. Based on this concept, we reconstructed the regulatory network related to FDH1, realizing significantly increased CO2 utilization by C. ljungdahlii. IMPORTANCE Microbial CO2 fixation and conversion constitute a potential solution to both utilization of greenhouse gas or industrial waste gases and sustainable production of bulk chemicals and fuels. Autotrophic gas-fermenting bacteria play central roles in this bioprocess. This study provides new insights regarding the metabolic regulatory mechanisms underlying CO2 reduction in Clostridium ljungdahlii, a representative gas-fermenting bacterium. A critical formate dehydrogenase (FDH1) responsible for fixing CO2 and a dominant reversible lysine acetylation system, At2/Dat1, were identified. Furthermore, FDH1 was found to be interactively regulated by both the At2/Dat1 system and the global transcriptional factor CcpA, and the two regulatory systems are mutually restricted. Reconstruction of this multilevel metabolic regulatory module led to improved CO2 metabolism by C. ljungdahlii. These findings not only substantively expand our understanding but also provide a potentially useful metabolic engineering strategy for microbial carbon fixation.

The master regulator CcpA (catabolite control protein A) manages a large and complex regulatory network that is essential for cellular physiology and metabolism in Gram-positive bacteria. Although CcpA can affect the expression of target genes by binding to a -acting catabolite-responsive element (), whether and how the expression of CcpA is regulated remain poorly explored. Here, we report a novel dual- motif that is employed by the CcpA in , a typical solventogenic species, for autoregulation. Two sites are involved in CcpA autoregulation, and they reside in the promoter and coding regions of CcpA. In this dual- motif, , in the promoter region, positively regulates transcription, whereas , in the coding region, negatively regulates this transcription, thus enabling two-way autoregulation of CcpA. Although CcpA bound more strongly than , the assay showed that -based repression dominates CcpA autoregulation during the entire fermentation. Finally, a synonymous mutation of was made within the coding region, achieving an increased intracellular CcpA expression and improved cellular performance. This study provides new insights into the regulatory role of CcpA in and, moreover, contributes a new engineering strategy for this industrial strain. CcpA is known to be a key transcription factor in Gram-positive bacteria. However, it is still unclear whether and how the intracellular CcpA level is regulated, which may be essential for maintaining normal cell physiology and metabolism. We discovered here that CcpA employs a dual- motif to autoregulate, enabling dynamic control of its own expression level during the entire fermentation process. This finding answers the questions above and fills a void in our understanding of the regulatory network of CcpA. Interference in CcpA autoregulation leads to improved cellular performance, providing a new useful strategy in genetic engineering of Since CcpA is widespread in Gram-positive bacteria, including pathogens, this dual--based CcpA autoregulation would be valuable for increasing our understanding of CcpA-based global regulation in bacteria.

This paper investigates the meshing behavior of the roller screw, a mechanical transmission device characterized by threaded rollers that transfer a load between the nut and the screw, by analyzing the meshing characteristics between screw and rollers. This study seeks to establish a more accurate mathematical model for the thread surface by creating a modeling process in which the max radiuses of the threads are calculated more precisely. The contact line distribution and the contact location were also calculated in order to confirm the cross section of the meshing points. In the research presented in this paper, the actual transmission ratio is analyzed and the study results in a new method to calculate the actual transmission ratio. In this study, the helical angle and the vertex angle are proven to be of great significance after a careful analysis of their influence is conducted.