Circular RNA: Biosynthesis in vitro

Circular RNA (circRNA) is a unique type of noncoding RNA molecule. Compared with traditional linear RNA, circRNA is a covalently closed circle produced by a process called backsplicing. CircRNA is abundant in many cells and has rich functions in cells, such as acting as miRNA sponge, protein sponge, protein scaffold, and mRNA regulator. With the continuous development of circRNA study, circRNA has also played an important role in medical applications, including circRNA vaccines and gene therapy. In this review, we illustrate the synthesis of circRNAs in vitro. We focus on biological ligation methods, such as enzymatic ligation from the bacteriophage T4 and ribozyme method. In addition, we summarize the current challenges in the design, synthesis, application, and production of circRNAs, and propose possible solutions in the future. CircRNA is expected to play an essential role in basic research and medical applications.

Integration of metabolomics and transcriptomics indicates changes in MRSA exposed to terpinen-4-ol

Background This study investigated the effects of terpinen-4-ol on methicillin-resistant Staphylococcus aureus (MRSA) and its biofilm, and the possible mechanisms governing this effect. Results We observed that terpinen-4-ol has good antibacterial activity and inhibits the formation of MRSA biofilm. The MIC and MBC values for terpinen-4-ol against S. aureus were 0.08% ~ 0.32%. And terpinen-4-ol at 0.32% could kill all bacteria and clear all biofilms. Untargeted metabolomic and transcriptomic analyses showed that terpinen-4-ol strongly inhibited DNA and RNA biosynthesis in MRSA at 2 h after treatment by affecting genes and metabolites related to purine and pyrimidine metabolic pathways. Some differential genes which play important roles in DNA synthesis and the production of eDNA from biofilm exposed to terpinen-4-ol was also significantly decreased compared with that of the control. Conclusions Terpinen-4-ol has good antibacterial activity and significantly inhibits the formation of MRSA biofilm by inhibiting purine and pyrimidine metabolism.

Targeting Pyrimidine Metabolism in the Era of Precision Cancer Medicine

Metabolic rewiring is considered as a primary feature of cancer. Malignant cells reprogram metabolism pathway in response to various intrinsic and extrinsic drawback to fuel cell survival and growth. Among the complex metabolic pathways, pyrimidine biosynthesis is conserved in all living organism and is necessary to maintain cellular fundamental function (i.e. DNA and RNA biosynthesis). A wealth of evidence has demonstrated that dysfunction of pyrimidine metabolism is closely related to cancer progression and numerous drugs targeting pyrimidine metabolism have been approved for multiple types of cancer. However, the non-negligible side effects and limited efficacy warrants a better strategy for negating pyrimidine metabolism in cancer. In recent years, increased studies have evidenced the interplay of oncogenic signaling and pyrimidine synthesis in tumorigenesis. Here, we review the recent conceptual advances on pyrimidine metabolism, especially dihydroorotate dehydrogenase (DHODH), in the framework of precision oncology medicine and prospect how this would guide the development of new drug precisely targeting the pyrimidine metabolism in cancer.

Advances in the Relationship Between Regulator of Ribosome Synthesis 1 (RRS1) and Diseases

A regulator of ribosome synthesis 1 (RRS1) was discovered in yeast and is mainly localized in the nucleolus and endoplasmic reticulum. It regulates ribosomal protein, RNA biosynthesis, and protein secretion and is closely involved in cellular senescence, cell cycle regulation, transcription, translation, oncogenic transformation etc., Mutations in the RRS1 gene are associated with the occurrence and development of Huntington’s disease and cancer, and overexpression of RRS1 promotes tumor growth and metastasis. In this review, the structure, function, and mechanisms of RRS1 in various diseases are discussed.

Integration of Metabolomics and Transcriptomics Indicates Changes in MRSA Exposed to Terpinen-4-ol

Background: This study investigated the effects of terpinen-4-ol on methicillin-resistant Staphylococcus aureus (MRSA) and its biofilm, and the possible mechanisms governing this effect.Results: We observed that terpinen-4-ol has good antibacterial activity and inhibits the formation of MRSA biofilm. The MIC and MBC values for terpinen-4-ol against S. aureus were 0.08%~0.32%. And terpinen-4-ol at 0.32% could kill all bacteria and clear all biofilms. Untargeted metabolomic and transcriptomic analyses showed that terpinen-4-ol strongly inhibited DNA and RNA biosynthesis in MRSA at 2 h after treatment by affecting genes and metabolites related to purine and pyrimidine metabolic pathways. Some differential genes which play important roles in DNA synthesis and the production of eDNA from biofilm exposed to terpinen-4-ol was also significantly decreased compared with that of the control. Conclusions: Terpinen-4-ol has good antibacterial activity and significantly inhibits the formation of MRSA biofilm by inhibiting purine and pyrimidine metabolism.

Chromatophores efficiently promote light-driven ATP synthesis and DNA transcription inside hybrid multicompartment artificial cells

The construction of energetically autonomous artificial protocells is one of the most ambitious goals in bottom-up synthetic biology. Here, we show an efficient manner to build adenosine 5′-triphosphate (ATP) synthesizing hybrid multicompartment protocells. Bacterial chromatophores from Rhodobacter sphaeroides accomplish the photophosphorylation of adenosine 5′-diphosphate (ADP) to ATP, functioning as nanosized photosynthetic organellae when encapsulated inside artificial giant phospholipid vesicles (ATP production rate up to ∼100 ATP∙s−1 per ATP synthase). The chromatophore morphology and the orientation of the photophosphorylation proteins were characterized by cryo-electron microscopy (cryo-EM) and time-resolved spectroscopy. The freshly synthesized ATP has been employed for sustaining the transcription of a DNA gene, following the RNA biosynthesis inside individual vesicles by confocal microscopy. The hybrid multicompartment approach here proposed is very promising for the construction of full-fledged artificial protocells because it relies on easy-to-obtain and ready-to-use chromatophores, paving the way for artificial simplified-autotroph protocells (ASAPs).

AMPK Regulates Developmental Plasticity through an Endogenous Small RNA Pathway in Caenorhabditis elegans

Caenorhabditis elegans larvae can undergo developmental arrest upon entry into the dauer stage in response to suboptimal growth conditions. Dauer larvae can exit this stage in replete conditions with no reproductive consequence. During this diapause stage, the metabolic regulator AMP-activated protein kinase (AMPK) ensures that the germ line becomes quiescent to maintain germ cell integrity. Animals that lack all AMPK signalling undergo germline hyperplasia upon entering dauer, while those that recover from this stage become sterile. Neuronal AMPK expression in otherwise AMPK-deficient animals is sufficient for germline quiescence and germ cell integrity and its effects are likely mediated through an endogenous small RNA pathway. Upon impairing small RNA biosynthesis, the post-dauer fertility is restored in AMPK mutants. These data suggest that AMPK may function in neurons to relay a message through small RNAs to the germ cells to alter their quiescence in the dauer stage, thus challenging the permeability of the Weismann barrier.

Time-resolved NMR monitoring of tRNA maturation

ABSTRACTAlthough the biological importance of post-transcriptional RNA modifications in gene expression is widely appreciated, methods to directly detect the introduction of these modifications during RNA biosynthesis are rare and do not easily provide information on the temporal nature of events. Here we introduce the application of NMR spectroscopy to observe the maturation of tRNAs in cell extracts. By following the maturation of yeast tRNAPhewith time-resolved NMR measurements, we found that modifications are introduced in a defined sequential order, and that the chronology is controlled by cross-talk between modification events. In particular, we uncovered a strong hierarchy in the introduction of the T54, Ψ55 and m1A58 modifications in the T-arm, and demonstrate that the modification circuits identified in yeast extract with NMR also impact the tRNA modification process in living cells. The NMR-based methodology presented here could be adapted to investigate different aspects of tRNA maturation and RNA modifications in general.

BIOSINTESA FOLAT OLEH BAKTERI ASAM LAKTAT

Folate, an important B-group vitamin, participates in many metabolic pathways such as DNA and RNA biosynthesis and amino acid inter-conversions. Mammalian cells cannot synthesize folate; therefore, an exogenous supply of this vitamin is necessary to prevent nutritional deficiency. Folic acid is a composite molecule, being made up of three parts: a pteridine ring system (6-methylpterin), para-aminobenzoic acid , and glutamic acid . The folate biosynthesis pathway in micro-organisms can be divided in several parts. The pteridine proportion of folate is made from GTP, that is synthesized in the purine biosynthesis pathway. p-Aminobenzoic acid originates from chorismate and can be synthesized via the same biosynthesis pathways required for the aromatic amino acids, involving glycolysis, pentose phosphate pathway and shikimate pathway. The third component of a folate molecule is glutamate, that is normally taken up from the medium. This review will focus on biosynthesis and folate production by lactic acid bacteria and the folate level production in fermented product.

BIOLOGICAL ACTIVITY OF YERSINIA PSEUDOTUBERCULOSIS TOXINS

Aim. Study of effect of heat-labile (HLT) and thermostable (HST) lethal toxins of Yersinia pseudotuberculosis on the development of embryos of sea urchin Strongylocentrotus intermedius, processes of biosynthesis of nucleic acids and protein in embryo cells and activity of nucleoside-kinases of sea urchin. Materials and methods. Y. pseudotuberculosis strains 2517 (pYV-) and 512 (pYV48MD, pYV82MD) were used for isolation of HLT and HST. Gametes and embryos of sea urchin S. intermedius were used to carry out the experiments and isolate nucleoside-kinases. Results. Both of the studied toxins of Y. pseudotuberculosis possessed spermiotoxic effect and reduced fertilizing ability of sea urchin spermies. HLT LD50 was 1 (ig/ml, and HST - 2 pg/ml. Toxins affected the development of embryos of sea urchin resulting in severe morphologic damages, cessation of the development of embryos at early stages of embryogenesis, destruction of cells and death of embryos. Wherein, damaging effect of HLT was observed at lower concentrations compared with HST. HLT inhibited DNA and RNA biosynthesis at concentrations of 1-2 pg/ml. HST did not affect biosynthesis of nucleic acids even at high concentrations, but inhibited protein biosynthesis in sea urchin embryos. HLT did not reduce the level of inclusion of labeled amino acids into embryo cells. HLT had inhibiting effect on the activity of thymidine- and uridine-kinase of sea urchin, whereas HST did not affect these enzymes. Conclusion. Both of Y. pseudotuberculosis protein toxins affect the development of sea urchin embryos, however, mechanisms of action of HLT and HST on embryos and processes occurring in them differ.