Mechanism of autonomous synchronization of the circadian KaiABC rhythm

AbstractThe cyanobacterial circadian clock can be reconstituted by mixing three proteins, KaiA, KaiB, and KaiC, in vitro. In this protein mixture, oscillations of the phosphorylation level of KaiC molecules are synchronized to show the coherent oscillations of the ensemble of many molecules. However, the molecular mechanism of this synchronization has not yet been fully elucidated. In this paper, we explain a theoretical model that considers the multifold feedback relations among the structure and reactions of KaiC. The simulated KaiC hexamers show stochastic switch-like transitions at the level of single molecules, which are synchronized in the ensemble through the sequestration of KaiA into the KaiC–KaiB–KaiA complexes. The proposed mechanism quantitatively reproduces the synchronization that was observed by mixing two solutions oscillating in different phases. The model results suggest that biochemical assays with varying concentrations of KaiA or KaiB can be used to test this hypothesis.

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Mechanism of autonomous synchronization of the circadian KaiABC rhythm

10.1101/2020.11.24.395566 ◽

2020 ◽

Author(s):

Masaki Sasai

Keyword(s):

Theoretical Model ◽

Circadian Clock ◽

Single Molecules ◽

Protein Mixture ◽

Phosphorylation Level ◽

Biochemical Assays ◽

Oscillating Solutions

ABSTRACTThe cyanobacterial circadian clock can be reconstituted by mixing three proteins, KaiA, KaiB, and KaiC, in vitro. In this protein mixture, oscillations of the phosphorylation level of KaiC molecules are synchronized to show the coherent oscillations of the ensemble of many molecules. However, the mechanism of this synchronization remains elusive. In this paper, we explain a theoretical model that considers the multifold feedback relations among the structure and reactions of KaiC. The simulated KaiC hexamers show stochastic switch-like transitions at the level of single molecules, which are synchronized in the ensemble through the sequestration of KaiA into the KaiC-KaiB-KaiA complexes. The proposed mechanism quantitatively reproduces the synchronization that was observed by mixing two oscillating solutions in different phases. The model results suggest that biochemical assays with varying concentrations of KaiA or KaiB can be used to test this hypothesis.

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ACIS, A Novel KepTide™, Binds to ACE-2 Receptor and Inhibits the Infection of SARS-CoV2 Virus in vitro in Primate Kidney Cells: Therapeutic Implications for COVID-19 (Preprint)

10.2196/preprints.25089 ◽

2020 ◽

Author(s):

Avik Sotira Scientific

Keyword(s):

Social Life ◽

Plaque Assay ◽

Host Cells ◽

Surface Glycoprotein ◽

Crystallographic Structure ◽

Therapeutic Implications ◽

Biochemical Assays ◽

Targeted Medicine

UNSTRUCTURED Coronavirus disease 2019 (COVID-19) is a severe acute respiratory syndrome (SARS) caused by a virus known as SARS-Coronavirus 2 (SARS-CoV2). Without a targeted-medicine, this disease has been causing a massive humanitarian crisis not only in terms of mortality, but also imposing a lasting damage to social life and economic progress of humankind. Therefore, an immediate therapeutic strategy needs to be intervened to mitigate this global crisis. Here, we report a novel KepTide™ (Knock-End Peptide) therapy that nullifies SARS-CoV2 infection. SARS-CoV2 employs its surface glycoprotein “spike” (S-glycoprotein) to interact with angiotensin converting enzyme-2 (ACE-2) receptor for its infection in host cells. Based on our in-silico-based homology modeling study validated with a recent X-ray crystallographic structure (PDB ID:6M0J), we have identified that a conserved motif of S-glycoprotein that intimately engages multiple hydrogen-bond (H-bond) interactions with ACE-2 enzyme. Accordingly, we designed a peptide, termed as ACIS (ACE-2 Inhibitory motif of Spike), that displayed significant affinity towards ACE-2 enzyme as confirmed by biochemical assays such as BLItz and fluorescence polarization assays. Interestingly, more than one biochemical modifications were adopted in ACIS in order to enhance the inhibitory action of ACIS and hence called as KEpTide™. Consequently, a monolayer invasion assay, plaque assay and dual immunofluorescence analysis further revealed that KEpTide™ efficiently mitigated the infection of SARS-CoV2 in vitro in VERO E6 cells. Finally, evaluating the relative abundance of ACIS in lungs and the potential side-effects in vivo in mice, our current study discovers a novel KepTide™ therapy that is safe, stable, and robust to attenuate the infection of SARS-CoV2 virus if administered intranasally. INTERNATIONAL REGISTERED REPORT RR2-https://doi.org/10.1101/2020.10.13.337584

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Formulation and in vitro evaluation of self-nanoemulsifying liquisolid tablets of furosemide

Scientific Reports ◽

10.1038/s41598-020-79940-5 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Lena Dalal ◽

Abdul Wahab Allaf ◽

Hind El-Zein

Keyword(s):

Theoretical Model ◽

Castor Oil ◽

In Vivo Studies ◽

Coating Materials ◽

Peg 400 ◽

The Mean ◽

Usp Apparatus ◽

Solid System

AbstractSelf-nanoemulsifying drug delivery systems (SNEDDS) were used to enhance the dissolution rate of furosemide as a model for class IV drugs and the system was solidified into liquisolid tablets. SNEDDS of furosemide contained 10% Castor oil, 60% Cremophor EL, and 30% PEG 400. The mean droplets size was 17.9 ± 4.5 nm. The theoretical model was used to calculate the amounts of the carrier (Avicel PH101) and coating materials (Aerosil 200) to prepare liquisolid powder. Carrier/coating materials ratio of 5/1 was used and Ludipress was added to the solid system, thus tablets with hardness of 45 ± 2 N were obtained. Liquisolid tablets showed 2-folds increase in drug release as compared to the generic tablets after 60 min in HCl 0.1 N using USP apparatus-II. Furosemide loaded SNEDDS tablets have great prospects for further in vivo studies, and the theoretical model is useful for calculating the adequate amounts of adsorbents required to solidify these systems.

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miR-29a sensitizes the response of glioma cells to temozolomide by modulating the P53/MDM2 feedback loop

Cellular & Molecular Biology Letters ◽

10.1186/s11658-021-00266-9 ◽

2021 ◽

Vol 26 (1) ◽

Author(s):

Qiudan Chen ◽

Weifeng Wang ◽

Shuying Chen ◽

Xiaotong Chen ◽

Yong Lin

Keyword(s):

Molecular Mechanism ◽

Feedback Loop ◽

Glioma Cells ◽

Luciferase Reporter ◽

Migration And Invasion ◽

Transcriptional Level ◽

Aberrant Expression ◽

Altered Expression ◽

P53 Signaling

AbstractRecently, pivotal functions of miRNAs in regulating common tumorigenic processes and manipulating signaling pathways in brain tumors have been recognized; notably, miR‐29a is closely associated with p53 signaling, contributing to the development of glioma. However, the molecular mechanism of the interaction between miR-29a and p53 signaling is still to be revealed. Herein, a total of 30 glioma tissues and 10 non-cancerous tissues were used to investigate the expression of miR‐29a. CCK-8 assay and Transwell assay were applied to identify the effects of miR-29a altered expression on the malignant biological behaviors of glioma cells in vitro, including proliferation, apoptosis, migration and invasion. A dual-luciferase reporter assay was used to further validate the regulatory effect of p53 or miR-29a on miR-29a or MDM2, respectively, at the transcriptional level. The results showed that miR-29a expression negatively correlated with tumor grade of human gliomas; at the same time it inhibited cell proliferation, migration, and invasion and promoted apoptosis of glioma cells in vitro. Mechanistically, miR-29a expression was induced by p53, leading to aberrant expression of MDM2 targeted by miR-29a, and finally imbalanced the activity of the p53-miR-29a-MDM2 feedback loop. Moreover, miR-29a regulating p53/MDM2 signaling sensitized the response of glioma cells to temozolomide treatment. Altogether, the study demonstrated a potential molecular mechanism in the tumorigenesis of glioma, while offering a possible target for treating human glioma in the future.

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PDGFRα: Expression and Function during Mitral Valve Morphogenesis

Journal of Cardiovascular Development and Disease ◽

10.3390/jcdd8030028 ◽

2021 ◽

Vol 8 (3) ◽

pp. 28

Author(s):

Kelsey Moore ◽

Diana Fulmer ◽

Lilong Guo ◽

Natalie Koren ◽

Janiece Glover ◽

...

Keyword(s):

Mitral Valve ◽

Primary Cilia ◽

Growth Factor Receptor ◽

Akt Phosphorylation ◽

Valve Development ◽

Biochemical Assays ◽

Factor Receptor Alpha ◽

Mesenchymal Transformation ◽

And Function

Mitral valve prolapse (MVP) is a common form of valve disease and can lead to serious secondary complications. The recent identification of MVP causal mutations in primary cilia-related genes has prompted the investigation of cilia-mediated mechanisms of disease inception. Here, we investigate the role of platelet-derived growth factor receptor-alpha (PDGFRα), a receptor known to be present on the primary cilium, during valve development using genetically modified mice, biochemical assays, and high-resolution microscopy. While PDGFRα is expressed throughout the ciliated valve interstitium early in development, its expression becomes restricted on the valve endocardium by birth and through adulthood. Conditional ablation of Pdgfra with Nfatc1-enhancer Cre led to significantly enlarged and hypercellular anterior leaflets with disrupted endothelial adhesions, activated ERK1/2, and a dysregulated extracellular matrix. In vitro culture experiments confirmed a role in suppressing ERK1/2 activation while promoting AKT phosphorylation. These data suggest that PDGFRα functions to suppress mesenchymal transformation and disease phenotypes by stabilizing the valve endocardium through an AKT/ERK pathway.

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Molecular mechanism of in vitro 30 S ribosome assembly. II. Conformational changes of ribosomal proteins.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(18)36004-6 ◽

1979 ◽

Vol 254 (16) ◽

pp. 7712-7716

Author(s):

J.M. Dunn ◽

K.P. Wong

Keyword(s):

Molecular Mechanism ◽

Conformational Changes ◽

Ribosomal Proteins ◽

Ribosome Assembly

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Synthesis and Activity of New Schiff Bases of Furocoumarin

Heterocyclic Communications ◽

10.1515/hc-2020-0115 ◽

2020 ◽

Vol 26 (1) ◽

pp. 176-184

Author(s):

Huijun Xie ◽

Chao Niu ◽

Zeyang Chao ◽

Nuramina Mamat ◽

Haji Akber Aisa

Keyword(s):

Animal Models ◽

Amphotericin B ◽

Molecular Mechanism ◽

Schiff Bases ◽

Positive Control ◽

Excellent Performance ◽

Activation Rate ◽

Antibacterial Spectrum ◽

Better Than

AbstractFurocoumarins, such as 8-MOP, are the most common medications used to relieve the symptoms of vitiligo clinically. Some furocoumarins also showed excellent performance in an anti-bacterial assay. This paper describes the synthesis of a series of novel Schiff bases (6a-6k), and their promotion in melanogenesis and anti-bacterial properties were studied in vitro.The pigment production of B16 cells and bacterial inhibition ring assay were applied for the bioactivity of 6a-6k. According to the results, a stronger promotion on pigment content was observed, when six compounds co-cultured with cells, compared with positive control (8-MOP). Significantly, compound 6k (237%) as the most active was found to increase the amount of melanin more than 1.7 times compared with 8-MOP activation rate (136%). All the compounds could moderately retard C. albicans growth. Interestingly, aldehyde 5, which possessed a broader antibacterial spectrum, showed the highest inhibition against C. albicans as well and much better than the positive control (Amphotericin B).Studies of 6k in animal models of vitiligo and related molecular mechanism are presently under way, with the aim of discovering an anti-vitiligo leading compound.

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Circadian clock control of eIF2α phosphorylation is necessary for rhythmic translation initiation

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1918459117 ◽

2020 ◽

Vol 117 (20) ◽

pp. 10935-10945 ◽

Cited By ~ 1

Author(s):

Shanta Karki ◽

Kathrina Castillo ◽

Zhaolan Ding ◽

Olivia Kerr ◽

Teresa M. Lamb ◽

...

Keyword(s):

Circadian Clock ◽

Translation Initiation ◽

Translation Initiation Factor ◽

Initiation Factor ◽

Nutrient Metabolism ◽

Eif2α Phosphorylation ◽

Eif2α Kinase ◽

The Impact

The circadian clock in eukaryotes controls transcriptional and posttranscriptional events, including regulation of the levels and phosphorylation state of translation factors. However, the mechanisms underlying clock control of translation initiation, and the impact of this potential regulation on rhythmic protein synthesis, were not known. We show that inhibitory phosphorylation of eIF2α (P-eIF2α), a conserved translation initiation factor, is clock controlled in Neurospora crassa, peaking during the subjective day. Cycling P-eIF2α levels required rhythmic activation of the eIF2α kinase CPC-3 (the homolog of yeast and mammalian GCN2), and rhythmic activation of CPC-3 was abolished under conditions in which the levels of charged tRNAs were altered. Clock-controlled accumulation of P-eIF2α led to reduced translation during the day in vitro and was necessary for the rhythmic synthesis of select proteins in vivo. Finally, loss of rhythmic P-eIF2α levels led to reduced linear growth rates, supporting the idea that partitioning translation to specific times of day provides a growth advantage to the organism. Together, these results reveal a fundamental mechanism by which the clock regulates rhythmic protein production, and provide key insights into how rhythmic translation, cellular energy, stress, and nutrient metabolism are linked through the levels of charged versus uncharged tRNAs.

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