scholarly journals The circadian oscillator analysed at the single‐transcript level

2021 ◽  
Vol 17 (3) ◽  
Author(s):  
Nicholas E Phillips ◽  
Alice Hugues ◽  
Jake Yeung ◽  
Eric Durandau ◽  
Damien Nicolas ◽  
...  
Keyword(s):  
Transcript Level ◽  
Circadian Oscillator ◽  
Single Transcript

scholarly journals Multiplex in situ hybridization within a single transcript: RNAscope reveals dystrophin mRNA dynamics

2019 ◽  
Author(s):  
J.C.W. Hildyard ◽  
F. Rawson ◽  
D.J. Wells ◽  
R.J. Piercy
Keyword(s):  
Transcript Level ◽  
Vital Role ◽  
Dystrophic Muscle ◽  
Transcriptional Dynamics ◽  
Novel Approach ◽  
Highly Sensitive ◽  
Health And Disease ◽  
Single Transcript ◽  
Long Genes

AbstractDystrophin plays a vital role in maintaining muscle health, yet low mRNA expression, lengthy transcription time and the limitations of traditional in-situ hybridization (ISH) methodologies mean that the dynamics of dystrophin transcription remain poorly understood. RNAscope is highly sensitive ISH method that can be multiplexed, allowing detection of individual transcripts at sub-cellular resolution, with different target mRNAs assigned to distinct fluorophores. We present a novel approach, instead using RNAscope probes targeted to 5’ and 3’ regions of the same transcript: labelling muscle dystrophin mRNA in this manner allows transcriptional dynamics to be deciphered in health and disease, resolving both nascent myonuclear transcripts and exported mature mRNAs (the latter absent in dystrophic muscle, yet restored following therapeutic intervention). We show that even in healthy muscle, immature dystrophin mRNA predominates (60-80% of total), with the surprising implication that the half-life of a mature transcript is markedly shorter than the time invested in transcription: at the transcript level, supply may exceed demand. Our findings provide unique spatiotemporal insight into the behaviour of this long transcript (with implications for therapeutic approaches), and further suggests this modified multiplex ISH approach is well-suited to long genes, offering a highly tractable means to reveal complex transcriptional dynamics.

Glioblastoma invasion and NMDA receptors: A novel prospect

2019 ◽  
Vol 106 (3) ◽  
pp. 250-260 ◽  
Author(s):  
DN Nandakumar ◽  
P Ramaswamy ◽  
C Prasad ◽  
D Srinivas ◽  
K Goswami
Keyword(s):  
Glutamate Receptors ◽  
Cell Lines ◽  
Intracellular Signaling ◽  
Transcript Level ◽  
Glioblastoma Cells ◽  
Invasion And Migration ◽  
Matrigel Invasion ◽  
Nmdar Activation ◽  
And Migration

Purpose Glioblastoma cells create glutamate-rich tumor microenvironment, which initiates activation of ion channels and modulates downstream intracellular signaling. N-methyl-D-aspartate receptors (NMDARs; a type of glutamate receptors) have a high affinity for glutamate. The role of NMDAR activation on invasion of glioblastoma cells and the crosstalk with α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs) is yet to be explored. Main methods LN18, U251MG, and patient-derived glioblastoma cells were stimulated with NMDA to activate NMDAR glutamate receptors. The role of NMDAR activation on invasion and migration and its crosstalk with AMPAR were evaluated. Invasion and migration of glioblastoma cells were investigated by in vitro trans-well Matrigel invasion and trans-well migration assays, respectively. Expression of NMDARs and AMPARs at transcript level was evaluated by quantitative real-time polymerase chain reaction. Results We determined that NMDA stimulation leads to enhanced invasion in LN18, U251MG, and patient-derived glioblastoma cells, whereas inhibition of NMDAR using MK-801, a non-competitive antagonist of the NMDAR, significantly decreased the invasive capacity. Concordant with these findings, migration was significantly augmented by NMDAR in both cell lines. Furthermore, NMDA stimulation upregulated the expression of GluN2 and GluA1 subunits at the transcript level. Conclusions This study demonstrated the previously unexplored role of NMDAR in invasion of glioblastoma cells. Furthermore, the expression of the GluN2 subunit of NMDAR and the differential overexpression of the GluA1 subunit of AMPAR in both cell lines provide a plausible rationale of crosstalk between these calcium-permeable subunits in the glutamate-rich microenvironment of glioblastoma.

scholarly journals The Purple Leaf (pl6) Mutation Regulates Leaf Color by Altering the Anthocyanin and Chlorophyll Contents in Rice

Plants ◽  
2020 ◽  
Vol 9 (11) ◽  
pp. 1477
Author(s):  
Asadullah Khan ◽  
Sanaullah Jalil ◽  
Huan Cao ◽  
Yohannes Tsago ◽  
Mustapha Sunusi ◽  
...  
Keyword(s):  
Gamma Ray ◽  
Anthocyanin Biosynthesis ◽  
Light Attenuation ◽  
Transcript Level ◽  
Underlying Mechanism ◽  
Anthocyanin Accumulation ◽  
Morphological Marker ◽  
Chlorophyll Contents ◽  
Rice Mutant ◽  
Purple Coloration

The anthocyanin biosynthesis attracts strong interest due to the potential antioxidant value and as an important morphological marker. However, the underlying mechanism of anthocyanin accumulation in plant tissues is not clearly understood. Here, a rice mutant with a purple color in the leaf blade, named pl6, was developed from wild type (WT), Zhenong 41, with gamma ray treatment. By map-based cloning, the OsPL6 gene was located on the short arm of chromosome 6. The multiple mutations, such as single nucleotide polymorphism (SNP) at −702, −598, −450, an insertion at −119 in the promoter, three SNPs and one 6-bp deletion in the 5′-UTR region, were identified, which could upregulate the expression of OsPL6 to accumulate anthocyanin. Subsequently, the transcript level of structural genes in the anthocyanin biosynthesis pathway, including OsCHS, OsPAL, OsF3H and OsF3′H, was elevated significantly. Histological analysis revealed that the light attenuation feature of anthocyanin has degraded the grana and stroma thylakoids, which resulted in poor photosynthetic efficiency of purple leaves. Despite this, the photoabatement and antioxidative activity of anthocyanin have better equipped the pl6 mutant to minimize the oxidative damage. Moreover, the contents of abscisic acid (ABA) and cytokanin (CK) were elevated along with anthocyanin accumulation in the pl6 mutant. In conclusion, our results demonstrate that activation of OsPL6 could be responsible for the purple coloration in leaves by accumulating excessive anthocyanin and further reveal that anthocyanin acts as a strong antioxidant to scavenge reactive oxygen species (ROS) and thus play an important role in tissue maintenance.

scholarly journals Identification of Susceptibility Genes in Castanea sativa and Their Transcription Dynamics following Pathogen Infection

Plants ◽  
2021 ◽  
Vol 10 (5) ◽  
pp. 913
Author(s):  
Vera Pavese ◽  
Andrea Moglia ◽  
Paolo Gonthier ◽  
Daniela Torello Marinoni ◽  
Emile Cavalet-Giorsa ◽  
...  
Keyword(s):  
Phylogenetic Analyses ◽  
Castanea Sativa ◽  
Transcript Level ◽  
Cryphonectria Parasitica ◽  
Plant Responses ◽  
Functional Domain ◽  
Loss Of Function ◽  
Pathogen Infection ◽  
Domain Identification ◽  
S Genes

Castanea sativa is one of the main multipurpose tree species valued for its timber and nuts. This species is susceptible to two major diseases, ink disease and chestnut blight, caused by Phytophthora spp. and Cryphonectria parasitica, respectively. The loss-of-function mutations of genes required for the onset of pathogenesis, referred to as plant susceptibility (S) genes, are one mechanism of plant resistance against pathogens. On the basis of sequence homology, functional domain identification, and phylogenetic analyses, we report for the first time on the identification of S-genes (mlo1, dmr6, dnd1, and pmr4) in the Castanea genus. The expression dynamics of S-genes were assessed in C. sativa and C. crenata plants inoculated with P. cinnamomi and C. parasitica. Our results highlighted the upregulation of pmr4 and dmr6 in response to pathogen infection. Pmr4 was strongly expressed at early infection phases of both pathogens in C. sativa, whereas in C. crenata, no significant upregulation was observed. The infection of P. cinnamomi led to a higher increase in the transcript level of dmr6 in C. sativa compared to C. crenata-infected samples. For a better understanding of plant responses, the transcript levels of defense genes gluB and chi3 were also analyzed.

scholarly journals The relative abundance of wheat Rubisco activase isoforms is post-transcriptionally regulated

2021 ◽  
Author(s):  
Juan Alejandro Perdomo ◽  
Peter Buchner ◽  
Elizabete Carmo-Silva
Keyword(s):  
Relative Abundance ◽  
Light Availability ◽  
Transcript Level ◽  
Rubisco Activase ◽  
Small Subunit ◽  
Diel Cycle ◽  
Diurnal Rhythms ◽  
Crop Species ◽  
Β Protein ◽  
Wheat Leaves

AbstractDiurnal rhythms and light availability affect transcription–translation feedback loops that regulate the synthesis of photosynthetic proteins. The CO2-fixing enzyme Rubisco is the most abundant protein in the leaves of major crop species and its activity depends on interaction with the molecular chaperone Rubisco activase (Rca). In Triticum aestivum L. (wheat), three Rca isoforms are present that differ in their regulatory properties. Here, we tested the hypothesis that the relative abundance of the redox-sensitive and redox-insensitive Rca isoforms could be differentially regulated throughout light–dark diel cycle in wheat. While TaRca1-β expression was consistently negligible throughout the day, transcript levels of both TaRca2-β and TaRca2-α were higher and increased at the start of the day, with peak levels occurring at the middle of the photoperiod. Abundance of TaRca-β protein was maximal 1.5 h after the peak in TaRca2-β expression, but the abundance of TaRca-α remained constant during the entire photoperiod. The redox-sensitive TaRca-α isoform was less abundant, representing 85% of the redox-insensitive TaRca-β at the transcript level and 12.5% at the protein level. Expression of Rubisco large and small subunit genes did not show a consistent pattern throughout the diel cycle, but the abundance of Rubisco decreased by up to 20% during the dark period in fully expanded wheat leaves. These results, combined with a lack of correlation between transcript and protein abundance for both Rca isoforms and Rubisco throughout the entire diel cycle, suggest that the abundance of these photosynthetic enzymes is post-transcriptionally regulated.

scholarly journals Strategies to Inhibit Hepatitis B Virus at the Transcript Level

Viruses ◽  
2021 ◽  
Vol 13 (7) ◽  
pp. 1327
Author(s):  
Bingqian Qu ◽  
Richard J. P. Brown
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Gene Transcription ◽  
Antigen Expression ◽  
Transcript Level ◽  
Viral Gene ◽  
Small Interfering Rnas ◽  
Viral Gene Transcription ◽  
Hbv Vaccination ◽  
B Virus

Approximately 240 million people are chronically infected with hepatitis B virus (HBV), despite four decades of effective HBV vaccination. During chronic infection, HBV forms two distinct templates responsible for viral transcription: (1) episomal covalently closed circular (ccc)DNA and (2) host genome-integrated viral templates. Multiple ubiquitous and liver-specific transcription factors are recruited onto these templates and modulate viral gene transcription. This review details the latest developments in antivirals that inhibit HBV gene transcription or destabilize viral transcripts. Notably, nuclear receptor agonists exhibit potent inhibition of viral gene transcription from cccDNA. Small molecule inhibitors repress HBV X protein-mediated transcription from cccDNA, while small interfering RNAs and single-stranded oligonucleotides result in transcript degradation from both cccDNA and integrated templates. These antivirals mediate their effects by reducing viral transcripts abundance, some leading to a loss of surface antigen expression, and they can potentially be added to the arsenal of drugs with demonstrable anti-HBV activity. Thus, these candidates deserve special attention for future repurposing or further development as anti-HBV therapeutics.

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