In Vitro Liver Tissue Model Established from Transgenic Mice: Role of HIF-1alpha on Hypoxic Gene Expression

2006 ◽  
Vol 12 (11) ◽  
pp. 3135-3147 ◽  
Author(s):  
Jared W. Allen ◽  
Salman R. Khetani ◽  
Randall S. Johnson ◽  
Sangeeta N. Bhatia
Keyword(s):  
Gene Expression ◽  
Transgenic Mice ◽  
Liver Tissue ◽  
Tissue Model

In VitroLiver Tissue Model Established from Transgenic Mice: Role of HIF1alpha on Hypoxic Gene Expression

2006 ◽  
Vol 0 (0) ◽  
pp. 061004065151004
Author(s):  
Jared W. Allen ◽  
Salman R. Khetani ◽  
Randall S. Johnson ◽  
Sangeeta N. Bhatia
Keyword(s):  
Gene Expression ◽  
Transgenic Mice ◽  
Tissue Model

Notochord to endoderm signaling is required for pancreas development

Development ◽  
1997 ◽  
Vol 124 (21) ◽  
pp. 4243-4252 ◽  
Author(s):  
S.K. Kim ◽  
M. Hebrok ◽  
D.A. Melton
Keyword(s):  
Gene Expression ◽  
Developmental Stage ◽  
Pancreas Development ◽  
Chick Embryos ◽  
Carboxypeptidase A ◽  
Bud Development ◽  
Dorsal Pancreas ◽  
Stepwise Manner

The role of the notochord in inducing and patterning adjacent neural and mesodermal tissues is well established. We provide evidence that the notochord is also required for one of the earliest known steps in the development of the pancreas, an endodermally derived organ. At a developmental stage in chick embryos when the notochord touches the endoderm, removal of notochord eliminates subsequent expression of several markers of dorsal pancreas bud development, including insulin, glucagon and carboxypeptidase A. Pancreatic gene expression can be initiated and maintained in prepancreatic chick endoderm grown in vitro with notochord. Non-pancreatic endoderm, however, does not express pancreatic genes when recombined with the same notochord. The results suggest that the notochord provides a permissive signal to endoderm to specify pancreatic fate in a stepwise manner.

scholarly journals Postrecruitment Function of Yeast Med6 Protein during the Transcriptional Activation by Mediator Complex

2018 ◽  
Vol 2018 ◽  
pp. 1-9
Author(s):  
Gwang Sik Kim ◽  
Young Chul Lee
Keyword(s):  
Gene Expression ◽  
Reporter Gene ◽  
Transcriptional Activation ◽  
Reporter Gene Expression ◽  
Temperature Sensitive ◽  
Internal Deletion ◽  
Cell Extracts ◽  
Evolutionarily Conserved

Med6 protein (Med6p) is a hallmark component of evolutionarily conserved Mediator complexes, and the genuine role of Med6p in Mediator functions remains elusive. For the functional analysis ofSaccharomyces cerevisiaeMed6p (scMed6p), we generated a series of scMed6p mutants harboring a small internal deletion. Genetic analysis of these mutants revealed that three regions (amino acids 33–42 (Δ2), 125–134 (Δ5), and 157–166 (Δ6)) of scMed6p are required for cell viability and are located at highly conserved regions of Med6 homologs. Notably, the Med6p-Δ2 mutant was barely detectable in whole-cell extracts and purified Mediator, suggesting a loss of Mediator association and concurrent rapid degradation. Consistent with this, the recombinant forms of Med6p having these mutations partially (Δ2) restore or fail (Δ5 and Δ6) to restore in vitro transcriptional defects caused by temperature-sensitivemed6mutation. In an artificial recruitment assay, Mediator containing a LexA-fused wild-type Med6p or Med6p-Δ5 was recruited to thelexAoperator region with TBP and activated reporter gene expression. However, the recruitment of Mediator containing LexA-Med6p-Δ6 tolexAoperator region resulted in neither TBP recruitment nor reporter gene expression. This result demonstrates a pivotal role of Med6p in the postrecruitment function of Mediator, which is essential for transcriptional activation by Mediator.

scholarly journals Differential role of cAMP, cGMP and Ca2+ and involvement of kinases and CBP-CREB-CRE pathway in regulation of arylalkylamine N-acetyltransferase 2 gene in the pineal organ of air-breathing catfish, Clarias gariepinus

2021 ◽  
Author(s):  
Hijam Nonibala ◽  
Braj Bansh Prasad Gupta
Keyword(s):  
Gene Expression ◽  
Map Kinase ◽  
Gene Transcription ◽  
Pineal Organ ◽  
Clarias Gariepinus ◽  
Specific Inhibitor ◽  
P38 Map Kinase ◽  
Camp And Cgmp

Abstract Transcription of arylalkylamine N-acetyltransferase 2 (aanat2) gene leads to formation of AANAT2 - the rate-limiting enzyme in melatonin synthesis pathway in photosensitive fish pineal organ. However, unlike in avian and mammalian pineal gland, there is practically no information on signal transduction pathway(s) involved in regulation of aanat2 gene transcription in the fish pineal organ. Therefore, we investigated the role of important molecular components of signalling via cAMP, cGMP, Ca2+ involving PKA, PKG, PKC, MeK and p38 MAP kinase as well as possible role of serine/threonine phosphatases, CREB and CBP using their specific inhibitors and/or activators in aanat2 gene transcription in the fish pineal organ maintained under in vitro culture-conditions. db-cAMP and db-cGMP stimulated the expression of aanat2 gene. db-cAMP- and cGMP-induced aanat2 gene expression was significantly reduced in the presence of H-89 (specific inhibitor of PKA), KT5823 (specific inhibitor of PKG), chelerythrine chloride (specific inhibitor of PKC), U0126 ethanolate (specific inhibitor of MeK) and SB 202190 monohydrochloride hydrate (specific inhibitor of p38 MAP kinase). Inhibitors of PP1 and PP2A significantly increased aanat2 gene expression as well as significantly reduced cAMP- and cGMP-induced gene transcription, while inhibitor of PP2B had no effect on aanat2 gene expression. Inhibitors of both CREB and CBP-CREB interaction completely blocked cAMP-induced aanat2 gene transcription. Based on these findings, we suggest that cAMP, cGMP and Ca2+ stimulate aanat2 gene transcription via PKA, PKG and PKC, respectively. Further, protein phosphatases and CBP-CREB-CRE pathway are actively involved in regulation of on aanat2 gene expression in the fish pineal organ.

scholarly journals The Mycobacterium tuberculosis sRNA F6 regulates expression of groEL/S

2020 ◽  
Author(s):  
Joanna Houghton ◽  
Angela Rodgers ◽  
Graham Rose ◽  
Kristine B. Arnvig
Keyword(s):  
Gene Expression ◽  
Mycobacterium Tuberculosis ◽  
Small Rnas ◽  
Transcriptional Control ◽  
Cold Shock ◽  
Control Of Gene Expression ◽  
Rna Biology ◽  
Mouse Model Of Infection

ABSTRACTAlmost 140 years after the identification of Mycobacterium tuberculosis as the etiological agent of tuberculosis, important aspects of its biology remain poorly described. Little is known about the role of post-transcriptional control of gene expression and RNA biology, including the role of most of the small RNAs (sRNAs) identified to date. We have carried out a detailed investigation of the M. tuberculosis sRNA, F6, and show it to be dependent on SigF for expression and significantly induced during in vitro starvation and in a mouse model of infection. However, we found no evidence of attenuation of a ΔF6 strain within the first 20 weeks of infection. A further exploration of F6 using in vitro models of infection suggests a role for F6 as a highly specific regulator of the heat shock repressor, HrcA. Our results point towards a role for F6 during periods of low metabolic activity similar to cold shock and associated with nutrient starvation such as that found in human granulomas in later stages of infection.

P–181 Morphine regulates BMP4 growth factor and is involved in in-vitro early embryo development and PGCs formation

2021 ◽  
Vol 36 (Supplement_1) ◽  
Author(s):  
I Muñoa ◽  
M Araolaza-Lasa ◽  
I Urizar-Arenaza ◽  
M Gianzo Citores ◽  
N Subiran Ciudad
Keyword(s):  
Gene Expression ◽  
Growth Factor ◽  
Environmental Factors ◽  
Embryo Development ◽  
Early Embryo ◽  
Epigenetic Changes ◽  
Morphine Treatment ◽  
Early Embryo Development

Abstract Study question To elucidate if morphine can alter embryo development. Summary answer Chronic morphine treatment regulates BMP4 growth factor, in terms of gene expression and H3K27me3 enrichment and promotes in-vitro blastocysts development and PGC formation. What is known already BMP4 is a member of the bone morphogenetic protein family, which acts mainly through SMAD dependent pathway, to play an important role in early embryo development. Indeed, BMP4 enhances pluripotency in mouse embryonic stem cells (mESCs) and, specifically, is involved in blastocysts formation and primordial germ cells (PGCs) generation. Although, external morphine influence has been previously reported on the early embryo development, focus on implantation and uterus function, there is a big concern in understanding how environmental factors can cause stable epigenetic changes, which could be maintained during development and lead to health problems. Study design, size, duration First, OCT4-reported mESCs were chronically treated with morphine during 24h, 10–5mM. After morphine removal, mESCs were collected for RNA-seq and H3K27me3 ChIP-seq study. To elucidate the role of morphine in early embryo development, two cell- embryos stage were chronically treated with morphine for 24h and in-vitro cultured up to the blastocyst stage in the absence of morphine. Furthermore, after morphine treatment mESCs were differentiated to PGCs, to elucidate the role of morphine in PGC differentiation. Participants/materials, setting, methods Transcriptomic analyses and H3K27me3 genome wide distribution were carried out by RNA-Sequencing and Chip-Sequencing respectively. Validations were performed by RNA-RT-qPCR and Chip-RT-qPCR. Main results and the role of chance Dynamic transcriptional analyses identified a total of 932 differentially expressed genes (DEGs) after morphine treatment on mESCs, providing strong evidence of a transcriptional epigenetic effect induced by morphine. High-throughput screening approaches showed up Bmp4 as one of the main morphine targets on mESCs. Morphine caused an up-regulation of Bmp4 gene expression together with a decrease of H3K27me3 enrichment at promoter level. However, no significant differences were observed on gene expression and H3K27me3 enrichment on BMP4 signaling pathway components (such as Smad1, Smad4, Smad5, Smad7, Prdm1 and Prmd14) after morphine treatment. On the other hand, the Bmp4 gene expression was also up-regulated in in-vitro morphine treated blastocyst and in-vitro morphine treated PGCs. These results were consistent with the increase in blastocyst rate and PGC transformation rate observed after morphine chronic treatment. Limitations, reasons for caution To perform the in-vitro analysis. Further studies are needed to describe the whole signaling pathways underlying BMP4 epigenetic regulation after morphine treatment. Wider implications of the findings: Our findings confirmed that mESCs and two-cell embryos are able to memorize morphine exposure and promote both blastocyst development and PGCs formation through potentially BMP4 epigenetic regulation. These results provide insights understanding how environmental factors can cause epigenetic changes during the embryo development, leading to alterations and producing health problems/diseases Trial registration number Not applicable

Construction and optimization of herpes simplex virus vectors for central nervous system gene delivery based on CRISPR/Cas9-mediated genome editing

2021 ◽  
Vol 21 ◽  
Author(s):  
Xinwei Huang ◽  
Xiuqing Li ◽  
Lijuan Yang ◽  
Pengfei Wang ◽  
Jingyuan Yan ◽  
...  
Keyword(s):  
Gene Expression ◽  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Transgene Expression ◽  
Mouse Hippocampus ◽  
Simplex Virus ◽  
Hsv 1

Aims: We aim to define parameters affecting the safety and long-term transgene expression of attenuated HSV-1 vectors and optimize the expression cassettes to achieve robust and sustained expression in CNS. Background: Engineered, attenuated Herpes simplex virus (HSV) vectors are promising vehicles for gene delivery to the peripheral and central nervous systems. The virus latent promoter (LAP) is commonly used to drive exogenous gene expression; however, parameters affecting the safety and long-term transgene expression of attenuated HSV-1 vectors have not been fully understood. Objective: This study aimed to construct attenuated HSV-1 vectors using the CRISPR-Cas9 system and examine the influence of transgene cassette construction and insertion site on transgene expression and vector safety. Method: In this study, we used a CRISPR-Cas9 system to accurately and efficiently edit attenuated HSV-1 strain 1716, and constructed two series of recombinant virus LMR and LMRx with different sets of gene cassettes insertion in Exon1(LAP2) and 2.0 kb intron downstream of LAP, respectively. The transgene expression and viral gene transcriptional kinetics were compared in in-vitro cell lines. The reporter gene expression and safety profiles of each vector were further evaluated in the mouse hippocampus gene transduction model. Result: The in-vitro cell line analysis indicated that the insertion of a gene expression cassette would disrupt virus gene transcription. Mouse hippocampus transducing analysis suggested that complete expression cassette insertion at 2.0 kb intron could achieve robust and longtime gene expression than the other constructs. Recombinants with gene expression cassettes lacked Poly (A), which induced significant neuronal inflammation due to persistent viral antigen expression and microglia activation. Conclusion: Our results indicated that the integrity of LAT transcripts was not necessary for the establishment of long-term latent expression. Exogenous strong promoters (like cBh promoter) could remain active during latency when placed in Exon1 or 2.0 Kb Intron of LAT locus, although their transcriptional activity declined with time. Consistent with previous research, the foreign gene expression would last much longer when the gene cassette was located downstream of Exon1, which suggested a role of LAP2 in maintaining promoter activity during latency. Besides, over-transcription of the downstream part of LAT may induce continuous activation of the attenuated vectors, suggesting an important role of LAT in maintaining viral reactivation potential.

scholarly journals Increased Gene Expression of RUNX2 and SOX9 in Mesenchymal Circulating Progenitors Is Associated with Autophagy during Physical Activity

2019 ◽  
Vol 2019 ◽  
pp. 1-14 ◽  
Author(s):  
L. Dalle Carbonare ◽  
M. Mottes ◽  
S. Cheri ◽  
M. Deiana ◽  
F. Zamboni ◽  
...  
Keyword(s):  
Gene Expression ◽  
Physical Activity ◽  
Physical Exercise ◽  
Cellular Functions ◽  
Enhanced Expression ◽  
Chondrocytic Differentiation ◽  
Marathon Performance ◽  
Array Analyses

Lack of physical exercise is considered an important risk factor for chronic diseases. On the contrary, physical exercise reduces the morbidity rates of obesity, diabetes, bone disease, and hypertension. In order to gain novel molecular and cellular clues, we analyzed the effects of physical exercise on differentiation of mesenchymal circulating progenitor cells (M-CPCs) obtained from runners. We also investigated autophagy and telomerase-related gene expression to evaluate the involvement of specific cellular functions in the differentiation process. We performed cellular and molecular analyses in M-CPCs, obtained by a depletion method, of 22 subjects before (PRE RUN) and after (POST RUN) a half marathon performance. In order to prove our findings, we performed also in vitro analyses by testing the effects of runners’ sera on a human bone marrow-derived mesenchymal stem (hBM-MSC) cell line. PCR array analyses of PRE RUN versus POST RUN M-CPC total RNAs put in evidence several genes which appeared to be modulated by physical activity. Our results showed that physical exercise promotes differentiation. Osteogenesis-related genes as RUNX2, MSX1, and SPP1 appeared to be upregulated after the run; data showed also increased levels of BMP2 and BMP6 expressions. SOX9, COL2A1, and COMP gene enhanced expression suggested the induction of chondrocytic differentiation as well. The expression of telomerase-associated genes and of two autophagy-related genes, ATG3 and ULK1, was also affected and correlated positively with MSC differentiation. These data highlight an attractive cellular scenario, outlining the role of autophagic response to physical exercise and suggesting new insights into the benefits of physical exercise in counteracting chronic degenerative conditions.

scholarly journals PRMT5 Is Required for Bovine Leukemia Virus Infection In Vivo and Regulates BLV Gene Expression, Syncytium Formation, and Glycosylation In Vitro

Viruses ◽  
2020 ◽  
Vol 12 (6) ◽  
pp. 650 ◽  
Author(s):  
Wlaa Assi ◽  
Tomoya Hirose ◽  
Satoshi Wada ◽  
Ryosuke Matsuura ◽  
Shin-nosuke Takeshima ◽  
...  
Keyword(s):  
Gene Expression ◽  
Small Molecule ◽  
Bovine Leukemia Virus ◽  
Proviral Load ◽  
Leukemia Virus ◽  
Syncytium Formation ◽  
High Proviral Load

Bovine leukemia virus (BLV) is the causative agent of enzootic bovine leukosis, which is the most common neoplastic disease of cattle and is closely related to human T-cell leukemia viruses. We investigated the role of a new host protein, PRMT5, in BLV infection. We found that PRMT5 is overexpressed only in BLV-infected cattle with a high proviral load, but not in those with a low proviral load. Furthermore, this upregulation continued to the lymphoma stage. PRMT5 expression was upregulated in response to experimental BLV infection; moreover, PRMT5 upregulation began in an early stage of BLV infection rather than after a long period of proviral latency. Second, siRNA-mediated PRMT5 knockdown enhanced BLV gene expression at the transcript and protein levels. Additionally, a selective small-molecule inhibitor of PRMT5 (CMP5) enhanced BLV gene expression. Interestingly, CMP5 treatment, but not siRNA knockdown, altered the gp51 glycosylation pattern and increased the molecular weight of gp51, thereby decreasing BLV-induced syncytium formation. This was supported by the observation that CMP5 treatment enhanced the formation of the complex type of N-glycan more than the high mannose type. In conclusion, PRMT5 overexpression is related to the development of BLV infection with a high proviral load and lymphoma stage and PRMT5 inhibition enhances BLV gene expression. This is the first study to investigate the role of PRMT5 in BLV infection in vivo and in vitro and to reveal a novel function for a small-molecule compound in BLV-gp51 glycosylation processing.

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