scholarly journals Genetic Control of MAP3K1 in Eye Development and Sex Differentiation

Cells ◽  
2021 ◽  
Vol 11 (1) ◽  
pp. 34
Author(s):  
Jingjing Wang ◽  
Eiki Kimura ◽  
Maureen Mongan ◽  
Ying Xia
Keyword(s):  
Biochemical Characterization ◽  
Eye Development ◽  
Sex Differentiation ◽  
Kinase Domain ◽  
Loss Of Function ◽  
Disease Etiology ◽  
Developmental Models ◽  
Environmental Signals ◽  
Mapk Cascades

The MAP3K1 is responsible for transmitting signals to activate specific MAP2K-MAPK cascades. Following the initial biochemical characterization, genetic mouse models have taken center stage to elucidate how MAP3K1 regulates biological functions. To that end, mice were generated with the ablation of the entire Map3k1 gene, the kinase domain coding sequences, or ubiquitin ligase domain mutations. Analyses of the mutants identify diverse roles that MAP3K1 plays in embryonic survival, maturation of T/B cells, and development of sensory organs, including eye and ear. Specifically in eye development, Map3k1 loss-of-function was found to be autosomal recessive for congenital eye abnormalities, but became autosomal dominant in combination with Jnk and RhoA mutations. Additionally, Map3k1 mutation increased eye defects with an exposure to environmental agents such as dioxin. Data from eye developmental models reveal the nexus role of MAP3K1 in integrating genetic and environmental signals to control developmental activities. Here, we focus the discussions on recent advances in understanding the signaling mechanisms of MAP3K1 in eye development in mice and in sex differentiation from human genomics findings. The research works featured here lead to a deeper understanding of the in vivo signaling network, the mechanisms of gene–environment interactions, and the relevance of this multifaceted protein kinase in disease etiology and pathogenesis.

scholarly journals The Human FSGS-Causing ANLN R431C Mutation Induces Dysregulated PI3K/AKT/mTOR/Rac1 Signaling in Podocytes

2018 ◽  
Vol 29 (8) ◽  
pp. 2110-2122 ◽  
Author(s):  
Gentzon Hall ◽  
Brandon M. Lane ◽  
Kamal Khan ◽  
Igor Pediaditakis ◽  
Jianqiu Xiao ◽  
...  
Keyword(s):  
Biochemical Characterization ◽  
Adaptor Protein ◽  
Pi3k Pathway ◽  
Actin Binding ◽  
Loss Of Function ◽  
Signaling Axis ◽  
Phosphoinositide 3 Kinase

BackgroundWe previously reported that mutations in the anillin (ANLN) gene cause familial forms of FSGS. ANLN is an F-actin binding protein that modulates podocyte cell motility and interacts with the phosphoinositide 3-kinase (PI3K) pathway through the slit diaphragm adaptor protein CD2-associated protein (CD2AP). However, it is unclear how the ANLN mutations cause the FSGS phenotype. We hypothesized that the R431C mutation exerts its pathogenic effects by uncoupling ANLN from CD2AP.MethodsWe conducted in vivo complementation assays in zebrafish to determine the effect of the previously identified missense ANLN variants, ANLNR431C and ANLNG618C during development. We also performed in vitro functional assays using human podocyte cell lines stably expressing wild-type ANLN (ANLNWT) or ANLNR431C.ResultsExperiments in anln-deficient zebrafish embryos showed a loss-of-function effect for each ANLN variant. In human podocyte lines, expression of ANLNR431C increased cell migration, proliferation, and apoptosis. Biochemical characterization of ANLNR431C-expressing podocytes revealed hyperactivation of the PI3K/AKT/mTOR/p70S6K/Rac1 signaling axis and activation of mTOR-driven endoplasmic reticulum stress in ANLNR431C-expressing podocytes. Inhibition of mTOR, GSK-3β, Rac1, or calcineurin ameliorated the effects of ANLNR431C. Additionally, inhibition of the calcineurin/NFAT pathway reduced the expression of endogenous ANLN and mTOR.ConclusionsThe ANLNR431C mutation causes multiple derangements in podocyte function through hyperactivation of PI3K/AKT/mTOR/p70S6K/Rac1 signaling. Our findings suggest that the benefits of calcineurin inhibition in FSGS may be due, in part, to the suppression of ANLN and mTOR. Moreover, these studies illustrate that rational therapeutic targets for familial FSGS can be identified through biochemical characterization of dysregulated podocyte phenotypes.

scholarly journals A point mutation in the catalytic domain of c-kit induces growth factor independence, tumorigenicity, and differentiation of mast cells

Blood ◽  
1996 ◽  
Vol 87 (8) ◽  
pp. 3117-3123 ◽  
Author(s):  
X Piao ◽  
A Bernstein
Keyword(s):  
Mast Cell ◽  
Point Mutation ◽  
Kinase Domain ◽  
Loss Of Function ◽  
Wild Type ◽  
Steel Factor ◽  
Kit Receptor ◽  
Cytoplasmic Kinase Domain

The murine W and Steel loci encode the Kit receptor tyrosine kinase and its ligand, Steel factor, respectively. Loss of function mutations at either the W or Sl loci lead to a variety of pleiotropic developmental defects, including mast cell deficiency and severe macrocytic anemia. In addition to these loss-of-function mutations, gain-of-function mutations in c-kit, leading to constitutive activation of the Kit receptor, have also been identified in both rodent and human mastocytomas. In this study, we have examined the transforming potential and biologic effects of a point mutation that results in substitution of the aspartic acid at codon 814 in the cytoplasmic kinase domain to tyrosine (D814Y) by introducing either wild-type (Kit) or mutant KitD814Y (KDY) cDNA into an interleukin-3-dependent mast cell line IC2. Stimulation of cells expressing the wild-type Kit receptor (IC2/Kit) with Steel factor in vitro resulted in a short-term growth response, whereas IC2/KDY cells were capable of sustained proliferation in a ligand-independent manner. In addition, expression of KDY resulted in the oncogenic transformation of IC2 cells, as determined by colony formation in vitro in the absence of exogenous growth factors and the formation of mastocytomas in vivo in syngeneic DBA/2 mice. Surprisingly, KDY expression in IC2 cells triggered dramatic changes in cell size and the extent of granulation. In addition, KDY induced the expression of mouse mast cell protease-4 (MMCP-4) and MMCP-6. In contrast, neither of these molecular or cellular changes was observed in IC2/Kit cells treated with Steel factor. These results show that the D814Y mutation in the cytoplasmic kinase domain of the Kit receptor induces ligand-independent mast cell growth in vitro, tumorigenicity in vivo, and mast cell differentiation.

Brassinosteroid-independent function of BRI1/CLV1 chimeric receptors

2006 ◽  
Vol 33 (8) ◽  
pp. 723 ◽  
Author(s):  
Anne Diévart ◽  
Matthew J. Hymes ◽  
Jianming Li ◽  
Steven E. Clark
Keyword(s):  
Kinase Domain ◽  
Chimeric Proteins ◽  
Chimeric Receptor ◽  
Receptor Function ◽  
Loss Of Function ◽  
Chimeric Receptors ◽  
Defence Responses ◽  
Leucine Rich Repeats ◽  
Lrr Domain

CLAVATA1 (CLV1) and BRASSINOSTEROID INSENSITIVE 1 (BRI1) belong to the leucine-rich repeat receptor-like kinase (LRR-RLK) family, comprising more than 200 members in Arabidopsis thaliana (L.) Heynh. and playing important roles in development and defence responses in many plant species (Diévart and Clark 2003, 2004; Shiu and Bleecker 2001a, b). To dissect the mechanisms of receptor function, we assessed the ability of chimeric proteins containing regions from two different receptors to function in vivo. Using domains from the receptor-kinases CLAVATA1 and BRASSINOSTEROID INSENSITIVE1, we tested the ability of the resulting chimeric receptors to replace CLV1 function. Receptors with the BRI1 extracellular domain and CLV1 kinase domain were able to partially replace CLV1 function. Both loss-of-function and gain-of-function mutations within the BRI1 leucine-rich repeats (LRRs) altered the extent of rescue. Chimeric receptor function was unaffected by addition of either exogenous brassinosteroids (BR) or BR biosynthesis inhibitors, suggesting that the chimeric receptors function in a ligand-independent fashion. We propose that the BRI1 LRR domain drives chimeric receptor homodimerisation, and that the BRI1 LRR domain mutations influence homodimerisation efficiency independent of ligand binding.

scholarly journals Regulation of Chk1 by Its C-terminal Domain

2008 ◽  
Vol 19 (11) ◽  
pp. 4546-4553 ◽  
Author(s):  
Ana Kosoy ◽  
Matthew J. O'Connell
Keyword(s):  
Catalytic Domain ◽  
Kinase Domain ◽  
Temperature Sensitive ◽  
Loss Of Function ◽  
Terminal Domain ◽  
C Terminus ◽  
The Face ◽  
Chk1 Kinase

Chk1 is a protein kinase that is the effector molecule in the G2 DNA damage checkpoint. Chk1 homologues have an N-terminal kinase domain, and a C-terminal domain of ∼200 amino acids that contains activating phosphorylation sites for the ATM/R kinases, though the mechanism of activation remains unknown. Structural studies of the human Chk1 kinase domain show an open conformation; the activity of the kinase domain alone is substantially higher in vitro than full-length Chk1, and coimmunoprecipitation studies suggest the C-terminal domain may contain an autoinhibitory activity. However, we show that truncation of the C-terminal domain inactivates Chk1 in vivo. We identify additional mutations within the C-terminal domain that activate ectopically expressed Chk1 without the need for activating phosphorylation. When expressed from the endogenous locus, activated alleles show a temperature-sensitive loss of function, suggesting these mutations confer a semiactive state to the protein. Intragenic suppressors of these activated alleles cluster to regions in the catalytic domain on the face of the protein that interacts with substrate, suggesting these are the regions that interact with the C-terminal domain. Thus, rather than being an autoinhibitory domain, the C-terminus of Chk1 also contains domains critical for adopting an active configuration.

The in vivo distribution and dynamics of DNA topoisomerase II in Drosophila embryonic nuclei and chromosomes

1993 ◽  
Vol 51 ◽  
pp. 74-75
Author(s):  
Jason R. Swedlow ◽  
Neil Osheroff ◽  
Tim Karr ◽  
John W. Sedat ◽  
David A. Agard
Keyword(s):  
Topoisomerase Ii ◽  
Biochemical Characterization ◽  
Developmental Cycle ◽  
Dna Topoisomerase Ii ◽  
Chromosomal Distribution ◽  
Dna Topoisomerase ◽  
Ideal System ◽  
Dnase Treatment

DNA topoisomerase II is an ATP-dependent double-stranded DNA strand-passing enzyme that is necessary for full condensation of chromosomes and for complete segregation of sister chromatids at mitosis in vivo and in vitro. Biochemical characterization of chromosomes or nuclei after extraction with high-salt or detergents and DNAse treatment showed that topoisomerase II was a major component of this remnant, termed the chromosome scaffold. The scaffold has been hypothesized to be the structural backbone of the chromosome, so the localization of topoisomerase II to die scaffold suggested that the enzyme might play a structural role in the chromosome. However, topoisomerase II has not been studied in nuclei or chromosomes in vivo. We have monitored the chromosomal distribution of topoisomerase II in vivo during mitosis in the Drosophila embryo. This embryo forms a multi-nucleated syncytial blastoderm early in its developmental cycle. During this time, the embryonic nuclei synchronously progress through 13 mitotic cycles, so this is an ideal system to follow nuclear and chromosomal dynamics.

scholarly journals microRNA-186 in extracellular vesicles from bone marrow mesenchymal stem cells alleviates idiopathic pulmonary fibrosis via interaction with SOX4 and DKK1

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Jing Zhou ◽  
Yang Lin ◽  
Xiuhua Kang ◽  
Zhicheng Liu ◽  
Wei Zhang ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Idiopathic Pulmonary Fibrosis ◽  
Pulmonary Fibrosis ◽  
Extracellular Vesicles ◽  
Luciferase Assay ◽  
Therapeutic Effects ◽  
Loss Of Function ◽  
Fibroblast Activation ◽  
Promising Therapeutic Approach

Abstract Background Previous reports have identified that human bone marrow mesenchymal stem cell-derived extracellular vesicles (BMSC-EVs) with their cargo microRNAs (miRNAs) are a promising therapeutic approach for the treatment of idiopathic pulmonary fibrosis (IPF). Therefore, we explored whether delivery of microRNA-186 (miR-186), a downregulated miRNA in IPF, by BMSC EVs could interfere with the progression of IPF in a murine model. Methods In a co-culture system, we assessed whether BMSC-EVs modulated the activation of fibroblasts. We established a mouse model of PF to evaluate the in vivo therapeutic effects of BMSC-EVs and determined miR-186 expression in BMSC-EVs by polymerase chain reaction. Using a loss-of-function approach, we examined how miR-186 delivered by BMSC-EVs affected fibroblasts. The putative relationship between miR-186 and SRY-related HMG box transcription factor 4 (SOX4) was tested using luciferase assay. Next, we investigated whether EV-miR-186 affected fibroblast activation and PF by targeting SOX4 and its downstream gene, Dickkopf-1 (DKK1). Results BMSC-EVs suppressed lung fibroblast activation and delayed IPF progression in mice. miR-186 was downregulated in IPF but enriched in the BMSC-EVs. miR-186 delivered by BMSC-EVs could suppress fibroblast activation. Furthermore, miR-186 reduced the expression of SOX4, a target gene of miR-186, and hence suppressed the expression of DKK1. Finally, EV-delivered miR-186 impaired fibroblast activation and alleviated PF via downregulation of SOX4 and DKK1. Conclusion In conclusion, miR-186 delivered by BMSC-EVs suppressed SOX4 and DKK1 expression, thereby blocking fibroblast activation and ameliorating IPF, thus presenting a novel therapeutic target for IPF.

scholarly journals TRIM27 interacts with Iκbα to promote the growth of human renal cancer cells through regulating the NF-κB pathway

BMC Cancer ◽  
2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Chengwu Xiao ◽  
Wei Zhang ◽  
Meimian Hua ◽  
Huan Chen ◽  
Bin Yang ◽  
...  
Keyword(s):  
Cell Lines ◽  
Prognostic Indicator ◽  
The Cancer Genome Atlas ◽  
Mrna Levels ◽  
Loss Of Function ◽  
Protein Levels ◽  
Kaplan Meier ◽  
Cancer Genome Atlas

Abstract Background The tripartite motif (TRIM) family proteins exhibit oncogenic roles in various cancers. The roles of TRIM27, a member of the TRIM super family, in renal cell carcinoma (RCC) remained unexplored. In the current study, we aimed to investigate the clinical impact and roles of TRIM27 in the development of RCC. Methods The mRNA levels of TRIM27 and Kaplan–Meier survival of RCC were analyzed from The Cancer Genome Atlas database. Real-time PCR and Western blotting were used to measure the mRNA and protein levels of TRIM27 both in vivo and in vitro. siRNA and TRIM27 were exogenously overexpressed in RCC cell lines to manipulate TRIM27 expression. Results We discovered that TRIM27 was elevated in RCC patients, and the expression of TRIM27 was closely correlated with poor prognosis. The loss of function and gain of function results illustrated that TRIM27 promotes cell proliferation and inhibits apoptosis in RCC cell lines. Furthermore, TRIM27 expression was positively associated with NF-κB expression in patients with RCC. Blocking the activity of NF-κB attenuated the TRIM27-mediated enhancement of proliferation and inhibition of apoptosis. TRIM27 directly interacted with Iκbα, an inhibitor of NF-κB, to promote its ubiquitination, and the inhibitory effects of TRIM27 on Iκbα led to NF-κB activation. Conclusions Our results suggest that TRIM27 exhibits an oncogenic role in RCC by regulating NF-κB signaling. TRIM27 serves as a specific prognostic indicator for RCC, and strategies targeting the suppression of TRIM27 function may shed light on future therapeutic approaches.

scholarly journals Roles for RNA export factor, Nxt1, in ensuring muscle integrity and normal RNA expression in Drosophila

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Kevin van der Graaf ◽  
Katia Jindrich ◽  
Robert Mitchell ◽  
Helen White-Cooper
Keyword(s):  
Mrna Export ◽  
Rna Stability ◽  
Muscle Degeneration ◽  
Loss Of Function ◽  
Cellular Functions ◽  
Partial Loss ◽  
Export Pathway ◽  
Pathway Gene ◽  
Rna Export

Abstract The mRNA export pathway is responsible for the transport of mRNAs from the nucleus to the cytoplasm, and thus is essential for protein production and normal cellular functions. A partial loss of function allele of the mRNA export factor Nxt1 in Drosophila shows reduced viability and sterility. A previous study has shown that the male fertility defect is due to a defect in transcription and RNA stability, indicating the potential for this pathway to be implicated in processes beyond the known mRNA transport function. Here we investigate the reduced viability of Nxt1 partial loss of function mutants, and describe a defect in growth and maintenance of the larval muscles, leading to muscle degeneration. RNA-seq revealed reduced expression of a set of mRNAs, particularly from genes with long introns in Nxt1 mutant carcass. We detected differential expression of circRNA, and significantly fewer distinct circRNAs expressed in the mutants. Despite the widespread defects in gene expression, muscle degeneration was rescued by increased expression of the costamere component tn (abba) in muscles. This is the first report of a role for the RNA export pathway gene Nxt1 in the maintenance of muscle integrity. Our data also links the mRNA export pathway to a specific role in the expression of mRNA and circRNA from common precursor genes, in vivo.

scholarly journals Periplaneta americana Oligosaccharides Exert Anti-Inflammatory Activity through Immunoregulation and Modulation of Gut Microbiota in Acute Colitis Mice Model

Molecules ◽  
2021 ◽  
Vol 26 (6) ◽  
pp. 1718
Author(s):  
Kaimin Lu ◽  
Jing Zhou ◽  
Jie Deng ◽  
Yangjun Li ◽  
Chuanfang Wu ◽  
...  
Keyword(s):  
Side Effects ◽  
Pathogenic Bacteria ◽  
Periplaneta Americana ◽  
Biochemical Characterization ◽  
Antioxidant Activities ◽  
Inflammatory Activity ◽  
Mice Model ◽  
Acute Colitis ◽  
Anti Inflammatory

The incidence and prevalence of inflammatory bowel disorders (IBD) are increasing around the world due to bacterial infection, abnormal immune response, etc. The conventional medicines for IBD treatment possess serious side effects. Periplaneta americana (P. americana), a traditional Chinese medicine, has been used to treat arthritis, fever, aches, inflammation, and other diseases. This study aimed to evaluate the anti-inflammatory effects of oligosaccharides from P. Americana (OPA) and its possible mechanisms in vivo. OPA were purified and biochemical characterization was analyzed by HPGPC, HPLC, FT-IR, and GC–MS. Acute colitis mice model was established, the acute toxicity and anti-inflammatory activity were tested in vivo. The results showed OPA with molecular mass of 1.0 kDa were composed of 83% glucose, 6% galactose, 11% xylose, and the backbone was (1→4)-Glcp. OPA had potent antioxidant activities in vitro and significantly alleviated the clinical symptoms of colitis, relieved colon damage without toxic side effects in vivo. OPA exhibited anti-inflammatory activity by regulating Th1/Th2, reducing oxidative stress, preserving intestinal barrier integrity, and inhibiting TLR4/MAPK/NF-κB pathway. Moreover, OPA protected gut by increasing microbial diversity and beneficial bacteria, and reducing pathogenic bacteria in feces. OPA might be the candidate of complementary and alternative medicines of IBD with low-cost and high safety.

scholarly journals β-elemene alleviates airway stenosis via the ILK/Akt pathway modulated by MIR143HG sponging miR-1275

2021 ◽  
Vol 26 (1) ◽  
Author(s):  
Guoying Zhang ◽  
Cheng Xue ◽  
Yiming Zeng
Keyword(s):  
Cell Cycle ◽  
Cell Viability ◽  
Cell Model ◽  
Protective Efficacy ◽  
Rabbit Model ◽  
Akt Pathway ◽  
Loss Of Function ◽  
Airway Stenosis

Abstract Background We have previously found that β-elemene could inhibit the viability of airway granulation fibroblasts and prevent airway hyperplastic stenosis. This study aimed to elucidate the underlying mechanism and protective efficacy of β-elemene in vitro and in vivo. Methods Microarray and bioinformatic analysis were used to identify altered pathways related to cell viability in a β-elemene-treated primary cell model and to construct a β-elemene-altered ceRNA network modulating the target pathway. Loss of function and gain of function approaches were performed to examine the role of the ceRNA axis in β-elemene's regulation of the target pathway and cell viability. Additionally, in a β-elemene-treated rabbit model of airway stenosis, endoscopic and histological examinations were used to evaluate its therapeutic efficacy and further verify its mechanism of action. Results The hyperactive ILK/Akt pathway and dysregulated LncRNA-MIR143HG, which acted as a miR-1275 ceRNA to modulate ILK expression, were suppressed in β-elemene-treated airway granulation fibroblasts; β-elemene suppressed the ILK/Akt pathway via the MIR143HG/miR-1275/ILK axis. Additionally, the cell cycle and apoptotic phenotypes of granulation fibroblasts were altered, consistent with ILK/Akt pathway activity. In vivo application of β-elemene attenuated airway granulation hyperplasia and alleviated scar stricture, and histological detections suggested that β-elemene's effects on the MIR143HG/miR-1275/ILK axis and ILK/Akt pathway were in line with in vitro findings. Conclusions MIR143HG and ILK may act as ceRNA to sponge miR-1275. The MIR143HG/miR-1275/ILK axis mediates β-elemene-induced cell cycle arrest and apoptosis of airway granulation fibroblasts by modulating the ILK/Akt pathway, thereby inhibiting airway granulation proliferation and ultimately alleviating airway stenosis.

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