scholarly journals USP21 modulates Goosecoid function through deubiquitination

2019 ◽  
Vol 39 (7) ◽  
Author(s):  
Fuwei Liu ◽  
Qian Fu ◽  
Yunpeng Li ◽  
Kai Zhang ◽  
Mingyue Tang ◽  
...  
Keyword(s):  
Cell Viability ◽  
Protein Stability ◽  
Reporter Gene ◽  
Gene Transcription ◽  
Recombinant Plasmid ◽  
Homeobox Gene ◽  
Craniofacial Development ◽  
Reporter Gene System ◽  
Carboxyl Terminal ◽  
And Function

Abstract The homeobox gene Goosecoid (GSC), which is known to regulate craniofacial development, is activated by mono-ubiquitination; however, the deubiquitylase responsible for GSC deubiquitination and inhibition has yet to be identified. In the present study, we constructed the recombinant plasmid pFlag-CMV-2-GSC and the SRY (sex-determining region Y)-box 6 (Sox6) reporter gene system to identify deubiquitylases that regulate GSC expression. We demonstrate that the ubiquitin carboxyl-terminal hydrolase 21 (USP21) regulates the deubiquitination of GSC negatively, as demonstrated by its inhibition of Sox6 reporter gene transcription. USP21 interacted with GSC to promote GSC deubiquitination while having no effect on GSC protein stability. Cell viability, migration, and function in ATDC5 cells were probably influenced by USP21 through GSC. These findings suggest that USP21 modulates GSC function through deubiquitination.

scholarly journals NKL Homeobox Gene VENTX Is Part of a Regulatory Network in Human Conventional Dendritic Cells

2021 ◽  
Vol 22 (11) ◽  
pp. 5902
Author(s):  
Stefan Nagel ◽  
Claudia Pommerenke ◽  
Corinna Meyer ◽  
Hans G. Drexler
Keyword(s):  
Dendritic Cells ◽  
Transcription Factors ◽  
Cell Lines ◽  
Expression Profiling ◽  
Regulatory Network ◽  
Homeobox Gene ◽  
Leukemia Cell Line ◽  
Specific Gene ◽  
Rna Seq ◽  
And Function

Recently, we documented a hematopoietic NKL-code mapping physiological expression patterns of NKL homeobox genes in human myelopoiesis including monocytes and their derived dendritic cells (DCs). Here, we enlarge this map to include normal NKL homeobox gene expressions in progenitor-derived DCs. Analysis of public gene expression profiling and RNA-seq datasets containing plasmacytoid and conventional dendritic cells (pDC and cDC) demonstrated HHEX activity in both entities while cDCs additionally expressed VENTX. The consequent aim of our study was to examine regulation and function of VENTX in DCs. We compared profiling data of VENTX-positive cDC and monocytes with VENTX-negative pDC and common myeloid progenitor entities and revealed several differentially expressed genes encoding transcription factors and pathway components, representing potential VENTX regulators. Screening of RNA-seq data for 100 leukemia/lymphoma cell lines identified prominent VENTX expression in an acute myelomonocytic leukemia cell line, MUTZ-3 containing inv(3)(q21q26) and t(12;22)(p13;q11) and representing a model for DC differentiation studies. Furthermore, extended gene analyses indicated that MUTZ-3 is associated with the subtype cDC2. In addition to analysis of public chromatin immune-precipitation data, subsequent knockdown experiments and modulations of signaling pathways in MUTZ-3 and control cell lines confirmed identified candidate transcription factors CEBPB, ETV6, EVI1, GATA2, IRF2, MN1, SPIB, and SPI1 and the CSF-, NOTCH-, and TNFa-pathways as VENTX regulators. Live-cell imaging analyses of MUTZ-3 cells treated for VENTX knockdown excluded impacts on apoptosis or induced alteration of differentiation-associated cell morphology. In contrast, target gene analysis performed by expression profiling of knockdown-treated MUTZ-3 cells revealed VENTX-mediated activation of several cDC-specific genes including CSFR1, EGR2, and MIR10A and inhibition of pDC-specific genes like RUNX2. Taken together, we added NKL homeobox gene activities for progenitor-derived DCs to the NKL-code, showing that VENTX is expressed in cDCs but not in pDCs and forms part of a cDC-specific gene regulatory network operating in DC differentiation and function.

scholarly journals Marine Fish Primary Hepatocyte Isolation and Culture: New Insights to Enzymatic Dissociation Pancreatin Digestion

2021 ◽  
Vol 18 (4) ◽  
pp. 1380
Author(s):  
Neusa Figueiredo ◽  
Beatriz Matos ◽  
Mário Diniz ◽  
Vasco Branco ◽  
Marta Martins
Keyword(s):  
Cell Viability ◽  
Cell Cultures ◽  
Marine Fish ◽  
Sparus Aurata ◽  
Cost Effective ◽  
Primary Hepatocyte ◽  
Digestion Method ◽  
Trypan Blue Exclusion ◽  
And Function

Primary cell cultures from wild organisms have been gaining relevance in ecotoxicology as they are considered more sensitive than immortalized cell lines and retain the biochemical pathways found in vivo. In this study, the efficacy of two methods for primary hepatocyte cell isolation was compared using liver from two marine fish (Sparus aurata and Psetta maxima): (i) two-step collagenase perfusion and (ii) pancreatin digestion with modifications. Cell cultures were incubated in L-15 medium at 17 ± 1 °C and monitored for up to six days for cell viability and function using the trypan blue exclusion test, MTT test, lactate dehydrogenase (LDH) activity, and ethoxyresorufin O-deethylase (EROD) activity after Benzo[a]Pyrene exposure. The results showed significant differences between the number of viable cells (p < 0.05), the highest number being obtained for the pancreatin digestion method (average = 4.5 ± 1.9 × 107 cells). Moreover, the hepatocytes showed solid adherence to the culture plate and the rounded shape, changing into a triangular/polygonal shape. The cell viability and function obtained by pancreatin digestion were maintained for five days, and the EROD induction after exposure to the B[a]P showed that cells were metabolically active. This study shows that the optimized pancreatin digestion method is a valid, cost-effective, and simple alternative to the standard perfusion method for the isolation of primary hepatocytes from fish and is suitable for ecotoxicological studies involving marine pollutants, such as PAHs.

The Pleiotropic Intricacies of Hedgehog Signaling: From Craniofacial Patterning to Carcinogenesis

FACE ◽  
2021 ◽  
pp. 273250162110243
Author(s):  
Mikhail Pakvasa ◽  
Andrew B. Tucker ◽  
Timothy Shen ◽  
Tong-Chuan He ◽  
Russell R. Reid
Keyword(s):  
Intracellular Signaling ◽  
Limb Development ◽  
Hedgehog Signaling ◽  
Target Genes ◽  
Craniofacial Development ◽  
Signaling Cascade ◽  
Signaling Mechanisms ◽  
Intracellular Signaling Cascade ◽  
And Function

Hedgehog signaling was discovered more than 40 years ago in experiments demonstrating that it is a fundamental mediator of limb development. Since that time, it has been shown to be important in development, homeostasis, and disease. The hedgehog pathway proceeds through a pathway highly conserved throughout animals beginning with the extracellular diffusion of hedgehog ligands, proceeding through an intracellular signaling cascade, and ending with the activation of specific target genes. A vast amount of research has been done elucidating hedgehog signaling mechanisms and regulation. This research has found a complex system of genetics and signaling that helps determine how organisms develop and function. This review provides an overview of what is known about hedgehog genetics and signaling, followed by an in-depth discussion of the role of hedgehog signaling in craniofacial development and carcinogenesis.

Application of P450RGS reporter gene system as a bioindicator of sediment PAH contamination in the vicinity of Incheon Harbor, Korea

Biomarkers ◽  
1997 ◽  
Vol 2 (3) ◽  
pp. 181-188 ◽  
Author(s):  
Gi Beum Kim ◽  
Jack W. Anderson ◽  
Kristen Bothner ◽  
Jong Hyeon Lee ◽  
Chul Hwan Koh ◽  
...  
Keyword(s):  
Reporter Gene ◽  
Gene System ◽  
Reporter Gene System

Reciprocal relation between reporter gene transcription and translation efficiency in fission yeast

Plasmid ◽  
2021 ◽  
Vol 115 ◽  
pp. 102557
Author(s):  
Suchita Srivastava ◽  
Satinderdeep Kaur ◽  
Hemant K. Verma ◽  
Suman Rani ◽  
Manisha Thakur ◽  
...  
Keyword(s):  
Fission Yeast ◽  
Reporter Gene ◽  
Gene Transcription ◽  
Reciprocal Relation ◽  
Translation Efficiency

scholarly journals Hsp90 inhibition destabilizes Ezh2 protein in alloreactive T cells and reduces graft-versus-host disease in mice

Blood ◽  
2017 ◽  
Vol 129 (20) ◽  
pp. 2737-2748 ◽  
Author(s):  
Qingrong Huang ◽  
Shan He ◽  
Yuanyuan Tian ◽  
Yuting Gu ◽  
Pan Chen ◽  
...  
Keyword(s):  
T Cells ◽  
Protein Stability ◽  
Graft Versus Host Disease ◽  
Hsp90 Inhibition ◽  
Host Disease ◽  
Graft Versus Host ◽  
Alloreactive T Cells ◽  
Graft Versus Leukemia ◽  
Key Points ◽  
And Function

Key Points Ezh2 requires Hsp90 to maintain Ezh2 protein stability and function in alloreactive T cells. Pharmacological inhibition of Hsp90 destabilizes Ezh2 protein in alloreactive T cells and reduces GVHD but preserves graft-versus-leukemia effects.

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