Recombinant expression, characterization, and quantification in human cancer cell lines of the Anaplastic Large-Cell Lymphoma-characteristic NPM-ALK fusion protein.
Actins
/ genetics
Adolescent
Cell Line, Tumor
Child
Electrophoresis, Gel, Two-Dimensional
Enzyme-Linked Immunosorbent Assay
Gene Expression
Humans
Lymphoma, Large-Cell, Anaplastic
/ genetics
Protein-Tyrosine Kinases
/ genetics
Recombination, Genetic
/ genetics
Translocation, Genetic
/ genetics
Young Adult
Journal
Scientific reports
ISSN: 2045-2322
Titre abrégé: Sci Rep
Pays: England
ID NLM: 101563288
Informations de publication
Date de publication:
19 03 2020
19 03 2020
Historique:
received:
12
03
2019
accepted:
05
03
2020
entrez:
21
3
2020
pubmed:
21
3
2020
medline:
15
12
2020
Statut:
epublish
Résumé
Systemic anaplastic large cell lymphoma (ALCL) is an aggressive T-cell lymphoma most commonly seen in children and young adults. The majority of pediatric ALCLs are associated with the t(2;5)(p23;q35) translocation which fuses the Anaplastic Lymphoma Kinase (ALK) gene with the Nucleophosmin (NPM) gene. The NPM-ALK fusion protein is a constitutively-active tyrosine kinase, and plays a major role in tumor pathogenesis. In an effort to advance novel diagnostic approaches and the understanding of the function of this fusion protein in cancer cells, we expressed in E. coli, purified and characterized human NPM-ALK fusion protein to be used as a standard for estimating expression levels in cultured human ALCL cells, a key tool in ALCL pathobiology research. We estimated that NPM-ALK fusion protein is expressed at substantial levels in both Karpas 299 and SU-DHL-1 cells (ca. 4-6 million molecules or 0.5-0.7 pg protein per cell; based on our in-house developed NPM-ALK ELISA; LOD of 40 pM) as compared to the ubiquitous β-actin protein (ca. 64 million molecules or 4.5 pg per lymphocyte). We also compared NPM-ALK/ β-actin ratios determined by ELISA to those independently determined by two-dimensional electrophoresis and showed that the two methods are in good agreement.
Identifiants
pubmed: 32193476
doi: 10.1038/s41598-020-61936-w
pii: 10.1038/s41598-020-61936-w
pmc: PMC7081362
doi:
Substances chimiques
Actins
0
p80(NPM-ALK) protein
EC 2.7.1.-
Protein-Tyrosine Kinases
EC 2.7.10.1
Types de publication
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
5078Subventions
Organisme : NCI NIH HHS
ID : P50 CA126752
Pays : United States
Organisme : NIAID NIH HHS
ID : R21 AI111120
Pays : United States
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