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Genetic evidence that uptake of the fluorescent analog 2NBDG occurs independently of known glucose transporters

ABSTRACT: Genetic evidence that uptake of the fluorescent analog 2NBDG occurs independently of known glucose transporters

PROVIDER: PRJNA834952 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR19076889_1.fastq.gz	Fastqsanger.gz
	SRR19076889_2.fastq.gz	Fastqsanger.gz
	SRR19076890_1.fastq.gz	Fastqsanger.gz
	SRR19076890_2.fastq.gz	Fastqsanger.gz
	SRR19076891_1.fastq.gz	Fastqsanger.gz

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Genetic evidence that uptake of the fluorescent analog 2NBDG occurs independently of known glucose transporters

Project description:RNA-sequencing was performed to examine for compensatory upregulation of Slc2 transporters in Slc2a1 gRNA- or Slc2a1, Slc2a3, Slc2a5, Slc2a6, and Slc2a8 gRNA-transduced 5TGM1 cells relative to a control (Rosa26) gRNA transduced 5TGM1-Cas9 cell line. Abstract: The fluorescent derivative of glucose, 2-Deoxy-2-[(7-nitro-2,1,3-benzoxadiazol-4-yl)-amino]-D-glucose (2NBDG), is a widely used surrogate reagent to visualize glucose uptake in live cells at single cell resolution. Using a model of CRISPR-Cas9 gene editing in 5TGM1 myeloma cells, we demonstrate that ablation of the glucose transporter gene Slc2a1 abrogates radioactive glucose uptake but has no effect on the magnitude or kinetics of 2NBDG import. Extracellular 2NBDG, but not NBD-fructose was transported by plasma cells into the cytoplasm suggesting a specific mechanism that is unlinked to glucose import and that of chemically similar compounds. Competing glucose in media or pharmacological inhibition of GLUT1 did not have any effect on 2NBDG uptake. RNA-Seq analysis of primary plasma cells and the 5TGM1 myeloma cell line revealed expression of other candidate glucose transporters. Yet, deletion of these transporters individually or in combination with one another also had no impact on 2NBDG uptake. Ablation of the genes in the Slc29 and Slc35 families of nucleoside and nucleoside sugar transporters as well as the ATP-binding cassette (ABC) transporter family also failed to impact 2NBDG import. Thus, cellular uptake of 2NBDG is promoted by an unknown mechanism and is not a faithful indicator of glucose transport.

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