Transposon-based mutagenesis identifies short RIP1 as an activator of NFκB

Maupali Dasgupta, Mukesh K. Agarwal, Patrick Varley, Tao Lu, George R. Stark, Eugene S. Kandel

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

We used a vector based on the Sleeping Beauty transposon to search for constitutive activators of NFκB in cultured cells. Dominant mutations were produced by random insertion of a tetracycline-regulated promoter, which provided robust and exceptionally well-regulated expression of downstream genes. The ability to regulate the mutant phenotype was used to attribute the latter to the insertional event. In one such mutant, the promoter was inserted in the middle of the gene encoding receptor-interacting protein kinase 1 (RIP1). The protein encoded by the hybrid transcript lacks the putative kinase domain of RIP1, but potently stimulates NFκB, AP-1 and Ets-1 activity. Similarly to TNFα treatment, expression of the short RIP1 was toxic to cells that failed to upregulate NFκB. The effects of short RIP1 did not require endogenous RIP1 or cytokine treatment and coincided with reduced responsiveness to TNFα. Additional evidence indicates that a similar short RIP1 could be produced naturally from the ripk1 locus. Interestingly, elevated expression of short RIP1 resulted in the loss of full length RIP1 from cells, pointing to a novel mechanism through which the abundance of RIP1 and the status of the related signaling cascades may be regulated.

Original languageEnglish (US)
Pages (from-to)2249-2256
Number of pages8
JournalCell Cycle
Volume7
Issue number14
StatePublished - Jul 15 2008
Externally publishedYes

Fingerprint

Receptor-Interacting Protein Serine-Threonine Kinases
Mutagenesis
Protein Kinases
Beauty
Gene encoding
Aptitude
Poisons
Transcription Factor AP-1
Tetracycline
Cultured Cells
Phosphotransferases
Up-Regulation
Genes
Cells
Cytokines
Phenotype
Gene Expression

Keywords

  • Forward genetics
  • Insertional mutagenesis
  • NFKB
  • RIP1
  • Sleeping beauty
  • Tumor necrosis factor alpha

ASJC Scopus subject areas

  • Cell Biology
  • Biochemistry
  • Molecular Biology

Cite this

Dasgupta, M., Agarwal, M. K., Varley, P., Lu, T., Stark, G. R., & Kandel, E. S. (2008). Transposon-based mutagenesis identifies short RIP1 as an activator of NFκB. Cell Cycle, 7(14), 2249-2256.

Transposon-based mutagenesis identifies short RIP1 as an activator of NFκB. / Dasgupta, Maupali; Agarwal, Mukesh K.; Varley, Patrick; Lu, Tao; Stark, George R.; Kandel, Eugene S.

In: Cell Cycle, Vol. 7, No. 14, 15.07.2008, p. 2249-2256.

Research output: Contribution to journalArticle

Dasgupta, M, Agarwal, MK, Varley, P, Lu, T, Stark, GR & Kandel, ES 2008, 'Transposon-based mutagenesis identifies short RIP1 as an activator of NFκB', Cell Cycle, vol. 7, no. 14, pp. 2249-2256.
Dasgupta M, Agarwal MK, Varley P, Lu T, Stark GR, Kandel ES. Transposon-based mutagenesis identifies short RIP1 as an activator of NFκB. Cell Cycle. 2008 Jul 15;7(14):2249-2256.
Dasgupta, Maupali ; Agarwal, Mukesh K. ; Varley, Patrick ; Lu, Tao ; Stark, George R. ; Kandel, Eugene S. / Transposon-based mutagenesis identifies short RIP1 as an activator of NFκB. In: Cell Cycle. 2008 ; Vol. 7, No. 14. pp. 2249-2256.
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