Physiological restraint of Bak by Bcl-xL is essential for cell survival

Erinna F Lee, Stephanie Grabow, Stephane Chappaz, Grant Dewson, Colin Hockings, Ruth M Kluck, Marlyse A. Debrincat, Daniel H. Gray, Matthew T. Witkowski, Marco Evangelista, Anne Pettikiriarachchi, Phillipe Bouillet, Rachael M. Lane, Peter E. Czabotar, Peter M. Colman, Brian J. Smith, Benjamin T. Kile, W. Douglas Fairlie

Research output: Contribution to journalArticleResearchpeer-review

Abstract

Due to the myriad interactions between prosurvival and proapoptotic members of the Bcl-2 family of proteins, establishing the mechanisms that regulate the intrinsic apoptotic pathway has proven challenging. Mechanistic insights have primarily been gleaned from in vitro studies because genetic approaches in mammals that produce unambiguous data are difficult to design. Here we describe a mutation in mouse and human Bak that specifically disrupts its interaction with the prosurvival protein Bcl-xL. Substitution of Glu75 in mBak (hBAK Q77) for leucine does not affect the three-dimensional structure of Bak or killing activity but reduces its affinity for Bcl-xL via loss of a single hydrogen bond. Using this mutant, we investigated the requirement for physical restraint of Bakby Bcl-xL in apoptotic regulation. In vitro, BakQ75L cellswere significantly more sensitive to various apoptotic stimuli. In vivo, loss of Bcl-xL binding to Bak led to significant defects in T-cell and blood platelet survival. Thus, we provide the first definitive in vivo evidence that prosurvival proteins maintain cellular viability by interacting with and inhibiting Bak.

Original languageEnglish
Pages (from-to)1240-1250
Number of pages11
JournalGenes and Development
Volume30
Issue number10
DOIs
Publication statusPublished - 19 May 2016
Externally publishedYes

Keywords

  • Apoptosis
  • Bak
  • Bcl-2
  • Bcl-x
  • BH3

Cite this

Lee, E. F., Grabow, S., Chappaz, S., Dewson, G., Hockings, C., Kluck, R. M., ... Fairlie, W. D. (2016). Physiological restraint of Bak by Bcl-xL is essential for cell survival. Genes and Development, 30(10), 1240-1250. https://doi.org/10.1101/gad.279414.116
Lee, Erinna F ; Grabow, Stephanie ; Chappaz, Stephane ; Dewson, Grant ; Hockings, Colin ; Kluck, Ruth M ; Debrincat, Marlyse A. ; Gray, Daniel H. ; Witkowski, Matthew T. ; Evangelista, Marco ; Pettikiriarachchi, Anne ; Bouillet, Phillipe ; Lane, Rachael M. ; Czabotar, Peter E. ; Colman, Peter M. ; Smith, Brian J. ; Kile, Benjamin T. ; Fairlie, W. Douglas. / Physiological restraint of Bak by Bcl-xL is essential for cell survival. In: Genes and Development. 2016 ; Vol. 30, No. 10. pp. 1240-1250.
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abstract = "Due to the myriad interactions between prosurvival and proapoptotic members of the Bcl-2 family of proteins, establishing the mechanisms that regulate the intrinsic apoptotic pathway has proven challenging. Mechanistic insights have primarily been gleaned from in vitro studies because genetic approaches in mammals that produce unambiguous data are difficult to design. Here we describe a mutation in mouse and human Bak that specifically disrupts its interaction with the prosurvival protein Bcl-xL. Substitution of Glu75 in mBak (hBAK Q77) for leucine does not affect the three-dimensional structure of Bak or killing activity but reduces its affinity for Bcl-xL via loss of a single hydrogen bond. Using this mutant, we investigated the requirement for physical restraint of Bakby Bcl-xL in apoptotic regulation. In vitro, BakQ75L cellswere significantly more sensitive to various apoptotic stimuli. In vivo, loss of Bcl-xL binding to Bak led to significant defects in T-cell and blood platelet survival. Thus, we provide the first definitive in vivo evidence that prosurvival proteins maintain cellular viability by interacting with and inhibiting Bak.",
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Lee, EF, Grabow, S, Chappaz, S, Dewson, G, Hockings, C, Kluck, RM, Debrincat, MA, Gray, DH, Witkowski, MT, Evangelista, M, Pettikiriarachchi, A, Bouillet, P, Lane, RM, Czabotar, PE, Colman, PM, Smith, BJ, Kile, BT & Fairlie, WD 2016, 'Physiological restraint of Bak by Bcl-xL is essential for cell survival' Genes and Development, vol. 30, no. 10, pp. 1240-1250. https://doi.org/10.1101/gad.279414.116

Physiological restraint of Bak by Bcl-xL is essential for cell survival. / Lee, Erinna F; Grabow, Stephanie; Chappaz, Stephane; Dewson, Grant; Hockings, Colin; Kluck, Ruth M; Debrincat, Marlyse A.; Gray, Daniel H.; Witkowski, Matthew T.; Evangelista, Marco; Pettikiriarachchi, Anne; Bouillet, Phillipe; Lane, Rachael M.; Czabotar, Peter E.; Colman, Peter M.; Smith, Brian J.; Kile, Benjamin T.; Fairlie, W. Douglas.

In: Genes and Development, Vol. 30, No. 10, 19.05.2016, p. 1240-1250.

Research output: Contribution to journalArticleResearchpeer-review

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AU - Lee, Erinna F

AU - Grabow, Stephanie

AU - Chappaz, Stephane

AU - Dewson, Grant

AU - Hockings, Colin

AU - Kluck, Ruth M

AU - Debrincat, Marlyse A.

AU - Gray, Daniel H.

AU - Witkowski, Matthew T.

AU - Evangelista, Marco

AU - Pettikiriarachchi, Anne

AU - Bouillet, Phillipe

AU - Lane, Rachael M.

AU - Czabotar, Peter E.

AU - Colman, Peter M.

AU - Smith, Brian J.

AU - Kile, Benjamin T.

AU - Fairlie, W. Douglas

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