Nix - Professional Heart Daily

Diverse mechanisms of Nix-mediated cell death
Gerald W Dorn II MD
FINANCIAL DISCLOSURE:
NHLBI R01s HL059888, HL080008, and HL087871
NHLBI P50s HL077101 and HL077113
UNLABELED/UNAPPROVED
USES DISCLOSURE:
NHLBI
None
r
h
Normal heart
Hypertrophic
genes
TAC/Gαq
r
h
Hypertrophied
heart
Cardiomyocyte
apoptosis
Nix
r
h
Cardiomyopathic
Dilation
What is Nix?
• Nix is a transcriptionally regulated member of the pro‐
apoptotic BNip subfamily of Bcl2 proteins.
• Unlike members of the pro‐apoptotic BH3‐only subfamily (Bad, Bid, Bim, Puma), the Nix pseudo‐BH3 domain is not required for its death effects.
• Whereas BH3‐only factors stimulate mitochondrial apoptosis, Nix stimulates atypical cell death having characteristics of both apoptosis and necrosis.
Cell Death and Diff 2009 16:939-46
Aouacheria, A. et al. Mol Biol Evol 2005 22:2395-2416
We know that:
• Nix is transcriptionally upregulated in experimental and clinical cardiac hypertrophy (Nat Med 2002; J Biol Chem 2006).
• Forced cardiomyocyte expression of Nix is sufficient to cause apoptotic cardiomyopathy, mediated via the mitochondrial pathway (Nat Med 2002; Circ Res 2004).
• Cardiomyocyte‐specific Nix gene ablation protects against ventricular remodeling and decreases cardiomyocyte apoptosis after TAC (Circulation 2008).
Nix stimulates mitochondrial pathway apoptosis
Mitochondrial apoptosis
Nix
FLAG-Nix
Overlay
Bax/Bak
Apoptosome Cyt c
Apaf-1 dATP
Casp 9
Mitochondria
Casp 3
DNA frag
(200 bp)
Cell Death
Nat Med. 2002 8:725-30
Nix localizes to mitochondria and ER/SR
Diwan A et al, J Clin Invest. 119:203-212, 2009
Transfected and endogenous cardiomyocyte Nix
localize to mitochondria and ER/SR
HEK cells
NRCM
Mito
ER/SR
Mouse heart (standard)
Mouse heart (sucrose grad)
Mito
ER/SR
Diwan A et al, J Clin Invest. 119:203-212, 2009
Nix overexpression increases SR calcium content
Conditional Nix
transgenic
Diwan A et al, J Clin Invest. 119:203-212, 2009
Nix ablation decreases SR calcium content
Conditional Nix
transgenic
Nix
knockout
Diwan A et al, J Clin Invest. 119:203-212, 2009
Nix localizes to ER and SR and increases SR Ca++ stores
Mitochondrial apoptosis
Nix
Regulation of
SR Ca++ stores
Bax/Bak
Apoptosome Cyt c
Apaf-1 dATP
Casp 9
Mitochondria
SR
Ca++
Casp 3
DNA frag
(200 bp)
Cell Death
Creation of organelle-specific Nix mutants
BH3
TM
Nix-wt
Nix-wt
Nix-ActA
mitochondria
Nix-cb5
ER/SR
Nix-trunc
No target
Mito
ER/SR
Diwan A et al, J Clin Invest. 119:203-212, 2009
Both mitochondrial- and SR-targeted Nix cause PCD.
Adenoviral Nix mutant expression in NRCM
SR-targeted, but not mitochondrial-Nix, decreases Δψm
No Nix (βgal)
Mito and SR Nix
Mito Nix-Act A
SR Nix-cb5
Diwan A et al, J Clin Invest. 119:203-212, 2009
cytosolic Nix trunc
Mitochondrial localization is sufficient for
Nix-mediated cell death.
Mitochondrial apoptosis
Nix-ActA
Bax/Bak
Apoptosome Cyt c
Apaf-1 dATP
Casp 9
Mitochondria
Casp 3
DNA frag
(200 bp)
Cell Death
Mitochondrial localization is dispensable for
Nix-mediated cell death.
Nix-cb5
Mitochondrial apoptosis
Apoptosome Cyt c
Apaf-1 dATP
Casp 9
Mitochondria
Δψm
Casp 3
DNA frag
(200 bp)
Cell Death
SR
What are the consequences of organelle‐specific Nix on cardiac function? Genetic mouse experiment
Gedankenexperiment
Hypothesis
Tissue culture experiment
Stay tuned to find out…………
Nix-cb5
Bax/Bak
SR
Mitochondria
Ca++
TP
MP
H20
ATP
depletion
Necrotic
Cell Death
Mfn1
Mfn2
Opa1
Cell, Volume 135, Issue 7, Pages 1165-1167
Genetic mouse experiment
Gedankenexperiment
Hypothesis
Tissue culture experiment
Genetic mouse experiment
Gedankenexperiment
Hypothesis
Tissue culture experiment
Genetic fly experiment
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Scot Matkovich
Abhinav Diwan
Yizheng Tu
Min Young Kang
Derek Van Booven
John Engelhard
Kyuwan Shim
Anna Hindes
Will Eschenbacher Mark Snee
Yun Chen
Lisa Dorn
Alina Kutsenko
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Jeanne Nerbonne
Steve Warner
Greg Longmore
Robi Mitra
Ken Polonsky
•
•
Cindy Sweet
Karen Gill
• Jeff Molkentin (ppif ‐/‐)
• David Chan (Mfn2 loxP)
• Litsa Kranias (Pln ‐/‐)