Truncation mutation in HRG4 (UNC119) leads to mitochondrial ANT-1-mediated photoreceptor synaptic and retinal degeneration by apoptosis.
OBJECTIVE
To characterize the time course of apoptosis and degeneration in a transgenic mouse model of retinal degeneration based on truncated mutant HRG4; to investigate the nature of binding of the mutant HRG4 to its target, ADP-ribosylation factor-like (ARL)2; to study its effects on the downstream molecules Binder-of-ARL2 (BART) and adenine nucleotide transporter (ANT)-1 and on the induction of apoptosis.


METHODS
Saturation binding, microscopic morphometric, Western blot, immunofluorescence, and TUNEL analyses were used.


RESULTS
Increased apoptosis did not occur until 20 months in the transgenic retina, consistent with the delayed-onset degeneration in this model. The truncated HRG4 protein exhibited approximately threefold greater affinity for ARL2 than the wild-type HRG4, likely resulting in nonfunctional sequestration of ARL2. A significant decrease in ARL2 was present by 20 months, accompanied by a 50% decrease in ANT-1 in the photoreceptor synaptic mitochondria, with evidence of mitochondrial dysfunction. Preapoptotic degeneration in the photoreceptor synapse was demonstrated with cytochrome c release and caspase 3 activation within the synapse-without evidence of TUNEL-positive apoptosis in the photoreceptor cell body-indicating an initial event in the synapse leading to apoptosis. Caspase 3 was activated in the accompanying secondary neuron, consistent with transsynaptic degeneration.


CONCLUSIONS
The results support a novel mechanism of retinal degeneration in which preapoptotic degeneration starts in the photoreceptor synapse because of a deficiency in ANT-1 and spreads to the secondary neuron transsynaptically, followed by apoptosis and degeneration in the cell body of the photoreceptor.
Mori
Naoki
N
Bascom Palmer Eye Institute, University of Miami Miller School of Medicine, Miami, Florida 33136, USA.
Ishiba
Yasutsugu
Y
Kubota
Shinya
S
Kobayashi
Akira
A
Higashide
Tomomi
T
McLaren
Margaret J
MJ
Inana
George
G
D048868:Adaptor Proteins, Signal Transducing; D033741:Adenine Nucleotide Translocator 1 / Q000172:deficiency*; D000818:Animals; D017209:Apoptosis; D015153:Blotting, Western; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D018389:Codon, Nonsense; D045304:Cytochromes c / Q000378:metabolism; D004789:Enzyme Activation; D020287:In Situ Nick-End Labeling; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics*; D051379:Mice; D008822:Mice, Transgenic; D008856:Microscopy, Fluorescence; D008868:Microtubule Proteins / Q000235:genetics*; D008928:Mitochondria / Q000378:metabolism*; D020419:Photoreceptor Cells, Vertebrate / Q000378:metabolism*; D012162:Retinal Degeneration / Q000235:genetics* / Q000378:metabolism; D013569:Synapses / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; D033741:Adenine Nucleotide Translocator 1; D018389:Codon, Nonsense; D047908:Intracellular Signaling Peptides and Proteins; D008868:Microtubule Proteins; C489115:Unc119 protein, mouse; D045304:Cytochromes c; C505409:Casp3 protein, mouse; D053148:Caspase 3; D020169:Caspases