Abstract
H1069Q substitution represents the most frequent mutation of the copper transporter ATP7B causing Wilson disease in Caucasian population. ATP7B localizes to the Golgi complex in hepatocytes but moves in response to copper overload to the endo-lysosomal compartment to support copper excretion via bile canaliculi. In heterologous or hepatoma-derived cell lines, overexpressed ATP7B-H1069Q is strongly retained in the ER and fails to move to the post-Golgi sites, resulting in toxic copper accumulation. However, this pathogenic mechanism has never been tested in patients’ hepatocytes, while animal models recapitulating this form of WD are still lacking. To reach this goal, we have reprogrammed skin fibroblasts of homozygous ATP7B-H1069Q patients into induced pluripotent stem cells and differentiated them into hepatocyte-like cells. Surprisingly, in HLCs we found one third of ATP7B-H1069Q localized in the Golgi complex and able to move to the endo-lysosomal compartment upon copper stimulation. However, despite normal mRNA levels, the expression of the mutant protein was only 20% compared to the control because of endoplasmic reticulum-associated degradation. These results pinpoint rapid degradation as the major cause for loss of ATP7B function in H1069Q patients, and thus as the primary target for designing therapeutic strategies to rescue ATP7B-H1069Q function.
Highlights
H1069Q substitution represents the most frequent mutation of the copper transporter ATP7B causing Wilson disease in Caucasian population
The main new result presented here is the characterization of the properties of ATP7B-H1069Q endogenously expressed in homozygous patient-derived hepatocyte-like cells (HLCs)
We investigated for the first time the dynamics of ATP7B-H1069Q mutant under isogenic conditions
Summary
H1069Q substitution represents the most frequent mutation of the copper transporter ATP7B causing Wilson disease in Caucasian population. In heterologous or hepatoma-derived cell lines, overexpressed ATP7B-H1069Q is strongly retained in the ER and fails to move to the post-Golgi sites, resulting in toxic copper accumulation. This pathogenic mechanism has never been tested in patients’ hepatocytes, while animal models recapitulating this form of WD are still lacking. Despite normal mRNA levels, the expression of the mutant protein was only 20% compared to the control because of endoplasmic reticulum-associated degradation These results pinpoint rapid degradation as the major cause for loss of ATP7B function in H1069Q patients, and as the primary target for designing therapeutic strategies to rescue ATP7B-H1069Q function. We and others have provided proof of principle for new correction strategies to contrast the disease: different treatments such as incubation at low temperature or with curcumin[9,11], expression of CRYAB12, inhibition of p38 and JNK kinase[13], were all effective in rescuing the localization and/or Cu response of the overexpressed mutant
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