- Email: [email protected]
68. Interaction between parkin and glucocerebrosidase: A possible link between Parkinson disease and Gaucher disease
Abstracts / Molecular Genetics and Metabolism 96 (2009) S12–S47 ogenesis of the disease through the study of PPT1 knockout mice. Analysis of the brain protein palmitoylome has revealed no major differences in the abundance of intact palmitoylated proteins, consistent with the role of PPT1 in the degradation of smaller palmitoylated peptides in the lysosome. Differences in lipid profiles in brain tissue have been uncovered, but these have been difficult to relate to the primary enzyme defect. Current work involves identification of disease markers and development of tools and reagents for enzyme replacement therapy, which poses considerable challenges.
S27
Reference [1] R.E. Infante etal., NPC2 facilitates bidirectional transfer of cholesterol between NPC1 and lipid bilayers, a step in cholesterol egress from lysosomes, Proc Natl Acad Sci USA 105 (2008) 15287–15292. doi:10.1016/j.ymgme.2008.11.070
doi:10.1016/j.ymgme.2008.11.068
70. Niemann-Pick disease type C and inflammatory Bowel diseases: Is there any correlation? 68. Interaction between parkin and glucocerebrosidase: A possible link between Parkinson disease and Gaucher disease
Elizabeth Jacklin, Jackie Imrie, Ed Wraith, Royal Manchester Childrens Hospital, Manchester, England
Mia Horowitz, Idit Ron, Ramat Aviv, Israel Gaucher disease, a sphingolipidosis characterized by impaired activity of the lysosomal enzyme glucocerebrosidase results from mutations in the glucocerebrosidase gene. We have shown that mutant glucocerebrosidase variants present variable degrees of ER retention and undergo ER associated degradation (ERAD) in the proteasome. Recently, an association between Gaucher disease and Parkinson disease has been demonstrated by the concurrence of Gaucher disease and parkinsonism in patients and the identification of glucocerebrosidase mutations in probands with sporadic Parkinson disease. Parkinson disease, a severe neurological disorder of movement, is the second most common neurodegenerative disorder. One of the major genes involved in Parkinson disease is parkin, a product of the causative gene PARK2 of autosomal recessive juvenile parkinsonism (ARJP). Parkin is an E3 ubiquitin-protein ligase, indicating that ARJP is linked to impediment of proteolysis mediated by the ubiquitin-proteasome system. Loss of parkin function leads to accumulation of its substrates, which may be the cause of neurodegeneration in parkin-related parkinsonism. Parkin also functions in the ERAD of misfolded ER protein, and it is up-regulated by unfolded-protein stress. We therefore, tested the possibility that the concurrence of Gaucher disease and Parkinson disease reflects the fact that parkin is the E3 ligase of misfolded mutant glucocerebrosidase variants. To test this possibility, we used SH-5YSY dopaminergic neuroblastoma cells that serve as an invitro cell model for Parkinson disease in tissue culture. Our results showed that the N370S mutant glucocerebrosidase variant associates with parkin. Overexpression of parkin reduced steady state levels of mutant glucocerebrosidase. However, the T240R parkin ring finger mutant failed to do so, indicating that the ring finger domain of parkin participates in parkin dependent degradation of mutant glucocerebrosidase. Parkin was also involved in ubiquitination of the N370S glucocerebrosidase mutant. We suggest that involvement of parkin in degradation of mutant glucocerebrosidase explains the concurrence of Gaucher disease and parkinsonism. doi:10.1016/j.ymgme.2008.11.069
69. NPC2 facilitates bidirectional transfer of cholesterol between NPC1 and lipid bilayers, a potential step in cholesterol egress from lysosomes Rodney Infante, Michael Wang, Arun Radhakrishnan, Hyock Kwon, Michael Brown, Joseph Goldstein, UT Southwestern Medical Center, Dallas, TX, USA The trafficking of lipoprotein-derived cholesterol from lysosomes is dependent on the polytopic membrane-bound Niemann-Pick C1 (NPC1) and the soluble Niemann-Pick C2 lysosomal proteins. The requirement for these proteins in the transport of cholesterol out of the lysosomes remains unclear. Previously, we demonstrated the ability of the N-terminal luminal domain of NPC1, hereinafter called NPC1 (NTD), to bind cholesterol with nM affinity. We and others also showed the ability of NPC2 to bind cholesterol with similar affinity. Recently, we established an invitro assay detecting the movement of [3H]cholesterol between these two proteins and model membranes. NPC2 readily transfers or receives [3H]cholesterol to and from phosphatidylcholine liposomes, relative to NPC1 (NTD). The bidirectional transport of [3H]cholesterol between NPC1 (NTD) and liposomes is accelerated over a 100-fold in the presence of NPC2. The naturally occurring mutant (Pro120Ser) form of NPC2 did not demonstrate significant cholesterol binding and had a reduction in the facilitation of cholesterol transport between NPC1 (NTD) and liposomes. These studies begin to define the dual requirement of both proteins in the egress of lipoprotein-derived cholesterol out of the lysosomes, thus explaining why mutations in either protein cause a similar clinical phenotype [1].
Introduction: It would appear that the incidence of inflammatory bowel disease in patients with Niemann-Pick type C is disproportionately high when compared to patients of similar age and ethnic backgrounds who are disease free. Hypothosis: This has led clinicians to ask the question if there is, therefore, some correlation between NPC and Crohn’s disease. There is only one paper published looking at the two diseases. Methods: Case History review on all patients with Niemann-Pick type C and inflammatory bowel disease known to us. Results: Documented findings on 6 patients with Niemann-Pick type C disease and inflammatory bowel conditions. Conclusion: The need for a multi-centre international case History review study on all patients diagnosed with Inflammatory bowel disease and NPC. doi:10.1016/j.ymgme.2008.11.071
71. Case report of chaperone therapy for Adult Tay-Sachs disease and comparison to an in-silico pharmacokinetic and cellular simulation John (Jack) Keimel, Lawrence Charnas, University of Minnesota, North Oaks, MN, USA Background: Individuals with adult Tay-Sachs disease (ATSD) develop ataxia and dysarthria by early teenage years and later lose ability to walk. No therapy has yet been shown clinically effective. ATSD is caused by inadequate Beta-hexosaminidase-A (HexA) activity and GM2 ganglioside accumulation in neuronal lysosomes. The most prevalent ATSD mutation, alphaG269S, is not believed to impact HexA enzyme function but instead cause post-translation misfolding and reduced enzyme stability. Others have shown invitro that pyrimethamine (PYR) is a potential pharmacological chaperone therapy for improving stability of G269S mutated HexA and increasing its transport to the lysosomes. Model description: Computer simulations were done to estimate PYR pharmacokinetics and GM2 response using three interlinked models: a pharmacokinetics model estimated neuronal PYR concentrations; an endoplasmic reticulum model predicted the relative amount of HexA transported to lysosomes; and a lysosomal model simulated GM2 degradation and HexA inhibition caused by PYR. The simulations also modulated HexA half-life as a function of accumulated GM2 substrate. Previously published parameter values were used when available. Case report: A male confirmed with ATSD and the G269S mutation, had been taking a substrate reduction therapy, miglustat (200 mg, tid), for greater than four years, but at age 25 showed accelerated decline in coordination and leg muscle strength, verified by EMG evaluations. At the request of the patient and parents, PYR 25 mg QD was begun for two weeks followed by 75 mg QD in combination with folic acid (5 mg QD) to offset partial PRY dihydrofolate reductase inhibition. Weekly leukocyte HexA assays and monthly hematology tests (CBC) were conducted. Results: No noticeable side-effects were observed during the first four weeks of treatment. Additional follow-up and comparisons to the modeled response will be presented. doi:10.1016/j.ymgme.2008.11.072
72. Proteostasis Jeffery Kelly, The Scripps Research Institute, La Jolla, CA, USA
S27
Reference [1] R.E. Infante etal., NPC2 facilitates bidirectional transfer of cholesterol between NPC1 and lipid bilayers, a step in cholesterol egress from lysosomes, Proc Natl Acad Sci USA 105 (2008) 15287–15292. doi:10.1016/j.ymgme.2008.11.070
doi:10.1016/j.ymgme.2008.11.068
70. Niemann-Pick disease type C and inflammatory Bowel diseases: Is there any correlation? 68. Interaction between parkin and glucocerebrosidase: A possible link between Parkinson disease and Gaucher disease
Elizabeth Jacklin, Jackie Imrie, Ed Wraith, Royal Manchester Childrens Hospital, Manchester, England
Mia Horowitz, Idit Ron, Ramat Aviv, Israel Gaucher disease, a sphingolipidosis characterized by impaired activity of the lysosomal enzyme glucocerebrosidase results from mutations in the glucocerebrosidase gene. We have shown that mutant glucocerebrosidase variants present variable degrees of ER retention and undergo ER associated degradation (ERAD) in the proteasome. Recently, an association between Gaucher disease and Parkinson disease has been demonstrated by the concurrence of Gaucher disease and parkinsonism in patients and the identification of glucocerebrosidase mutations in probands with sporadic Parkinson disease. Parkinson disease, a severe neurological disorder of movement, is the second most common neurodegenerative disorder. One of the major genes involved in Parkinson disease is parkin, a product of the causative gene PARK2 of autosomal recessive juvenile parkinsonism (ARJP). Parkin is an E3 ubiquitin-protein ligase, indicating that ARJP is linked to impediment of proteolysis mediated by the ubiquitin-proteasome system. Loss of parkin function leads to accumulation of its substrates, which may be the cause of neurodegeneration in parkin-related parkinsonism. Parkin also functions in the ERAD of misfolded ER protein, and it is up-regulated by unfolded-protein stress. We therefore, tested the possibility that the concurrence of Gaucher disease and Parkinson disease reflects the fact that parkin is the E3 ligase of misfolded mutant glucocerebrosidase variants. To test this possibility, we used SH-5YSY dopaminergic neuroblastoma cells that serve as an invitro cell model for Parkinson disease in tissue culture. Our results showed that the N370S mutant glucocerebrosidase variant associates with parkin. Overexpression of parkin reduced steady state levels of mutant glucocerebrosidase. However, the T240R parkin ring finger mutant failed to do so, indicating that the ring finger domain of parkin participates in parkin dependent degradation of mutant glucocerebrosidase. Parkin was also involved in ubiquitination of the N370S glucocerebrosidase mutant. We suggest that involvement of parkin in degradation of mutant glucocerebrosidase explains the concurrence of Gaucher disease and parkinsonism. doi:10.1016/j.ymgme.2008.11.069
69. NPC2 facilitates bidirectional transfer of cholesterol between NPC1 and lipid bilayers, a potential step in cholesterol egress from lysosomes Rodney Infante, Michael Wang, Arun Radhakrishnan, Hyock Kwon, Michael Brown, Joseph Goldstein, UT Southwestern Medical Center, Dallas, TX, USA The trafficking of lipoprotein-derived cholesterol from lysosomes is dependent on the polytopic membrane-bound Niemann-Pick C1 (NPC1) and the soluble Niemann-Pick C2 lysosomal proteins. The requirement for these proteins in the transport of cholesterol out of the lysosomes remains unclear. Previously, we demonstrated the ability of the N-terminal luminal domain of NPC1, hereinafter called NPC1 (NTD), to bind cholesterol with nM affinity. We and others also showed the ability of NPC2 to bind cholesterol with similar affinity. Recently, we established an invitro assay detecting the movement of [3H]cholesterol between these two proteins and model membranes. NPC2 readily transfers or receives [3H]cholesterol to and from phosphatidylcholine liposomes, relative to NPC1 (NTD). The bidirectional transport of [3H]cholesterol between NPC1 (NTD) and liposomes is accelerated over a 100-fold in the presence of NPC2. The naturally occurring mutant (Pro120Ser) form of NPC2 did not demonstrate significant cholesterol binding and had a reduction in the facilitation of cholesterol transport between NPC1 (NTD) and liposomes. These studies begin to define the dual requirement of both proteins in the egress of lipoprotein-derived cholesterol out of the lysosomes, thus explaining why mutations in either protein cause a similar clinical phenotype [1].
Introduction: It would appear that the incidence of inflammatory bowel disease in patients with Niemann-Pick type C is disproportionately high when compared to patients of similar age and ethnic backgrounds who are disease free. Hypothosis: This has led clinicians to ask the question if there is, therefore, some correlation between NPC and Crohn’s disease. There is only one paper published looking at the two diseases. Methods: Case History review on all patients with Niemann-Pick type C and inflammatory bowel disease known to us. Results: Documented findings on 6 patients with Niemann-Pick type C disease and inflammatory bowel conditions. Conclusion: The need for a multi-centre international case History review study on all patients diagnosed with Inflammatory bowel disease and NPC. doi:10.1016/j.ymgme.2008.11.071
71. Case report of chaperone therapy for Adult Tay-Sachs disease and comparison to an in-silico pharmacokinetic and cellular simulation John (Jack) Keimel, Lawrence Charnas, University of Minnesota, North Oaks, MN, USA Background: Individuals with adult Tay-Sachs disease (ATSD) develop ataxia and dysarthria by early teenage years and later lose ability to walk. No therapy has yet been shown clinically effective. ATSD is caused by inadequate Beta-hexosaminidase-A (HexA) activity and GM2 ganglioside accumulation in neuronal lysosomes. The most prevalent ATSD mutation, alphaG269S, is not believed to impact HexA enzyme function but instead cause post-translation misfolding and reduced enzyme stability. Others have shown invitro that pyrimethamine (PYR) is a potential pharmacological chaperone therapy for improving stability of G269S mutated HexA and increasing its transport to the lysosomes. Model description: Computer simulations were done to estimate PYR pharmacokinetics and GM2 response using three interlinked models: a pharmacokinetics model estimated neuronal PYR concentrations; an endoplasmic reticulum model predicted the relative amount of HexA transported to lysosomes; and a lysosomal model simulated GM2 degradation and HexA inhibition caused by PYR. The simulations also modulated HexA half-life as a function of accumulated GM2 substrate. Previously published parameter values were used when available. Case report: A male confirmed with ATSD and the G269S mutation, had been taking a substrate reduction therapy, miglustat (200 mg, tid), for greater than four years, but at age 25 showed accelerated decline in coordination and leg muscle strength, verified by EMG evaluations. At the request of the patient and parents, PYR 25 mg QD was begun for two weeks followed by 75 mg QD in combination with folic acid (5 mg QD) to offset partial PRY dihydrofolate reductase inhibition. Weekly leukocyte HexA assays and monthly hematology tests (CBC) were conducted. Results: No noticeable side-effects were observed during the first four weeks of treatment. Additional follow-up and comparisons to the modeled response will be presented. doi:10.1016/j.ymgme.2008.11.072
72. Proteostasis Jeffery Kelly, The Scripps Research Institute, La Jolla, CA, USA