Abstract: TH-PO534
Pin1 Isomerase Activity Determines PTH Gene Expression in Uremic Secondary Hyperparathyroidism
Session Information
- Bone and Mineral Metabolism: Basic
November 07, 2019 | Location: Exhibit Hall, Walter E. Washington Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Bone and Mineral Metabolism
- 401 Bone and Mineral Metabolism: Basic
Authors
- Hassan, Alia, Hadassah Hebrew University Medical Center, Jerusalem, Israel
- Levin, Rachel, Jerusalem College of Technology, Jerusalem, Israel
- Silver, Justin, Hadassah Hebrew University Medical Center, Jerusalem, Israel
- Nechama, Morris, Hadassah Hebrew University Medical Center, Jerusalem, Israel
- Naveh-Many, Tally, Hadassah Hebrew University Medical Center, Jerusalem, Israel
Background
Secondary hyperparathyroidism (SHP) is a common complication of CKD that correlates with morbidity and mortality. In experimental SHP there is increased PTH secretion, gene expression and parathyroid cell proliferation. The high PTH gene expression is due to increased PTH mRNA stability mediated by the balanced protein-PTH mRNA interaction of AUF1 (AU-rich binding protein 1) that stabilizes and KSRP (K-homology splicing regulatory protein) that destabilizes PTH mRNA. Pin1 binds to and isomerizes phosphorylated Ser/Thr-Pro motifs in target proteins, including mRNA binding proteins. Pin1-KSRP interaction leads to dephosphorylation of KSRP at Ser181 that then binds to PTH mRNA with higher affinity to induce PTH mRNA decay. In SHP, Pin1 isomerase activity is decreased and phosphorylated KSRP fails to bind PTH mRNA, resulting in increased PTH mRNA stability and levels. Pin1 activity is regulated by phosphorylation at Ser16 and Ser71 that disrupts its interaction with target proteins and catalytic isomerase activity.
Methods
We performed proteome and phospho-proteome analysis of parathyroids from normal and adenine high phosphorus induced CKD rats. Pin1 phosphorylation was demonstrated by immunofluorescence staining. The PKA activator forskolin or PKA inhibitor H89 were added to mouse thyroparathyroid glands in culture or HEK293 cells transfected with a PTH expression plasmid. Secreted PTH was measured by Elisa and mRNA levels by qRT-PCR.
Results
Phospho-proteome analysis confirmed KSRP hyper-phosphorylation in parathyroids of SHP rats, that would prevent PTH mRNA-KSRP binding and decay. It also identified new Pin1 targets and signaling pathways. Phosphorylation of both Pin1 Ser16 and Ser71 was increased in parathyroids of SHP rats, correlating with the decreased Pin1 activity. Accordingly, parathyroid extracts from SHP rats showed increased in vitro phosphorylation activity towards recombinant GST-Pin1. PKA activation, that leads to Pin1 Ser16 phosphorylation, increased PTH secretion in parathyroid organ cultures and PTH mRNA in transfected HEK293 cells. PKA inhibition had the opposite effect.
Conclusion
Pin1 activity is central to the pathogenesis of SHP by orchestrating PTH mRNA-protein interaction and thus mRNA decay. The resulting increased PTH mRNA stability leads to the high serum PTH levels in SHP.
Funding
- Government Support - Non-U.S.