A parathyroid hormone/salt-inducible kinase signaling axis controls renal vitamin D activation and organismal calcium homeostasis
Sung-Hee Yoon, … , Michael Mannstadt, Marc N. Wein
Sung-Hee Yoon, … , Michael Mannstadt, Marc N. Wein
Published March 2, 2023
Citation Information: J Clin Invest. 2023;133(9):e163627. https://doi.org/10.1172/JCI163627.
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Research Article Bone biology Nephrology

A parathyroid hormone/salt-inducible kinase signaling axis controls renal vitamin D activation and organismal calcium homeostasis

Abstract

The renal actions of parathyroid hormone (PTH) promote 1,25-vitamin D generation; however, the signaling mechanisms that control PTH-dependent vitamin D activation remain unknown. Here, we demonstrated that salt-inducible kinases (SIKs) orchestrated renal 1,25-vitamin D production downstream of PTH signaling. PTH inhibited SIK cellular activity by cAMP-dependent PKA phosphorylation. Whole-tissue and single-cell transcriptomics demonstrated that both PTH and pharmacologic SIK inhibitors regulated a vitamin D gene module in the proximal tubule. SIK inhibitors increased 1,25-vitamin D production and renal Cyp27b1 mRNA expression in mice and in human embryonic stem cell–derived kidney organoids. Global- and kidney-specific Sik2/Sik3 mutant mice showed Cyp27b1 upregulation, elevated serum 1,25-vitamin D, and PTH-independent hypercalcemia. The SIK substrate CRTC2 showed PTH and SIK inhibitor–inducible binding to key Cyp27b1 regulatory enhancers in the kidney, which were also required for SIK inhibitors to increase Cyp27b1 in vivo. Finally, in a podocyte injury model of chronic kidney disease–mineral bone disorder (CKD-MBD), SIK inhibitor treatment stimulated renal Cyp27b1 expression and 1,25-vitamin D production. Together, these results demonstrated a PTH/SIK/CRTC signaling axis in the kidney that controls Cyp27b1 expression and 1,25-vitamin D synthesis. These findings indicate that SIK inhibitors might be helpful for stimulation of 1,25-vitamin D production in CKD-MBD.

Authors

Sung-Hee Yoon, Mark B. Meyer, Carlos Arevalo, Murat Tekguc, Chengcheng Zhang, Jialiang S. Wang, Christian D. Castro Andrade, Katelyn Strauss, Tadatoshi Sato, Nancy A. Benkusky, Seong Min Lee, Rebecca Berdeaux, Marc Foretz, Thomas B. Sundberg, Ramnik J. Xavier, Charles H. Adelmann, Daniel J. Brooks, Anthony Anselmo, Ruslan I. Sadreyev, Ivy A. Rosales, David E. Fisher, Navin Gupta, Ryuji Morizane, Anna Greka, J. Wesley Pike, Michael Mannstadt, Marc N. Wein

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Figure 3

PTH and SIK inhibitors increase Cyp27b1 expression and 1,25-vitamin D synthesis in human kidney organoids.

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PTH and SIK inhibitors increase Cyp27b1 expression and 1,25-vitamin D sy...
(A) H9 human embryonic stem cell–derived kidney organoids were treated with TMR-labeled PTH and then stained with LTL. (B) Kidney organoids were treated with PTH1-34 for 30 minutes and immunostained with pPKA substrate antibody to show PTH responsiveness (Lotus Tetragonolobus Lectin [LTL] for proximal tubule; CDH1 for distal tubule; podocalyxin [PODXL] for podocytes; Sytox Blue as the nuclear dye). Scale bars: 100 μm. Quantification is shown in the histogram on the right. Student’s t test was performed for statistical analysis. (C) Day 28 organoids were treated for the indicated times with vehicle or PTH (242 nM), and 1,25-vitamin D was measured in the culture media by ELISA. (D) PTH-induced 1,25-vitamin D production was inhibited by cotreatment with FGF23 (100 nM). (E–G) Cyp27b1 gene expression was measured by RT-qPCR in response to treatment for the indicated times with PTH (242 nM) and SIK inhibitors, YKL-05-099 (10 μM) and SK-124 (20 μM). (H) Similar experiments in which 1,25-vitamin D was measured in the culture media by ELISA. (I) Pretreatment with H89 (10 μM), YM-254890 (10 μM), and bisindolylmaleimide I (10 μM) for 1 hour prior to 4-hour PTH treatment (242 nM) showed that cAMP/PKA pathway is responsible for PTH-induced Cyp27b1 upregulation. Data are shown as the mean ± SD, and each dot represents independent sample of 3–4 organoids pooled together. For C, Student’s t test were performed between PTH and Veh at each time point. For D, Student’s t test was performed against Veh (indicated by ***P < 0.001, ****P < 0.0001) and to test the effect of FGF23 (indicated by ##P < 0.001). For E–H, 1-way ANOVA was performed, with Dunnett’s post hoc test, against Veh (*P < 0.05; **P < 0.01, ***P < 0.001, ****P < 0.0001).