Epithelial Systems Biology Laboratory

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Vasopressin and its
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Phosphoproteomics Deep Sequencing
in Epithelia
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Integrated Antibody Design Kidney Systems Biology Project Isolated Perfused

Quantitative Mass Spectrometry

A major part of our work is the development and exploitation of tools for quantitative protein mass spectrometry (LC-MS/MS) for application to signaling mechanisms in epithelia. We are exploiting TMT-labeling, MRM and label-free approaches for native epithelia, as well as SILAC for studies in cultured epithelia. The NHLBI Proteomics Core Facility provides state-of-the-art mass spectrometers for these studies and those of other PIs in the NHLBI Intramural Program. Major focus is on quantification of cellular processes relevant to vasopressin-induced signaling in the renal collecting duct. Opportunities also exist for development of computational approaches to analysis and integration of data.


  1. Limbutara K, Kelleher A, Yang CR, Raghuram V, Knepper MA. Phosphorylation Changes in Response to Kinase Inhibitor H89 in PKA-Null Cells. Sci Rep. 2019; 9 :2814. PMID: 30808967.

  2. Sung CC, Chen L, Limbutara K, Jung HJ, Gilmer GG, Yang CR, Lin SH, Khositseth S, Chou CL, Knepper MA. RNA-Seq and protein mass spectrometry in microdissected kidney tubules reveal signaling processes initiating lithium-induced nephrogenic diabetes insipidus. Kidney Int. 2019. S0085-2538 30267-4. PMID: 31146973.

  3. Rinschen MM, Limbutara K, Knepper MA, Payne DM, Pisitkun T. From Molecules to Mechanisms: Functional Proteomics and Its Application to Renal Tubule Physiology. Physiol Rev. 2018; 98: 2571-2606. PMID: 30182799.

  4. Chou CL, Hwang G, Hageman DJ, Han L, Agrawal P, Pisitkun T, Knepper MA. Identification of UT-A1- and AQP2-interacting proteins in rat inner medullary collecting duct. Am J Physiol Cell Physiol. 2018; 314:C99-C117. PMID: 29046292.

  5. Isobe K, Jung HJ, Yang CR, Claxton J, Sandoval P, Burg MB, Raghuram V, Knepper MA. Systems-level identification of PKA-dependent signaling in epithelial cells. Proc Natl Acad Sci USA. 2017; 114:E8875-E8884. PMID: 28973931.

  6. Hwang JR, Chou CL, Medvar B, Knepper MA, Jung HJ. Identification of β-catenin-interacting proteins in nuclear fractions of native rat collecting duct cells. Am J Physiol Renal Physiol. 2017; 313:F30-F46. PMID: 28298358.

  7. LeMaire SM, Raghuram V, Grady CR, Pickering CM, Chou CL, Umejiego EN, Knepper MA. Serine/threonine phosphatases and aquaporin-2 regulation in renal collecting duct. Am J Physiol Renal Physiol. 2017; 312:F84-F95. PMID: 27784696.

  8. Huebner AR, Cheng L, Somparn P, Knepper MA, Fenton RA, Pisitkun T. Deubiquitylation of Protein cargo Is Not an Essential Step in Exosome Formation. Mol Cell Proteomics. 2016; 15:1556-71. PMID: 26884507.

  9. Cheng L, Pisitkun T, Knepper MA, Hoffert JD. Peptide Labeling Using Isobaric Tagging Reagents for Quantitative Phosphoproteomics. Methods Mol Biol. 2016; 1355:53-70. PMID: 26584918.

  10. Pickering CM, Grady C, Medvar B, Emamian M, Sandoval PC, Zhao Y, Yang CR, Jung HJ, Chou CL, Knepper MA. Proteomic profiling of nuclear fractions from native renal inner medullary collecting duct cells. Physiol Genomics. 2016; 48: 154-66. PMID: 26508704.

  11. Yang CR, Raghuram V, Emamian M, Sandoval PC, Knepper MA. Deep proteomic profiling of vasopressin-sensitive collecting duct cells. II. Bioinformatic analysis of vasopressin signaling. Am J Physiol Cell Physiol. 2015; 309:C799-812. PMID: 26310817.

  12. Yang CR, Tongyoo P, Emamian M, Sandoval PC, Raghuram V, Knepper MA. Deep proteomic profiling of vasopressin-sensitive collecting duct cells. I. Virtual Western blots and molecular weight distributions. Am J Physiol Cell Physiol. 2015; 309:C785-98. PMID: 26310816.

  13. Trepiccione F, Pisitkun T, Hoffert JD, Poulsen SB, Capasso G, Nielsen S, Knepper MA, Fenton RA, Christensen BM. Early targets of lithium in rat kidney inner medullary collecting duct include p38 and ERK1/2. Kidney Int. 2014; 86:757-67. PMID: 24786704.

  14. Bradford D, Raghuram V, Wilson JL, Chou CL, Hoffert JD, Knepper MA, Pisitkun T. Use of LC-MS/MS and Bayes' theorem to identify protein kinases that phosphorylate aquaporin-2 at Ser256. Am J Physiol Cell Physiol. 2014; 307:C123-39. PMID: 24598363.

  15. Schenk LK, Bolger SJ, Luginbuhl K, Gonzales PA, Rinschen MM, Yu MJ, Hoffert JD, Pisitkun T, Knepper MA. Quantitative proteomics identifies vasopressin-responsive nuclear proteins in collecting duct cells. J Am Soc Nephrol. 2012; 23:1008-18. PMID: 22440904.

  16. Hoffert JD, Pisitkun T, Saeed F, Song JH, Chou CL, Knepper MA. Dynamics of the G protein-coupled vasopressin V2 receptor signaling network revealed by quantitative phosphoproteomics. Mol Cell Proteomics. 2012; 11:M111.014613 PMID: 22108457.

  17. Rinschen MM, Yu MJ, Wang G, Boja ES, Hoffert JD, Pisitkun T, Knepper MA. Quantitative phosphoproteomic analysis reveals vasopressin V2 receptor-dependent signaling pathways in renal collecting duct cells. Proc Natl Acad Sci USA. 2010; 107:3882-7. PMID: 20139300.