The VPS33B-binding protein VPS16B is required in megakaryocyte and platelet α-granule biogenesis

Denisa Urban, Ng Li, Hilary Christensen, Fred G. Pluthero, Shao Zun Chen, Michael Puhacz, Parvesh M. Garg, Kiran K. Lanka, James J. Cummings, Helmut Kramer, James D. Wasmuth, John Parkinson, Walter H A Kahr

Research output: Contribution to journalArticle

52 Scopus citations

Abstract

Patients with platelet α or dense δ-granule defects have bleeding problems. Although several proteins are known to be required for δ-granule development, less is known about α-granule biogenesis. Our previous work showed that the BEACH protein NBEAL2 and the Sec1/Munc18 protein VPS33B are required for α-granule biogenesis. Using a yeast two-hybrid screen, mass spectrometry, coimmunoprecipitation, and bioinformatics studies, we identified VPS16B as a VPS33B-binding protein. Immunoblotting confirmed VPS16B expression in various human tissues and cells including megakaryocytes and platelets, and also in megakaryocytic Dami cells. Characterization of platelets from a patient with arthrogryposis, renal dysfunction, and cholestasis (ARC) syndrome containing mutations in C14orf133 encoding VPS16B revealed pale-appearing platelets in blood films and electron microscopy revealed a complete absence of α-granules, whereas δ-granules were observed. Soluble and membrane-bound α-granule proteins were reduced or undetectable, suggesting that both releasable and membranebound α-granule constituents were absent. Immunofluorescence microscopy of Dami cells stably expressing GFP-VPS16B revealed that similar to VPS33B, GFPVPS16B colocalized with markers of the trans-Golgi network, late endosomes and α-granules. We conclude that VPS16B, similar to its binding partner VPS33B, is essential for megakaryocyte and platelet α-granule biogenesis.

Original languageEnglish (US)
Pages (from-to)5032-5040
Number of pages9
JournalBlood
Volume120
Issue number25
DOIs
StatePublished - Dec 13 2012

ASJC Scopus subject areas

  • Biochemistry
  • Immunology
  • Hematology
  • Cell Biology

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    Urban, D., Li, N., Christensen, H., Pluthero, F. G., Chen, S. Z., Puhacz, M., Garg, P. M., Lanka, K. K., Cummings, J. J., Kramer, H., Wasmuth, J. D., Parkinson, J., & Kahr, W. H. A. (2012). The VPS33B-binding protein VPS16B is required in megakaryocyte and platelet α-granule biogenesis. Blood, 120(25), 5032-5040. https://doi.org/10.1182/blood-2012-05-431205