VPS33B regulates protein sorting into and maturation of α-granule progenitor organelles in mouse megakaryocytes

Danai Bem; Holly Smith; Blerida Banushi; Jemima J Burden; Ian J White; Joanna Hanley; Nadia Jeremiah; Frédéric Rieux-Laucat; Ruth Bettels; Gema Ariceta; Andrew D Mumford; Steven G Thomas; Steve P Watson; Paul Gissen

doi:10.1182/blood-2014-12-614677

VPS33B regulates protein sorting into and maturation of α-granule progenitor organelles in mouse megakaryocytes

Danai Bem, Holly Smith, Blerida Banushi, Jemima J Burden, Ian J White, Joanna Hanley, Nadia Jeremiah, Frédéric Rieux-Laucat, Ruth Bettels, Gema Ariceta, Andrew D Mumford, Steven G Thomas, Steve P Watson, Paul Gissen

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Abstract

Arthrogryposis, Renal dysfunction and Cholestasis (ARC) syndrome is caused by deficiencies in the trafficking proteins VPS33B or VIPAR, and is associated with a bleeding diathesis and a marked reduction in platelet α-granules. We generated a tamoxifen-inducible mouse model of VPS33B deficiency, Vps33b(fl/fl)-ER(T2), and studied the platelet phenotype and α-granule biogenesis. Ultrastructural analysis of Vps33b(fl/fl)-ER(T2) platelets identified a marked reduction in α-granule count and the presence of small granule-like structures in agreement with the platelet phenotype observed in ARC patients. A reduction in the order of 65-75% was observed in the α-granule proteins von Willebrand factor and P-selectin. Although platelet aggregation responses were not affected, a defect in δ-granule secretion was observed. Under arteriolar shear conditions, Vps33b(fl/fl)-ER(T2) platelets were unable to form stable aggregates, and tail bleeding measurement revealed a bleeding diathesis. Analysis of bone marrow-derived megakaryocytes by conventional and immunoelectron microscopy from Vps33b(fl/fl)-ER(T2) mice revealed a reduction in mature type-II multivesicular bodies (MVB II) and an accumulation of large vacuoles. Proteins that are normally stored in α-granules were under-represented in MVB II and proplatelet extensions. These results demonstrate that abnormal protein trafficking and impairment in MVB maturation in megakaryocytes underlie the α-granule deficiency in Vps33b(fl/fl)-ER(T2) mouse and ARC patients.

Original language	English
Journal	Blood
DOIs	https://doi.org/10.1182/blood-2014-12-614677
Publication status	Published - 6 May 2015

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@article{c15e86e286254ab1a3bb9f8c2341a9ab,

title = "VPS33B regulates protein sorting into and maturation of α-granule progenitor organelles in mouse megakaryocytes",

abstract = "Arthrogryposis, Renal dysfunction and Cholestasis (ARC) syndrome is caused by deficiencies in the trafficking proteins VPS33B or VIPAR, and is associated with a bleeding diathesis and a marked reduction in platelet α-granules. We generated a tamoxifen-inducible mouse model of VPS33B deficiency, Vps33b(fl/fl)-ER(T2), and studied the platelet phenotype and α-granule biogenesis. Ultrastructural analysis of Vps33b(fl/fl)-ER(T2) platelets identified a marked reduction in α-granule count and the presence of small granule-like structures in agreement with the platelet phenotype observed in ARC patients. A reduction in the order of 65-75\% was observed in the α-granule proteins von Willebrand factor and P-selectin. Although platelet aggregation responses were not affected, a defect in δ-granule secretion was observed. Under arteriolar shear conditions, Vps33b(fl/fl)-ER(T2) platelets were unable to form stable aggregates, and tail bleeding measurement revealed a bleeding diathesis. Analysis of bone marrow-derived megakaryocytes by conventional and immunoelectron microscopy from Vps33b(fl/fl)-ER(T2) mice revealed a reduction in mature type-II multivesicular bodies (MVB II) and an accumulation of large vacuoles. Proteins that are normally stored in α-granules were under-represented in MVB II and proplatelet extensions. These results demonstrate that abnormal protein trafficking and impairment in MVB maturation in megakaryocytes underlie the α-granule deficiency in Vps33b(fl/fl)-ER(T2) mouse and ARC patients.",

author = "Danai Bem and Holly Smith and Blerida Banushi and Burden, \{Jemima J\} and White, \{Ian J\} and Joanna Hanley and Nadia Jeremiah and Fr{\'e}d{\'e}ric Rieux-Laucat and Ruth Bettels and Gema Ariceta and Mumford, \{Andrew D\} and Thomas, \{Steven G\} and Watson, \{Steve P\} and Paul Gissen",

note = "Copyright {\textcopyright} 2015 American Society of Hematology.",

year = "2015",

month = may,

day = "6",

doi = "10.1182/blood-2014-12-614677",

language = "English",

journal = "Blood",

issn = "0006-4971",

publisher = "Elsevier B.V.",

}

TY - JOUR

T1 - VPS33B regulates protein sorting into and maturation of α-granule progenitor organelles in mouse megakaryocytes

AU - Bem, Danai

AU - Smith, Holly

AU - Banushi, Blerida

AU - Burden, Jemima J

AU - White, Ian J

AU - Hanley, Joanna

AU - Jeremiah, Nadia

AU - Rieux-Laucat, Frédéric

AU - Bettels, Ruth

AU - Ariceta, Gema

AU - Mumford, Andrew D

AU - Thomas, Steven G

AU - Watson, Steve P

AU - Gissen, Paul

PY - 2015/5/6

Y1 - 2015/5/6

N2 - Arthrogryposis, Renal dysfunction and Cholestasis (ARC) syndrome is caused by deficiencies in the trafficking proteins VPS33B or VIPAR, and is associated with a bleeding diathesis and a marked reduction in platelet α-granules. We generated a tamoxifen-inducible mouse model of VPS33B deficiency, Vps33b(fl/fl)-ER(T2), and studied the platelet phenotype and α-granule biogenesis. Ultrastructural analysis of Vps33b(fl/fl)-ER(T2) platelets identified a marked reduction in α-granule count and the presence of small granule-like structures in agreement with the platelet phenotype observed in ARC patients. A reduction in the order of 65-75% was observed in the α-granule proteins von Willebrand factor and P-selectin. Although platelet aggregation responses were not affected, a defect in δ-granule secretion was observed. Under arteriolar shear conditions, Vps33b(fl/fl)-ER(T2) platelets were unable to form stable aggregates, and tail bleeding measurement revealed a bleeding diathesis. Analysis of bone marrow-derived megakaryocytes by conventional and immunoelectron microscopy from Vps33b(fl/fl)-ER(T2) mice revealed a reduction in mature type-II multivesicular bodies (MVB II) and an accumulation of large vacuoles. Proteins that are normally stored in α-granules were under-represented in MVB II and proplatelet extensions. These results demonstrate that abnormal protein trafficking and impairment in MVB maturation in megakaryocytes underlie the α-granule deficiency in Vps33b(fl/fl)-ER(T2) mouse and ARC patients.

AB - Arthrogryposis, Renal dysfunction and Cholestasis (ARC) syndrome is caused by deficiencies in the trafficking proteins VPS33B or VIPAR, and is associated with a bleeding diathesis and a marked reduction in platelet α-granules. We generated a tamoxifen-inducible mouse model of VPS33B deficiency, Vps33b(fl/fl)-ER(T2), and studied the platelet phenotype and α-granule biogenesis. Ultrastructural analysis of Vps33b(fl/fl)-ER(T2) platelets identified a marked reduction in α-granule count and the presence of small granule-like structures in agreement with the platelet phenotype observed in ARC patients. A reduction in the order of 65-75% was observed in the α-granule proteins von Willebrand factor and P-selectin. Although platelet aggregation responses were not affected, a defect in δ-granule secretion was observed. Under arteriolar shear conditions, Vps33b(fl/fl)-ER(T2) platelets were unable to form stable aggregates, and tail bleeding measurement revealed a bleeding diathesis. Analysis of bone marrow-derived megakaryocytes by conventional and immunoelectron microscopy from Vps33b(fl/fl)-ER(T2) mice revealed a reduction in mature type-II multivesicular bodies (MVB II) and an accumulation of large vacuoles. Proteins that are normally stored in α-granules were under-represented in MVB II and proplatelet extensions. These results demonstrate that abnormal protein trafficking and impairment in MVB maturation in megakaryocytes underlie the α-granule deficiency in Vps33b(fl/fl)-ER(T2) mouse and ARC patients.

U2 - 10.1182/blood-2014-12-614677

DO - 10.1182/blood-2014-12-614677

M3 - Article (Academic Journal)

C2 - 25947942

SN - 0006-4971

JO - Blood

JF - Blood

ER -

VPS33B regulates protein sorting into and maturation of α-granule progenitor organelles in mouse megakaryocytes

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