Survival protein anoctamin-6 controls multiple platelet responses including phospholipid scrambling, swelling, and protein cleavage.
Scott syndrome is a rare bleeding disorder, characterized by altered Ca(2+)-dependent platelet signaling with defective phosphatidylserine (PS) exposure and microparticle formation, and is linked to mutations in the ANO6 gene, encoding anoctamin (Ano)6. We investigated how the complex platelet phenotype of this syndrome is linked to defective expression of Anos or other ion channels. Mice were generated with heterozygous of homozygous deficiency in Ano6, Ano1, or Ca(2+)-dependent KCa3.1 Gardos channel. Platelets from these mice were extensively analyzed on molecular functions and compared with platelets from a patient with Scott syndrome. Deficiency in Ano1 or Gardos channel did not reduce platelet responses compared with control mice (P > 0.1). In 2 mouse strains, deficiency in Ano6 resulted in reduced viability with increased bleeding time to 28.6 min (control 6.4 min, P < 0.05). Platelets from the surviving Ano6-deficient mice resembled platelets from patients with Scott syndrome in: 1) normal collagen-induced aggregate formation (P > 0.05) with reduced PS exposure (-65 to 90%); 2) lowered Ca(2+)-dependent swelling (-80%) and membrane blebbing (-90%); 3) reduced calpain-dependent protein cleavage (-60%); and 4) moderately affected apoptosis-dependent PS exposure. In conclusion, mouse deficiency of Ano6 but not of other channels affects viability and phenocopies the complex changes in platelets from hemostatically impaired patients with Scott syndrome.
*Department of Cell Biochemistry of Thrombosis and Haemostasis Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), University of Maastricht, Maastricht, The Netherlands; Department of Experimental Biomedicine, University Hospital and Rudolf Virchow Center, University of Würzburg, Würzburg, Germany; Walter Brendel Centre of Experimental Medicine and German Centre of Cardiovascular Research, Munich Heart Alliance, Ludwig-Maximilians-Universität München, München, Germany; Department of Developmental Biology, Centre for Medical Biotechnology, University of Duisburg-Essen, Duisburg-Essen, Germany; Aragon Institute of Health Sciences I+CS/IIS and ARAID, Zaragoza, Spain; School of Physiology and Pharmacology, University of Bristol, Bristol, United Kingdom; Institute of Physiology, University of Regensburg, Regensburg, Germany; **Arthur Bloom Haemophilia Centre, School of Medicine, Cardiff University, Cardiff, United Kingdom.
Nadine J A
NJ
Mattheij
*Department of Cell Biochemistry of Thrombosis and Haemostasis Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), University of Maastricht, Maastricht, The Netherlands; Department of Experimental Biomedicine, University Hospital and Rudolf Virchow Center, University of Würzburg, Würzburg, Germany; Walter Brendel Centre of Experimental Medicine and German Centre of Cardiovascular Research, Munich Heart Alliance, Ludwig-Maximilians-Universität München, München, Germany; Department of Developmental Biology, Centre for Medical Biotechnology, University of Duisburg-Essen, Duisburg-Essen, Germany; Aragon Institute of Health Sciences I+CS/IIS and ARAID, Zaragoza, Spain; School of Physiology and Pharmacology, University of Bristol, Bristol, United Kingdom; Institute of Physiology, University of Regensburg, Regensburg, Germany; **Arthur Bloom Haemophilia Centre, School of Medicine, Cardiff University, Cardiff, United Kingdom.
Attila
A
Braun
*Department of Cell Biochemistry of Thrombosis and Haemostasis Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), University of Maastricht, Maastricht, The Netherlands; Department of Experimental Biomedicine, University Hospital and Rudolf Virchow Center, University of Würzburg, Würzburg, Germany; Walter Brendel Centre of Experimental Medicine and German Centre of Cardiovascular Research, Munich Heart Alliance, Ludwig-Maximilians-Universität München, München, Germany; Department of Developmental Biology, Centre for Medical Biotechnology, University of Duisburg-Essen, Duisburg-Essen, Germany; Aragon Institute of Health Sciences I+CS/IIS and ARAID, Zaragoza, Spain; School of Physiology and Pharmacology, University of Bristol, Bristol, United Kingdom; Institute of Physiology, University of Regensburg, Regensburg, Germany; **Arthur Bloom Haemophilia Centre, School of Medicine, Cardiff University, Cardiff, United Kingdom.
Roger
R
van Kruchten
*Department of Cell Biochemistry of Thrombosis and Haemostasis Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), University of Maastricht, Maastricht, The Netherlands; Department of Experimental Biomedicine, University Hospital and Rudolf Virchow Center, University of Würzburg, Würzburg, Germany; Walter Brendel Centre of Experimental Medicine and German Centre of Cardiovascular Research, Munich Heart Alliance, Ludwig-Maximilians-Universität München, München, Germany; Department of Developmental Biology, Centre for Medical Biotechnology, University of Duisburg-Essen, Duisburg-Essen, Germany; Aragon Institute of Health Sciences I+CS/IIS and ARAID, Zaragoza, Spain; School of Physiology and Pharmacology, University of Bristol, Bristol, United Kingdom; Institute of Physiology, University of Regensburg, Regensburg, Germany; **Arthur Bloom Haemophilia Centre, School of Medicine, Cardiff University, Cardiff, United Kingdom.
Elisabetta
E
Castoldi
*Department of Cell Biochemistry of Thrombosis and Haemostasis Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), University of Maastricht, Maastricht, The Netherlands; Department of Experimental Biomedicine, University Hospital and Rudolf Virchow Center, University of Würzburg, Würzburg, Germany; Walter Brendel Centre of Experimental Medicine and German Centre of Cardiovascular Research, Munich Heart Alliance, Ludwig-Maximilians-Universität München, München, Germany; Department of Developmental Biology, Centre for Medical Biotechnology, University of Duisburg-Essen, Duisburg-Essen, Germany; Aragon Institute of Health Sciences I+CS/IIS and ARAID, Zaragoza, Spain; School of Physiology and Pharmacology, University of Bristol, Bristol, United Kingdom; Institute of Physiology, University of Regensburg, Regensburg, Germany; **Arthur Bloom Haemophilia Centre, School of Medicine, Cardiff University, Cardiff, United Kingdom.
Joachim
J
Pircher
*Department of Cell Biochemistry of Thrombosis and Haemostasis Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), University of Maastricht, Maastricht, The Netherlands; Department of Experimental Biomedicine, University Hospital and Rudolf Virchow Center, University of Würzburg, Würzburg, Germany; Walter Brendel Centre of Experimental Medicine and German Centre of Cardiovascular Research, Munich Heart Alliance, Ludwig-Maximilians-Universität München, München, Germany; Department of Developmental Biology, Centre for Medical Biotechnology, University of Duisburg-Essen, Duisburg-Essen, Germany; Aragon Institute of Health Sciences I+CS/IIS and ARAID, Zaragoza, Spain; School of Physiology and Pharmacology, University of Bristol, Bristol, United Kingdom; Institute of Physiology, University of Regensburg, Regensburg, Germany; **Arthur Bloom Haemophilia Centre, School of Medicine, Cardiff University, Cardiff, United Kingdom.
Constance C F M J
CC
Baaten
*Department of Cell Biochemistry of Thrombosis and Haemostasis Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), University of Maastricht, Maastricht, The Netherlands; Department of Experimental Biomedicine, University Hospital and Rudolf Virchow Center, University of Würzburg, Würzburg, Germany; Walter Brendel Centre of Experimental Medicine and German Centre of Cardiovascular Research, Munich Heart Alliance, Ludwig-Maximilians-Universität München, München, Germany; Department of Developmental Biology, Centre for Medical Biotechnology, University of Duisburg-Essen, Duisburg-Essen, Germany; Aragon Institute of Health Sciences I+CS/IIS and ARAID, Zaragoza, Spain; School of Physiology and Pharmacology, University of Bristol, Bristol, United Kingdom; Institute of Physiology, University of Regensburg, Regensburg, Germany; **Arthur Bloom Haemophilia Centre, School of Medicine, Cardiff University, Cardiff, United Kingdom.
Manuela
M
Wülling
*Department of Cell Biochemistry of Thrombosis and Haemostasis Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), University of Maastricht, Maastricht, The Netherlands; Department of Experimental Biomedicine, University Hospital and Rudolf Virchow Center, University of Würzburg, Würzburg, Germany; Walter Brendel Centre of Experimental Medicine and German Centre of Cardiovascular Research, Munich Heart Alliance, Ludwig-Maximilians-Universität München, München, Germany; Department of Developmental Biology, Centre for Medical Biotechnology, University of Duisburg-Essen, Duisburg-Essen, Germany; Aragon Institute of Health Sciences I+CS/IIS and ARAID, Zaragoza, Spain; School of Physiology and Pharmacology, University of Bristol, Bristol, United Kingdom; Institute of Physiology, University of Regensburg, Regensburg, Germany; **Arthur Bloom Haemophilia Centre, School of Medicine, Cardiff University, Cardiff, United Kingdom.
Marijke J E
MJ
Kuijpers
*Department of Cell Biochemistry of Thrombosis and Haemostasis Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), University of Maastricht, Maastricht, The Netherlands; Department of Experimental Biomedicine, University Hospital and Rudolf Virchow Center, University of Würzburg, Würzburg, Germany; Walter Brendel Centre of Experimental Medicine and German Centre of Cardiovascular Research, Munich Heart Alliance, Ludwig-Maximilians-Universität München, München, Germany; Department of Developmental Biology, Centre for Medical Biotechnology, University of Duisburg-Essen, Duisburg-Essen, Germany; Aragon Institute of Health Sciences I+CS/IIS and ARAID, Zaragoza, Spain; School of Physiology and Pharmacology, University of Bristol, Bristol, United Kingdom; Institute of Physiology, University of Regensburg, Regensburg, Germany; **Arthur Bloom Haemophilia Centre, School of Medicine, Cardiff University, Cardiff, United Kingdom.
Ralf
R
Köhler
*Department of Cell Biochemistry of Thrombosis and Haemostasis Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), University of Maastricht, Maastricht, The Netherlands; Department of Experimental Biomedicine, University Hospital and Rudolf Virchow Center, University of Würzburg, Würzburg, Germany; Walter Brendel Centre of Experimental Medicine and German Centre of Cardiovascular Research, Munich Heart Alliance, Ludwig-Maximilians-Universität München, München, Germany; Department of Developmental Biology, Centre for Medical Biotechnology, University of Duisburg-Essen, Duisburg-Essen, Germany; Aragon Institute of Health Sciences I+CS/IIS and ARAID, Zaragoza, Spain; School of Physiology and Pharmacology, University of Bristol, Bristol, United Kingdom; Institute of Physiology, University of Regensburg, Regensburg, Germany; **Arthur Bloom Haemophilia Centre, School of Medicine, Cardiff University, Cardiff, United Kingdom.
Alastair W
AW
Poole
*Department of Cell Biochemistry of Thrombosis and Haemostasis Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), University of Maastricht, Maastricht, The Netherlands; Department of Experimental Biomedicine, University Hospital and Rudolf Virchow Center, University of Würzburg, Würzburg, Germany; Walter Brendel Centre of Experimental Medicine and German Centre of Cardiovascular Research, Munich Heart Alliance, Ludwig-Maximilians-Universität München, München, Germany; Department of Developmental Biology, Centre for Medical Biotechnology, University of Duisburg-Essen, Duisburg-Essen, Germany; Aragon Institute of Health Sciences I+CS/IIS and ARAID, Zaragoza, Spain; School of Physiology and Pharmacology, University of Bristol, Bristol, United Kingdom; Institute of Physiology, University of Regensburg, Regensburg, Germany; **Arthur Bloom Haemophilia Centre, School of Medicine, Cardiff University, Cardiff, United Kingdom.
Rainer
R
Schreiber
*Department of Cell Biochemistry of Thrombosis and Haemostasis Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), University of Maastricht, Maastricht, The Netherlands; Department of Experimental Biomedicine, University Hospital and Rudolf Virchow Center, University of Würzburg, Würzburg, Germany; Walter Brendel Centre of Experimental Medicine and German Centre of Cardiovascular Research, Munich Heart Alliance, Ludwig-Maximilians-Universität München, München, Germany; Department of Developmental Biology, Centre for Medical Biotechnology, University of Duisburg-Essen, Duisburg-Essen, Germany; Aragon Institute of Health Sciences I+CS/IIS and ARAID, Zaragoza, Spain; School of Physiology and Pharmacology, University of Bristol, Bristol, United Kingdom; Institute of Physiology, University of Regensburg, Regensburg, Germany; **Arthur Bloom Haemophilia Centre, School of Medicine, Cardiff University, Cardiff, United Kingdom.
Andrea
A
Vortkamp
*Department of Cell Biochemistry of Thrombosis and Haemostasis Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), University of Maastricht, Maastricht, The Netherlands; Department of Experimental Biomedicine, University Hospital and Rudolf Virchow Center, University of Würzburg, Würzburg, Germany; Walter Brendel Centre of Experimental Medicine and German Centre of Cardiovascular Research, Munich Heart Alliance, Ludwig-Maximilians-Universität München, München, Germany; Department of Developmental Biology, Centre for Medical Biotechnology, University of Duisburg-Essen, Duisburg-Essen, Germany; Aragon Institute of Health Sciences I+CS/IIS and ARAID, Zaragoza, Spain; School of Physiology and Pharmacology, University of Bristol, Bristol, United Kingdom; Institute of Physiology, University of Regensburg, Regensburg, Germany; **Arthur Bloom Haemophilia Centre, School of Medicine, Cardiff University, Cardiff, United Kingdom.
Peter W
PW
Collins
*Department of Cell Biochemistry of Thrombosis and Haemostasis Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), University of Maastricht, Maastricht, The Netherlands; Department of Experimental Biomedicine, University Hospital and Rudolf Virchow Center, University of Würzburg, Würzburg, Germany; Walter Brendel Centre of Experimental Medicine and German Centre of Cardiovascular Research, Munich Heart Alliance, Ludwig-Maximilians-Universität München, München, Germany; Department of Developmental Biology, Centre for Medical Biotechnology, University of Duisburg-Essen, Duisburg-Essen, Germany; Aragon Institute of Health Sciences I+CS/IIS and ARAID, Zaragoza, Spain; School of Physiology and Pharmacology, University of Bristol, Bristol, United Kingdom; Institute of Physiology, University of Regensburg, Regensburg, Germany; **Arthur Bloom Haemophilia Centre, School of Medicine, Cardiff University, Cardiff, United Kingdom.
Bernhard
B
Nieswandt
*Department of Cell Biochemistry of Thrombosis and Haemostasis Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), University of Maastricht, Maastricht, The Netherlands; Department of Experimental Biomedicine, University Hospital and Rudolf Virchow Center, University of Würzburg, Würzburg, Germany; Walter Brendel Centre of Experimental Medicine and German Centre of Cardiovascular Research, Munich Heart Alliance, Ludwig-Maximilians-Universität München, München, Germany; Department of Developmental Biology, Centre for Medical Biotechnology, University of Duisburg-Essen, Duisburg-Essen, Germany; Aragon Institute of Health Sciences I+CS/IIS and ARAID, Zaragoza, Spain; School of Physiology and Pharmacology, University of Bristol, Bristol, United Kingdom; Institute of Physiology, University of Regensburg, Regensburg, Germany; **Arthur Bloom Haemophilia Centre, School of Medicine, Cardiff University, Cardiff, United Kingdom.
Karl
K
Kunzelmann
*Department of Cell Biochemistry of Thrombosis and Haemostasis Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), University of Maastricht, Maastricht, The Netherlands; Department of Experimental Biomedicine, University Hospital and Rudolf Virchow Center, University of Würzburg, Würzburg, Germany; Walter Brendel Centre of Experimental Medicine and German Centre of Cardiovascular Research, Munich Heart Alliance, Ludwig-Maximilians-Universität München, München, Germany; Department of Developmental Biology, Centre for Medical Biotechnology, University of Duisburg-Essen, Duisburg-Essen, Germany; Aragon Institute of Health Sciences I+CS/IIS and ARAID, Zaragoza, Spain; School of Physiology and Pharmacology, University of Bristol, Bristol, United Kingdom; Institute of Physiology, University of Regensburg, Regensburg, Germany; **Arthur Bloom Haemophilia Centre, School of Medicine, Cardiff University, Cardiff, United Kingdom.
Judith M E M
JM
Cosemans
*Department of Cell Biochemistry of Thrombosis and Haemostasis Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), University of Maastricht, Maastricht, The Netherlands; Department of Experimental Biomedicine, University Hospital and Rudolf Virchow Center, University of Würzburg, Würzburg, Germany; Walter Brendel Centre of Experimental Medicine and German Centre of Cardiovascular Research, Munich Heart Alliance, Ludwig-Maximilians-Universität München, München, Germany; Department of Developmental Biology, Centre for Medical Biotechnology, University of Duisburg-Essen, Duisburg-Essen, Germany; Aragon Institute of Health Sciences I+CS/IIS and ARAID, Zaragoza, Spain; School of Physiology and Pharmacology, University of Bristol, Bristol, United Kingdom; Institute of Physiology, University of Regensburg, Regensburg, Germany; **Arthur Bloom Haemophilia Centre, School of Medicine, Cardiff University, Cardiff, United Kingdom jwm.heemskerk@maastrichtuniversity.nl.
Johan W M
JW
Heemskerk
D000818:Animals; D000075369:Anoctamin-1; D000075368:Anoctamins; D001778:Blood Coagulation Disorders / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D001792:Blood Platelets / Q000378:metabolism* / Q000473:pathology; D002118:Calcium / Q000378:metabolism; D002462:Cell Membrane / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D018118:Chloride Channels / Q000235:genetics / Q000378:metabolism; D005260:Female; D006801:Humans; D051660:Intermediate-Conductance Calcium-Activated Potassium Channels / Q000235:genetics / Q000378:metabolism; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D009154:Mutation; D009363:Neoplasm Proteins / Q000235:genetics / Q000378:metabolism; D049294:Phospholipid Transfer Proteins / Q000235:genetics / Q000378:metabolism*; D010743:Phospholipids / Q000235:genetics / Q000378:metabolism*; D059748:Proteolysis
C476213:ANO1 protein, human; C531181:ANO1 protein, mouse; C576687:ANO6 protein, mouse; C555078:ANO6 protein, human; D000075369:Anoctamin-1; D000075368:Anoctamins; D018118:Chloride Channels; D051660:Intermediate-Conductance Calcium-Activated Potassium Channels; C496923:KCNN4 protein, human; C496925:Kcnn4 protein, mouse; D009363:Neoplasm Proteins; D049294:Phospholipid Transfer Proteins; D010743:Phospholipids; D002118:Calcium
Scott syndrome; TMEM16F; bleeding; embryonic lethality; phosphatidylserine