Blood, Vol. 92 No. 12 (December 15), 1998:
pp. 4671-4676
Impaired Cotranslational Processing as a Mechanism for Type I
Antithrombin Deficiency
Alison C. Fitches,
Ruth Appleby,
David A. Lane,
Valerio De
Stefano,
Giuseppe Leone, and
Robin J. Olds
From the Department of Pathology, Dunedin School of Medicine,
University of Otago, Dunedin, New Zealand; the Department of
Haematology, Imperial College School of Medicine, Charing Cross Campus,
London, UK; and the Department of Haematology, Catholic University,
Rome, Italy.
Most secretory proteins, including antithrombin (AT), are
synthesized with a signal peptide, which is cleaved before the mature protein is exported from the cell. The signal peptide is important in
the process whereby nascent protein is recognized as requiring subsequent modification within the endoplasmic reticulum (ER). We have
identified a novel mutation, 2436T
C L(-10)P, which affects the central hydrophobic domain of the AT signal peptide, in a proband
presenting with venous thrombotic disease and type I AT deficiency. We
investigated the basis of the phenotype by examining expression in
mammalian cells of a range of variant AT cDNAs with mutations affecting
the -10 residue. Glycosylated AT was secreted from COS-7 cells
transfected with wild-type AT, -10L deletion, -10V or -10M variants,
but not variants with P, T, R, or G at -10. Cell-free expression of
wild-type and variant AT cDNAs was then performed in the presence of
canine pancreatic microsomes, as a substitute for ER. Variant AT
proteins with P, T, R, or G at residue -10 did not undergo
posttranslational glycosylation, and their susceptibility to trypsin
digestion suggested they had not been translocated into microsomes. Our
results suggest that the ability of AT signal peptide to direct the
protein to ER for cotranslational processing events appears to be
critically dependent on maintaining the hydrophobic nature of the
region including residue -10. The investigations have defined impaired
cotranslational processing as a hitherto unrecognized cause of
hereditary AT deficiency.
