Blood, Vol. 93 No. 4 (February 15), 1999:
pp. 1237-1244
Characterization of Two Naturally Occurring Mutations in the Second
Epidermal Growth Factor-Like Domain of Factor VII
Mathilde Hunault,
Arnaldo A. Arbini,
Josephine A. Carew,
Flora Peyvandi, and
Kenneth A. Bauer
From the Hematology-Oncology Section, Department of Medicine,
Brockton-West Roxbury Department of Veterans Affairs Medical Center,
West Roxbury, MA; Beth Israel Deaconess Medical Center, Harvard Medical
School, Boston, MA; A. Bianchi Bonomi Hemophilia and Thrombosis Center,
Department of Internal Medicine, IRCCS Ospedale Maggiore, Maggiore,
Italy; and the University of Milan, Milan, Italy
We investigated the mechanisms responsible for severe factor VII
(FVII) deficiency in homozygous Italian patients with either Gly97Cys or Gln100Arg mutations in the second
epidermal growth factor domain of FVII. Transient expression of
complementary DNA coding for the mutations in COS-1 cells showed
impaired secretion of the mutant molecules. Using stably transfected
Chinese hamster ovary (CHO) cells, we performed pulse-chase labeling
studies, immunohistochemistry, and experiments with inhibitors of
protein degradation, showing that FVII-Cys97 did not
accumulate intracellularly but was degraded in a pre-Golgi, nonlysosomal compartment by a cysteine protease. In stably transfected CHO cells expressing FVII-Arg100, the level of
intracellular FVII was not increased by several inhibitors of protein
degradation, but FVII-Arg100 was retained in the
endoplasmic reticulum for a longer period of time than wild-type FVII.
FVII-Arg100 had a lower apparent molecular weight than did
wild-type FVII under nondenaturing conditions, which is attributable to
misfolding due to abnormal disulfide bond formation.
