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Coagulation & its Disorders
Christmas disease in a Hovawart family resembling human hemophilia B Leyden is caused by a single nucleotide deletion in a highly conserved transcription factor binding site of the F9 gene promoter
Ferrata Storti Foundation
Haematologica 2019 Volume 104(11):2307-2313
Bertram Brenig,1* Lilith Steingräber1*, Shuwen Shan,1 Fangzheng Xu,1 Marc Hirschfeld,1,2 Reiner Andag,3 Mirjam Spengeler,4 Elisabeth Dietschi,4 Reinhard Mischke5 and Tosso Leeb4
1University of Göttingen, Institute of Veterinary Medicine, Göttingen, Germany; 2Department of Obstetrics and Gynecology, Freiburg University Medical Center, Freiburg, Germany; 3University Medical Center Göttingen, Institute for Clinical Chemistry, Göttingen, Germany; 4Institute of Genetics, University of Bern, Bern, Switzerland and 5Small Animal Clinic, University of Veterinary Medicine Hannover Foundation, Hannover, Germany
*BB and LS contributed equally to this work.
ABSTRACT
Hemophilia B is a classical monogenic, X-chromosomal, recessively transmitted bleeding disorder caused by genetic variants within the coagulation factor IX gene (F9). Although hemophilia B has been described in dogs, it has not yet been reported in the Hovawart breed. Here we describe the identification of a Hovawart family transmitting typical signs of an X-linked bleeding disorder. Five males were reported to suffer from recurrent hemorrhagic episodes. A blood sample from one of these males with only 2% of the normal concentration of plasma factor IX together with samples from seven relatives were provided. Next-generation sequencing of the mother and grandmother revealed a single nucleotide deletion in the F9 promoter. Genotyping of the deletion in 1,298 dog spec- imens including 720 Hovawarts revealed that the mutant allele was only present in the aforementioned Hovawart family. The deletion is located 73 bp upstream of the F9 start codon in the conserved overlapping DNA bind- ing sites of hepatocyte nuclear factor 4α (HNF-4α) and androgen receptor (AR). The deletion only abolished binding of HNF-4α, while AR binding was unaffected as demonstrated by electrophoretic mobility shift assay using human HNF-4α and AR with double-stranded DNA probes encom- passing the mutant promoter region. Luciferase reporter assays using wild- type and mutated promoter fragment constructs transfected into Hep G2 cells showed a significant reduction in expression from the mutant promot- er. The data provide evidence that the deletion in the Hovawart family caused a rare type of hemophilia B resembling human hemophilia B Leyden.
Introduction
Hemophilia B (Christmas disease) is a recessive, X-linked bleeding disorder caused by genetic variants within the clotting factor IX gene (F9) resulting in the absence or insufficient levels of factor IX (FIX) in the blood.1 In humans hemophilia B is also known as the “royal disease” as it was transmitted into several European royal dynasties by Queen Victoria.2,3 As of present, 1,113 unique F9 variants have been described in humans.4 The majority of the pathogenic variants are located within exons (n=923) and intronic regions (n=137) of F9. Only 33 variants (2.96%) have been described in the 5’-UTR (n=28) and 3’-UTR (n=5) accounting for 2.52% and 0.45% of human pathogenic hemophilia B variants, respectively.4
Although the first reports about canine hemophilia B date back to the early 1960s
Correspondence:
BERTRAM BRENIG
bbrenig@gwdg.de
Received: December 26, 2018 Accepted: March 6, 2019. Pre-published: March 7, 2019.
doi:10.3324/haematol.2018.215426
Check the online version for the most updated information on this article, online supplements, and information on authorship & disclosures: www.haematologica.org/content/104/11/2307
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haematologica | 2019; 104(11)
2307
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