Profiling of Mutations in the F8 and F9, Causative Genes of Hemophilia A and Hemophilia B

Hemophilia, a common congenital coagulation disorder, is classified as hemophilia A (HA) and hemophilia B (HB), which result from a deficiency or dysfunction of coagulation factor VIII (FVIII) and factor IX (FIX), respectively. HA is known to be caused by heterogeneous mutations of the FVIII gene (F8), such as inversions, substitutions, deletions, insertions, etc. F8 (NM_000132.3) is located on the long arm of the Xq28 region of the X chromosome. F8 is extremely large (186 kb) and consists of 26 exons (Graw et al., 2005). The transcript of F8 is approximately 9010 bp and comprises a short 5′-untranslated region (5′-UTR; 150 bp), an open reading frame (ORF) plus stop codon (7056 bp), and a long 3′-UTR (1806 bp). The protein product of F8 is a cofactor of FIX, without enzyme activity. The ORF encodes a signal peptide with 19 amino acids at its N-terminus, which leads to the passage of FVIII through hepatocytes to blood vessels. The matured FVIII protein contains 2332 amino acids and a glycoprotein of approximately 250 kDa, and circulates as an inactive pro-cofactor. FVIII is a multi-domain protein composed of A1-A2-B-A3-C1-C2, named from the Nterminus. FVIII synthesized in hepatocytes is secreted into the circulation and readily assembled with von Willebrand factor (vWF), which is generated and secreted by endothelial cells. Besides vWF, FVIII protein can also interact with diverse proteins such as thrombin and FX. These interactions are important for effective hemostasis. However, F8 mutations can lead to the production of truncated proteins, which lead to disruption of FVIII function and suppress normal protein interaction with proteins involved in the coagulation cascade (Bowen, 2002). This inappropriate reaction causes bleeding tendency. F8 mutations can occur at diverse sites in a variety of types, such as structural variation (inversions of intron 22 or intron 1) and sequence variation (insertion, deletion, and substitution). The latter variation leads to nonsense, missense, and frameshift mutations. Recently, more than 1,200 types of F8 mutations were reported in the HAMSTeRS (Hemophilia A Mutation, Structure, Test and Resource Site) database (http://hadb.org.uk). The F9 gene (NM_000133.3) is also located on the X chromosome at Xq27.1–q27.2. In contrst to F8, the size of F9 gene is approximately 34 kb with only eight exons and the size of the transcript mRNA is 2803 bp. The F9 gene encodes the FIX protein, one of the vitamin