Hemophilia A (HA) is an X-linked, recessive coagulation disorder caused by lack of or a defective factor VIII (FVIII) (Salen P & Babiker HM 2019). FVIII values may be decreased in the circulation by mutations in F8 gene that affect protein synthesis and/or intracellular trafficking thus compromising its secretion (Pipe SW et al. 1996; Voorberg J et al. 1996; Roelse JC et al. 2000; Wei W et al. 2017, 2018). Nearly half of severe HA cases are caused by F8 intron 22 inversion (Inv22). This 0.6-Mb inversion splits the 186-kb F8 into two parts with opposite transcription directions (Pandey GS et al. 2013; Sauna ZE et al. 2015). The inverted 5′ part (141 kb) preserves the first 22 exons that are driven by the intrinsic F8 promoter, leading to a truncated F8 transcript due to the lack of the last 627 bp coding sequence of exons 23–26. Intron-22-inversion patients express the entire FVIII-amino-acid sequence intracellularly as 2 non-secreted polypeptides that are annotated here as F8 V2144_Y2351delinsHGVLENGKNWEPSYRW and F8 A20_M2143delinsMRIQDPGK (Pandey GS et al. 2013; Sauna ZE et al. 2015). Missense F8 variations that lead to disruption of N‐linked glycosylation attachment sites may affect the protein conformation and intracellular trafficking (Wei W et al. 2017; 2018). The F8 N601D/H/K and N2137Q mutations, which eliminate the native N-glycan of the A2 and C domains respectively dramatically decreased secretion (Wei W et al. 2017; 2018). Further, mutations near N-linked glycosylation sites, such as D599H/V, R602G, S603C/G, E600D/K, W604C/R etc., resulted in aberrant N601 glycosylation and also severely reduced the secretion of the A2 domain of FVIII, as no mutant proteins were detected in conditioned media (Wei W et al. 2017). A subgroup of missense mutations add new (ectopic) N-linked glycosylation sites to the non-B domian regions of FVIII. Investigation of all 19 such mutations found varied effects of these mutations on FVIII folding, stability, trafficking/secretion and activity, overall resulting in deficient forms of FVIII compared to the wild type protein (Wei W et al. 2018). Several other missense changes in exon 19 of the F8 gene (p.G2013R and p.R2016W) were found to cause splicing alterations thus impairing FVIII protein secretion and function (Donadon I et al. 2017). Further, F8 R2016W was quantitatively evaluated in lentiviral-mediated expression of FVIII protein and through F8 mRNA minigene and splicing assays to show that a nucleotide change may have pleiotropic effects impairing both protein biosynthesis/activity and also mRNA properties (Donadon I et al. 2017). Similar finding were reported for F8 D337N variant that also led to both splicing and synthesis abnormalities (Jourdy Y et al. 2019).
The Reactome event describes intracellular accumulation and/or decreased secretion of FVIII due to different HA-related genetic alterations spread throughout the F8 gene. The F8 variants are described in relation to changes in the protein sequence. Defective splicing events induced by F8 mutations are not shown here.