Splice effect of a synonymous variant in AP4B1: multiomics approach establishes the diagnosis in two sisters with...

Patients with suspected monogenic disorders often remain undiagnosed after exome sequencing. We report a family with two sisters affected by a complex spastic paraplegia. Initial exome sequencing had identified monoallelic pathogenic nonsense variants in AP4S1 and AP4B1, subunits of the adaptor protein complex 4 (AP-4), suggesting digenic inheritance. As digenic inheritance has not been established for AP-4-associated disorders, we applied a multiomics approach including genome sequencing, RNA sequencing and proteomics to clarify the genetic cause. By RNA sequencing a predicted synonymous variant (NM_006594.5:c.969G > A), compound heterozygous to the nonsense variant in AP4B1 and previously considered as benign, was re-prioritized as aberrant splicing was demonstrated. Proteomics showed reduced abundance of AP-4 components AP4B1 and AP4M1 and an upregulation of the cargo protein ATG9A, confirming AP-4 deficiency. Although the AP4S1 variant resulted in nonsense-mediated decay, the identification of biallelic causative variants in AP4B1 established the diagnosis of monogenic "Spastic paraplegia 47, autosomal recessive" while the initial hypothesis of digenic inheritance was refuted. This study illustrates the value of multiomics approaches in the diagnostic workflow of rare diseases and the potential for pathogenicity of synonymous variants.