Suzanne de Bruijn

295 General discussion and perspectives discarded, differential expression levels could be detected, and in RNA-seq data novel variants could be identified that are located in or close to retained intronic sequences (pseudoexons) not covered by WES. Multi-omics Genomics Proteomics and metabolomics Epigenomics Transcriptomics Single nucleotide variants Structural variants Epigenetic profiling 3D chromosome organization Alternative splicing Gene expression levels Proteome Metabolites 55 _ 0 _ H3K27ac H3K27me3 H3K4me3 Neo-TAD Wildtype Mutant m/z Lipids Sugars Amino acids C C C C C A A A A AA GG GG G T G TA Figure 2. A schematic overview of multi-omics approaches. A multi-omics approach combines the strengths of the different omics technologies: (1) genomics allows the detection of single nucleotide variants and structural variants, (2) epigenomics allows visualization of the effect of variants on the regulatory chromosome landscape (e.g. epigenetic profiling or 3D chromosome organization) whereas (3) transcriptomics detects changes on the RNA level, this entails both qualitative changes like alternative splicing and quantitative changes that lead to up- or downregulation of gene expression. (4) Proteomics and metabolomics can be employed to investigate consequences on the proteome, aberrant protein structure or overexpression, or changes in the metabolite composition (e.g. lipids, sugars or amino acids). A whole transcriptome analysis might not even be necessary to address all genetically unexplained cases or research questions. Mini- and midi-gene in vitro splice assays in HEK293T cells, or patient-derived induced pluripotent stem cells and photoreceptor precursor cells, have proven their value in the elucidation of putative splice-modulating variants in known disease genes (e.g. ABCA4 81-83 ), and the validation of candidate disease genes such as CLRN2, implicated in autosomal recessive HL. 84 To accelerate genetic diagnostics, the implementation of (whole) transcriptome analyses should be considered in future studies, and an informed decision should be made that includes weighing all the pros and cons of the diverse technologies to select the most suitable approach.