Systematic evaluation of MANE Plus Clinical transcripts: implications for genomic diagnostics

Differences between RefSeq and Ensembl/GENCODE transcript annotations have complicated data harmonisation, leading to the MANE project that defined a single MANE Select (MS) transcript per gene, supplemented by MANE Plus Clinical (MPC) transcripts for clinically-relevant alternative isoforms. However, the diagnostic impact of relying solely on MS versus also considering MPC transcripts has not been systematically evaluated. UCSC Genome Browser multi-region visualisation and interval-based analyses was used to compare transcript structures and coding differences for 65 genes in MANE v1.4 with both MS and MPC transcripts. The median unique coding sequence per MS/MPC pair was 115bp, but two outliers ( DST, SYNE1 ) had > 20kb specific to the MS. Data from gnomAD indicated that MPC-specific exons in BRAF, SYNE1 and REEP6 have reduced exome coverage, highlighting potential diagnostic blind spots. To assess clinical relevance, data from ClinVar, 100kGP and the Genome Medicine Service were evaluated and ~ 2.5% of (likely) pathogenic variants mapped exclusively to MPC-specific coding sequence, rising to ≥ 25% for 7/65 genes. For IQSEC2, MOCS2 and CDKN2A , MS and MPC transcripts encode entirely distinct proteins. Overlapping, opposing-strand start codons in MUTYH/TOE1 and seven other gene-pairs suggest novel mechanisms for phenotypic blending. Analysis of >4M ClinVar variants identified clinically-relevant exons outside current MANE annotations, notably an alternative cytoplasmic domain exon in PCDH15 harbouring 14 (likely) pathogenic variants, supporting inclusion of additional transcripts in future releases. In conclusion, transcript selection profoundly influences clinical variant interpretation, and we demonstrate the utility of UCSC multi-region visualisation to support transcript-aware analysis across diagnostics, research and education.