HNRNPH1 mutations in MCL cluster near exon 4 in poly-G motifs. (A) Somatic mutations found in genomic sequencing cases and targeted sequencing coverage of a representative sample. The prevalence and pattern of mutations in HNRNPH1 is compared between DLBCL and MCL. (B) Splice site and intronic mutations affecting poly-G motifs were observed both upstream and downstream of exon 4. Paired mutations (orange triangles) are those found to be somatic by sequencing matched constitutional DNA (n = 7). Unpaired mutations (blue triangles) are mutations found in tumor-only DNA sequencing (n = 12). (C) HNRNPH1 iCLIP binding peaks show that HNRNPH1 binds near exon 4 of the transcript (shown is Ref-seq isoform NM_001257293). (D) A representative Sashimi plot of splicing events in HNRNPH1 are indicated. The canonical splicing events are shown in blue, and the alternative (exon 4 skipping) splice event is shown in green. RNA-seq splicing ratios were calculated by the sum of reads supporting the alternative (green) event, divided by the number of reads supporting the canonical (blue) splicing event. (E) Mutated HNRNPH1 cases showed significantly lower exon-skipping ratios compared with unmutated cases, as measured by RNA-seq. Cases below the dotted horizontal line (skipping ratio ≤ median mutant skipping ratio) are referred to as mutantlike in further analyses. (F) Digital PCR was used to separately quantify alternative and canonical HNRNPH1 transcripts in mutant (n = 6) and wild-type (n = 30) cases. Mutant cases exhibit lower rate of exon skipping and higher overall abundance of HNRNPH1 mRNA.