Stable RNA interference of MMSET I and MMSET II isoforms in t(4;14) + MM cells. (A) Structure of transcripts originating from the MMSET locus. Boxes indicate protein-coding exons, beginning with exon 3 for MMSET I and II. Lines indicate 5′ and 3′ untranslated regions (not shown to scale). MMSET II splices from exon 10 to exons 12-24, whereas MMSET I splices from exon 10 to exon 11, which is shaded gray. The untranslated exons 10 and 12-14 of REIIBP are stippled. REIIBP translation begins in exon 15. Arrows indicate major translation initiation sites. The conserved protein domains PWWP, HMG, PHD, and SET are indicated above the corresponding exons. Thick bars indicate positions of MMSET shRNA 1 (black) and control (gray) short hairpin RNAs. (B) Real-time quantitative RT-PCR analysis of MMSET I, MMSET II, and REIIBP transcripts in KMS-11 cells stably transduced with MMSET shRNA lentivirus or control shRNA lentivirus. Transcript levels are relative to empty vector–transduced cells. Results are means (± SD) of at least 2 independent lentiviral infections (*P < .05, †P = .45 by Student t test). (C) Immunoblotting with antisera recognizing a C-terminal epitope of MMSET demonstrates reduction of full-length MMSET protein by 3 different shRNAs in KMS-11 (lanes 1-5) and NCI-H929 (lanes 6-10). The top arrow indicates full-length MMSET II, translated from the first ATG codon in exon 3. The lower arrows indicate MMSET II isoforms consistent with translation from downstream ATG codons in exons 4 and 6 in NCI-H929 cells, which have a translocation breakpoint between exons 3 and 4.