N-Terminal deleted isoforms of E3 ligase RNF220 are ubiquitously expressed and required for mouse muscle differentiation.

Four isoform peptides of the novel E3 ligase ring finger protein 220 (RNF220) have been identified in humans. However, all of the previous studies have predominantly focused on isoform 1 (the full-length form), which consists of 566 amino acids. Here, we show that a shorter isoform, which is 308 amino acids lacking most of the N-terminus (human isoform 4; mouse isoform 3; ΔN-RNF220), is the predominant and ubiquitously expressed variant that warrants functional investigation. Both isoform 1 and ΔN-RNF220 are expressed in the brain; however, ΔN-RNF220 is the major isoform expressed in all other tissues in mice. Consistently, H3K4me3 ChIP-seq data from ENCODE reveal that the transcription start site for ΔN-RNF220 demonstrates broader and stronger activity across human tissues than that of isoform 1. ΔN-RNF220 produces 2 peptides (4a and 4b) through alternative translation initiation, with isoform 4b displaying distinct subcellular localization, subnuclear structures and interaction with a nuclear protein WDR5. Notably, during embryonic stem cell differentiation into neural stem cells, isoform 1 expression increases, whereas ΔN-RNF220 expression decreases. In murine myoblasts, ΔN-RNF220 is the sole expressed isoform and is required for MyoD and myogenin expression, as well as for muscle differentiation. Our findings highlight ΔN-RNF220 as the ubiquitously and highly expressed variant, likely playing a fundamental role across tissues while exhibiting functional differences from isoform 1. These results emphasize the critical importance of ΔN-RNF220 in future studies investigating the biological functions of RNF220.