Alternative splicing generates a truncated isoform of human TNFRSF11A (RANK) with an altered capacity to activate NF-κB.

Alternative splicing (AS) is a major post-transcriptional modification taking place in all cells. Many members of the TNF receptor superfamily modulate their function through protein isoforms produced by alternative splicing. TNFRSF11A (RANK) gene, through alternative splicing produces multiple isoforms truncated in their intracellular domain, with distinct functions. Here, we report the identification and characterization of a novel human TNFRSF11A (RANK) variant from human normal brain, named RANK-e5a (TNFRSF11A_e5a). The novel variant lacks 42 nucleotides from exon 5, giving rise to a novel shorter form of exon 5, named exon 5a. The incorporation of the novel exon 5a in RANK mRNA does not affect the open reading frame, producing a truncation of thirteen amino acids of the third and fourth TNFR motifs of the extracellular part of the receptor. By western blot analysis and immunofluorescence staining we were able to further characterize the RANK-e5a isoform at the protein level. In addition, we performed an ELISA assay to characterize RANK/RANKL and RANK-e5a/RANKL binding capacities, and we identified a reduced affinity of RANK-e5a to bind RANKL. Finally, when RANK-e5a is stimulated by RANK ligand, its capability to activate NF-κB is reduced compared to the wild type RANK receptor. Overall, our data provide a novel regulatory mechanism for the RANK/RANKL system, at the RANK receptor level.