Desmoplakin tail domain position in the desmosomal plaque is isoform dependent

Desmoplakin (DP) is the anchoring subunit of desmosomes, macromolecular junctions that provide mechanical integrity to the skin and heart. DP has three isoforms, DPI, DPIa, and DPII that arise from alternative splicing. The isoforms are structurally identical excluding the length of their central rod domain. As desmosomes are macromolecular complexes, the precise arrangement of their component proteins, or architecture, is essential to maintain physiological function. Alterations of the tissue-specific expression of DP isoforms underlies rare human diseases impacting the skin and heart. Overall DP is oriented with its head domain closest to the plasma membrane and tail domain extending into the cytosol. However, the differences in the architecture of the DP isoforms within the desmosomal plaque remains unknown. Here, we sought to define the architectural arrangement of each DP isoform. To address this, we utilized direct stochastic optical reconstruction microscopy (dSTORM) and analysis of DP KO HaCaT cells stably expressing DPI, DPIa, or DPII with a C-terminal mEGFP tag. Our results show the DP head domain position in the desmosomal plaque is isoform independent and the DP tail domain position correlates with rod length. The tail domain of DPI, the isoform with the longest rod, is furthest from the plasma membrane and that of DPII, the isoform with the shortest rod, is closest. We propose a variable tail location model to describe the architectural arrangement of each isoform. In this model, the DP isoforms are arranged with their rod domains parallel at an angle between 21{degrees} to 25{degrees} from the plasma membrane. These results provide valuable insight into the role of DP isoforms in desmosomal architecture and function.