A novel correlation-based method for quantifying membrane-associated periodic skeleton periodicity

The advent of superresolution microscopy uncovered the presence of the membrane-associated periodic skeleton (MPS), a part of the neuronal cytoskeleton that is organized in a periodic localization pattern. Central to this structure are spectrin tetramers, which localize rings of actin filaments at an average distance of 190 nm. The MPS is particularly pronounced in the axon, where it appears able to recruit ion channels and signaling platforms into similar periodic localization patterns, suggesting the MPS plays a key role in orchestrating neuronal signaling. Despite this, precise quantification of MPS-associated periodicity has been lacking a unified quantification method. We here developed a novel, unbiased, user-friendly approach that enabled quantitative measures of MPS periodicity using autocorrelation. In addition, it was able to estimate co-distributions of periodic targets via cross-correlation. We employed the tool to analyze the nanoscale organization of the potassium channel TRAAK within the axon initial segment (AIS). Our analysis revealed that exogenously expressed TRAAK co-distributes with ankyrin G in a periodic localization pattern and that this localization depends on a C-terminal ankyrin G binding motif within the channel. Thus, our analytical tool can provide quantitative measures of periodic localization patterns to obtain a deeper understanding of the MPS and its implications in neuronal signaling and pathology.