Pearling drives mitochondrial DNA nucleoid distribution

Abstract The mitochondria of most eukaryotes carry an indispensable second genome (mtDNA), encoding genes engaged in oxidative phosphorylation 1 . The regular positioning and segregation of mtDNA-containing nucleoids is essential for mitochondrial function and inheritance, as well as cellular health 2–5 . However, the underlying mechanism driving nucleoid distribution and disaggregation remains unknown 6,7 . Our data reveal that mitochondria frequently undergo reversible pearling, a biophysical instability that undulates tubules into regularly spaced beads 8 , typically triggered by calcium influx. We discovered that physiological pearling imposes a characteristic length scale, simultaneously mediating nucleoid disaggregation and establishing inter-nucleoid distancing with near-maximally achievable precision. We found that lamellar cristae invaginations of the inner mitochondrial membrane play a dual role, determining pearling frequency and duration, and preserving the resulting nucleoid spacing after organelle recovery to a tubular form. Thus, disrupting cristae ultrastructure resulted in more frequent pearling, but also aberrant nucleoid clustering. Our results demonstrate that the distribution of mitochondrial genomes is governed by the interplay between rapid and reversible pearling and cristae ultrastructure, establishing a mechanism for this long-puzzling yet fundamental feature of eukaryotic life, and offering insights into its potential modulation.