Pearling drives mitochondrial DNA nucleoid distribution

The distribution of mitochondrial DNA-containing nucleoids is essential for mitochondrial function and genome inheritance; however, no known mechanisms can explain nucleoid segregation or their regular positioning. In this work, we found that mitochondria frequently undergo a reversible biophysical instability termed "pearling," transforming from a tubular into a regularly spaced beads morphology. Physiological pearling imposed a characteristic length scale and simultaneously mediated nucleoid disaggregation and established internucleoid distancing with high precision. Pearling onset was triggered by calcium influx, whereas the density of lamellar cristae invaginations modulated pearling prevalence and preserved nucleoid spacing following recovery. The dysregulation of mitochondrial calcium influx or inner membrane cristae integrity caused aberrant nucleoid clustering. Our results identify pearling as a mechanism governing nucleoid distribution and inheritance and offer insights into its regulation.