A combined 3D-SIM/SMLM approach allows centriole proteins to be localized with a precision of ∼4–5 nm

Centrioles are small barrel-shaped structures that form centrosomes and cilia [1]. Centrioles assemble around a central cartwheel comprising the Sas-6 and Ana2/STIL proteins. The amino termini of nine Sas-6 dimers form a central hub of ∼12 nm radius from which nine dimer spokes radiate, placing the Sas-6 carboxyl termini at the outer edge of the ∼60 nm radius cartwheel [2]. Several centriole proteins are distributed in a toroid around the cartwheel, and super-resolution light microscopy studies have measured the average radii of these ∼100–200 nm radius toroids with a ‘precision’ — or standard deviation (s.d. or 1σ) — of ±∼10–40 nm. The organization of Ana2/STIL within the cartwheel, however, has not been resolvable. Here, we develop methods to calculate the average toroidal radius of centriolar proteins in the ∼20–60 nm range with a s.d. of just ±∼4–5 nm, revealing that the amino and carboxyl termini of Ana2 are located in the outer cartwheel region. Centrioles are small structures that assemble around a central cartwheel comprising Sas-6, Ana2/STIL and Sas-4. The cartwheel cannot be resolved by conventional light microscopy. Here, Gartenmann et al. resolve the relative positions of the Sas-6, Ana2 and Sas-4 termini within the cartwheel using combined SIM/SMLM super-resolution microscopy.