Subtomogram averaging workflow followed using the Dynamo package.

(a) Schematic workflow of the subtomogram averaging procedure to generate the ribosome volume. (b) Two 60°-related views of ribosome reconstruction (transparent white), fitted with the structure of the V. necatrix ribosome (PDB ID: 6RM3, magenta). (c) Schematic workflow of the subtomogram averaging procedure used to create the reconstructions of the segments of polar tube outer layers. The scheme is shown for cargo-filled tubes, and a similar methodology was used for empty tubes. (d) Subtomogram averaging workflow used to reconstruct dimeric ribosomes from clustered particles in sporoplasm-filled tubes. (e) Low-pass filtered subtomogram averages of ribosome dimers placed back into their original location in germinated polar tubes. Averages are shown in yellow, and the central slice of the tomogram slice is shown in gray.

(PNG)