Three-dimensional flagella structures from animals’ closest unicellular relatives, the Choanoflagellates

Abstract

In most eukaryotic organisms, cilia and flagella perform a variety of life-sustaining roles related to environmental sensing and motility. Cryo-electron microscopy has provided considerable insight into the morphology and function of flagellar structures, but studies have been limited to less than a dozen of the millions of known eukaryotic species. Ultrastructural information is particularly lacking for unicellular organisms in the Opisthokonta clade, leaving a sizeable gap in our understanding of flagella evolution between unicellular species and multicellular metazoans (animals). Choanoflagellates are important aquatic heterotrophs, uniquely positioned within the opisthokonts as the metazoans’ closest living unicellular relatives. We performed cryo-focused ion beam milling and cryo-electron tomography on flagella from the choanoflagellate species Salpingoeca rosetta. We show that the axonemal dyneins, radial spokes, and central pair complex in S. rosetta more closely resemble metazoan structures than those of unicellular organisms from other suprakingdoms. In addition, we describe unique features of S. rosetta flagella, including microtubule holes, microtubule inner proteins, and the flagellar vane: a fine, net-like extension that has been notoriously difficult to visualize using other methods. Furthermore, we report barb-like structures of unknown function on the extracellular surface of the flagellar membrane. Together, our findings provide new insights into choanoflagellate biology and flagella evolution between unicellular and multicellular opisthokonts.

Macroevolutionary dynamics of gene family gain and loss along multicellular eukaryotic lineages

“…5c, d ). These patterns together suggest that the flagellar/ciliary apparatus of choanozoans, compared to other eukaryotes, is a derived feature underpinned by the recruitment of novel protein families 52 .

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An RFX transcription factor regulates ciliogenesis in the closest living relatives of animals

“…Choanoflagellate cells feature a distinctive “collar complex” composed of a single apical cilium surrounded by a collar of actin-filled microvilli 12 , 13 . Structural conservation of cilia across eukaryotic diversity suggests that the last common ancestor of eukaryotes had a cilium 1 , 14 and that cilia of choanoflagellates and animals are homologous 15 .…”

Correlative montage parallel array cryo-tomography for in situ structural cell biology

“…Cryo-electron tomography (cryo-ET) links three-dimensional (3D) contextual visualization and high-resolution structure determination of cryogenically preserved macromolecules in their native cellular environment 2 . Computationally extracted sub-tomograms can be averaged and classified to reveal sub-nanometer to nanometer resolution (3 Å to 4 nm) structures of in situ complexes 3 – 5 . Cryo-ET is generally restricted to investigations of small specimen volumes and the thin peripheral areas of cells (<500 nm) that are penetrable by the electron beam.…”