Biology subjects

Reetz, E.

Publications and source records attributed to Reetz, E..

5 recordsLinked to original sources

bioRxiv2022

Cryo-electron tomography of the apicomplexan invasion machinery in its native state reveals rigid body motion of the conoid and docked secretory machinery

The apical complex is a specialized collection of cytoskeletal and secretory machinery in apicomplexan parasites, which include the pathogens that cause malaria and toxoplasmosis. Its structure and mechanism of motion are poorly understood. We used cryo-FIB-milling and cryo-electron tomography to visualize the 3D-structure of the apical complex in its protruded and retracted states. Averages of conoid-fibers revealed their polarity and unusual nine-protofilament arrangement with associated proteins connecting and likely stabilizing the fibers. Neither the structure of the conoidfibers nor the architecture of the spiral-shaped conoid complex change during protrusion or retraction. Thus, the conoid moves as a rigid body, and is not spring-like and compressible, as previously suggested. Instead, the apical-polar-rings (APR), previously considered rigid, dilate during conoid protrusion. We identified actin-like filaments connecting the conoid and APR during protrusion, suggesting a role during conoid movements. Furthermore, our data capture the parasites in the act of secretion during conoid protrusion.

biophysics↗

bioRxiv2022

Triglyceride lipolysis driven by glucose restriction triggers liquid-crystalline phase transitions and proteome remodeling of lipid droplets

Lipid droplets (LDs) are reservoirs for triglycerides (TGs) and sterol-esters (SEs), but how these lipids are organized within LDs and influence its proteome remains unclear. Using in situ cryoelectron tomography, we show that glucose restriction triggers lipid phase transitions within LDs generating liquid-crystalline lattices inside them. Mechanistically this requires TG lipolysis, which decreases the LD TG:SE ratio, promoting SE transition to a liquid-crystalline phase. Molecular dynamics simulations reveal TG depletion promotes spontaneous TG and SE de-mixing in LDs, additionally altering the lipid packing of the phospholipid monolayer surface. Fluorescence imaging and proteomics further reveal that liquid-crystalline phases are associated with selective remodeling of the LD proteome. Some canonical LD proteins including Erg6 re-localize to the ER network, whereas others remain LD-associated. Model peptide LiveDrop also redistributes from LDs to the ER, suggesting liquid-crystalline-phases influence ER-LD inter organelle transport. Our data suggests glucose restriction drives TG mobilization, which alters the phase properties of LD lipids and selectively remodels the LD proteome.

cell biology↗

bioRxiv2022

Three-dimensional cilia structures from animals' closest unicellular relatives, the Choanoflagellates

In most eukaryotic organisms, cilia 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 ciliary 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 opisthokont clade, leaving a sizeable gap in our understanding of cilia 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 cilia 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 cilia, including microtubule holes, microtubule inner proteins, and the ciliary 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 ciliary membrane. Together, our findings provide new insights into choanoflagellate biology and cilia evolution between unicellular and multicellular opisthokonts.

cell biology↗

bioRxiv2021

Liquid-crystalline lipid phase transitions in lipid droplets selectively remodel the LD proteome

Lipid droplets (LDs) are reservoirs for triglycerides (TGs) and sterol-esters (SEs). How lipids are organized within LDs and influence the LD proteome remains unclear. Using in situ cryo-electron tomography, we show that glucose restriction triggers lipid phase transitions within LDs generating liquid-crystalline lattices inside them. Mechanistically, this requires TG lipolysis, which alters LD neutral lipid composition and promotes SE transition to a liquid-crystalline phase. Fluorescence imaging and proteomics further reveal that LD liquid-crystalline lattices selectively remodel the LD proteome. Some canonical LD proteins including Erg6 re-localize to the ER network, whereas others remain on LDs. Model peptide LiveDrop also redistributes from LDs to the ER, suggesting liquid-crystalline-phases influence ER-LD inter-organelle transport. Proteomics also indicates glucose restriction elevates peroxisome lipid oxidation, suggesting TG mobilization provides fatty acids for cellular energetics. This suggests glucose restriction drives TG mobilization, which alters the phase properties of LD lipids and selectively remodels the LD proteome.

cell biology↗

bioRxiv2021

3D structure and in situ arrangements of CatSper channel in the sperm flagellum

The sperm calcium channel CatSper plays a central role in successful fertilization as a primary Ca2+ gateway into the sperm flagellum. However, CatSpers complex subunit composition has impeded its reconstitution in vitro and structural elucidation. Here, we applied cryo-electron tomography to visualize the macromolecular organization of the native CatSper channel complex in intact mammalian sperm, as well as identified three additional CatSper-associated proteins. The repeating CatSper units form long zigzag-rows in four nanodomains along the flagella. In both mouse and human sperm, each CatSper repeat consists of a tetrameric pore complex. Murine CatSper contains an additional outwardly directed wing-structure connected to the tetrameric channel. The majority of the extracellular domains form a canopy above each pore-forming channel that interconnects to a zigzag-shaped roof. The intracellular domains link two neighboring channel complexes to a diagonal array. The loss of this intracellular link in Efcab9-/- sperm distorts the longitudinally aligned zigzag pattern and compromises flagellar movement. This work offers unique insights into the mechanisms underlying the assembly and transport of the CatSper complex to generate the nanodomains and provides a long-sought structural basis for understanding CatSper function in the regulation of sperm motility.

biophysics↗