Visualization of cell microtubules in their native state

Area of Science:

• Cellular Biology
• Structural Biology
• Microscopy

Background:

• Cryo-electron microscopy (cryo-EM) of vitrified specimens has elucidated in vitro microtubule structures.
• In vivo microtubule structure remains poorly understood due to chemical fixation artifacts in traditional electron microscopy.

Purpose of the Study:

• To investigate microtubule ultrastructure within their native cellular context using cryo-electron microscopy of vitreous sections.
• To achieve molecular-level resolution of microtubules in vivo.

Main Methods:

• Cryo-electron microscopy and cryo-electron tomography of vitreous sections.
• Preparation of vitreous sections from organotypic slices of rat hippocampus and cultured Chinese-hamster ovary cells.

Main Results:

• Microtubules were visualized in vivo, revealing protofilament ultrastructure and polarity.
• Molecular details comparable to in vitro studies were obtained, with a resolution of approximately 4 nm.
• Luminal particles of approximately 6 nm in diameter were detected within microtubules.

Conclusions:

• This study provides the first molecular-level description of microtubules in an unperturbed cellular context.
• The achieved resolution is comparable to plunge-frozen samples, enabling in vivo identification of macromolecular assemblies.
• This method advances in vivo structural biology by allowing visualization of cellular components without chemical fixation.