Microtubule nucleation and γTuRC centrosome localization in interphase cells require ch-TOG

Area of Science:

• Cell Biology
• Molecular Biology
• Cytoskeleton Dynamics

Background:

• Microtubule organization is crucial for cellular functions.
• The γ-tubulin ring complex (γTuRC) nucleates microtubules at microtubule organizing centers (MTOCs).
• Adapter proteins are hypothesized to recruit and activate γTuRC, but mechanisms are unclear.

Purpose of the Study:

• To elucidate the molecular mechanisms regulating γTuRC recruitment and activation at interphase centrosomes.
• To investigate the role of ch-TOG (CKAP5) in γTuRC localization and microtubule nucleation.
• To determine if ch-TOG's function is restricted to centrosomes.

Main Methods:

• Immunofluorescence microscopy to visualize protein localization.
• Analysis of ch-TOG mutants to assess its role in γTuRC recruitment.
• Biochemical assays to study ch-TOG and γTuRC interactions.

Main Results:

• ch-TOG, not adapter proteins, controls γTuRC recruitment and activation at interphase centrosomes.
• ch-TOG co-assembles with γTuRC to stimulate nucleation around centrioles.
• Absence of ch-TOG prevents γTuRC localization to centrosomal sites, but not within the centriole lumen.
• ch-TOG also promotes microtubule nucleation at the Golgi apparatus.
• ch-TOG's dynamic association with the pericentriolar material (PCM) is regulated by its TOG domains and C-terminal region.

Conclusions:

• Microtubule nucleation and γTuRC attachment at interphase centrosomes are coupled via transient ch-TOG recruitment.
• ch-TOG acts as a key regulator of γTuRC function and microtubule nucleation.
• ch-TOG's nucleation-promoting activity extends beyond centrosomes, including the Golgi apparatus.