Separation-of-function MCPH-associated mutations in CPAP affect centriole number and length

Area of Science:

• Cell Biology
• Molecular Biology
• Genetics

Background:

• Centrioles are essential microtubule-based structures regulating cell division.
• CPAP (CENPJ) is a key protein controlling centriole length via microtubule interactions.
• Mutations in CPAP's G-box domain are linked to primary microcephaly (MCPH).

Purpose of the Study:

• To investigate the distinct functional impacts of two MCPH-associated CPAP G-box variants (E1235V and D1196N).
• To elucidate the molecular mechanisms by which these CPAP mutations affect centriole biology and cell viability.
• To explore alternative pathways for CPAP recruitment to centrioles.

Main Methods:

• Analysis of CPAP variants E1235V and D1196N in MCPH.
• Assessment of CPAP centriole recruitment, length, and number.
• STIL binding assays and molecular dynamics simulations.
• Investigation of CEP152-dependent CPAP localization.

Main Results:

• E1235V reduces CPAP recruitment, leading to elongated centrioles.
• D1196N increases centriole numbers without altering localization.
• Both mutations abolish STIL binding, crucial for centriole duplication.
• An alternative CEP152-dependent route for CPAP recruitment was identified.

Conclusions:

• The CPAP G-box region outside microtubule-binding domains critically regulates centriole number and length.
• MCPH-associated CPAP mutations disrupt STIL interaction, causing spindle defects and reduced cell viability.
• Findings provide molecular insights into primary microcephaly pathogenesis and highlight CPAP's complex regulatory roles.