Ran GTPase regulates Mad2 localization to the nuclear pore complex.

Area of Science:

• Cell biology
• Molecular biology
• Biochemistry

Background:

• Spindle assembly checkpoint (SAC) proteins Mad1p and Mad2p localize to the nuclear pore complex (NPC) in interphase yeast and mammalian cells.
• The role of nucleocytoplasmic transport, specifically the Ran GTPase pathway, in regulating SAC protein localization and function is not fully understood.

Purpose of the Study:

• To investigate the relationship between the Ran GTPase pathway and the NPC localization of spindle assembly checkpoint proteins Mad1p and Mad2p in yeast.
• To determine if Ran levels influence Mad2p localization at the NPC and its subsequent effect on spindle checkpoint activation.

Main Methods:

• Utilized yeast cells with conditional mutations in the Ran GTPase pathway.
• Assessed the steady-state distribution of NPC-localized proteins and reporter molecules.
• Correlated Mad2p displacement from the NPC with spindle checkpoint activation during M-phase.

Main Results:

• Deletion of MAD1 or MAD2 did not alter the distribution of nuclear localization signal or nuclear export signal reporters, nor Ran localization.
• Mutations disrupting nuclear Ran concentration displaced Mad2p, but not Mad1p, from the NPC.
• Mad2p displacement from NPCs in M-phase cells correlated with spindle checkpoint activation.

Conclusions:

• Mad2p localization at the NPC is sensitive to nuclear Ran levels.
• Release of Mad2p from NPCs is closely linked to spindle assembly checkpoint activation in yeast.
• This study provides the first evidence that Ran affects Mad2p localization to the NPC.