Lte1 contributes to Bfa1 localization rather than stimulating nucleotide exchange by Tem1

Area of Science:

• Cell Biology
• Molecular Biology
• Genetics

Background:

• Lte1 is a known mitotic regulator in Saccharomyces cerevisiae.
• Lte1 was previously thought to function as a guanosine nucleotide exchange factor (GEF) for Tem1, a key component of the mitotic exit network.

Purpose of the Study:

• To reevaluate the proposed GEF function of Lte1.
• To elucidate the precise mechanism by which Lte1 regulates the mitotic exit network and cell polarization.

Main Methods:

• In vitro GEF activity assays.
• Mutational analysis of Lte1's putative GEF domain.
• Microscopy to assess subcellular localization of proteins (Lte1, Bfa1, polarity cap) and spindle position.
• Analysis of lte1 mutants and cells with misaligned spindles.

Main Results:

• No detectable GEF activity of Lte1 for Tem1 in vitro.
• Lte1's function depends on interaction with Ras for localization, not nucleotide exchange.
• Lte1 determines the subcellular localization of Bfa1 at spindle pole bodies (SPBs), promoting Bfa1 loss from the maternal SPB.
• Mislocalization of Lte1 in mother cells with misaligned spindles leads to Bfa1 loss and bypass of the spindle position checkpoint.
• lte1 mutants exhibit aberrant localization of the polarity cap, an organizer of the actin cytoskeleton.

Conclusions:

• Lte1 does not function as a GEF for Tem1; its role is related to localization and interaction with Ras.
• Lte1 regulates mitotic exit by controlling Bfa1 localization at SPBs and is crucial for the spindle position checkpoint.
• Lte1's function in cell polarization, evidenced by polarity cap localization, is integral to its role in mitotic regulation.