The alternate AP-1 adaptor subunit Apm2 interacts with the Mil1 regulatory protein and confers differential cargo...

Area of Science:

• Cell Biology
• Molecular Biology
• Protein Trafficking

Background:

• Heterotetrameric adaptor protein complexes (AP-1) mediate cargo sorting in clathrin-coated vesicles.
• Distinct AP-1 forms arise from alternate μ chain expression, altering cargo specificity, but functional differences remain unclear.

Purpose of the Study:

• Investigate the distinct cargo-sorting functions of the two yeast AP-1 forms, which differ only in their μ chain (Apm1 and Apm2).
• Determine if μ subunit differences dictate localization or cargo recognition.
• Identify regulatory factors influencing AP-1 function.

Main Methods:

• Yeast genetics to study loss-of-function phenotypes for Apm1 and Apm2.
• Fluorescence microscopy to assess protein localization (Apm1, Apm2, Snc1, Chs3).
• Co-immunoprecipitation and binding assays to identify protein interactions (Apm2 and Mil1).

Main Results:

• Loss of Apm2, but not Apm1, leads to increased cell surface levels of the v-SNARE Snc1.
• Apm2 cannot substitute for Apm1 in sorting Chs3, indicating μ-independent sorting signals.
• Apm1 and Apm2 colocalize at Golgi/early endosomes, refuting compartment-specific localization.
• A novel lipase, Mil1, binds Apm2 (not Apm1) and is required for Apm2-dependent sorting.

Conclusions:

• The yeast μ chains, Apm1 and Apm2, confer distinct cargo-sorting functions rather than specifying distinct cellular compartments.
• A novel protein, Mil1, acts as a specific regulator for Apm2-dependent sorting events.
• Regulatory interactions provide a mechanism for diversifying adaptor complex functions.