Retrograde transport from the yeast Golgi is mediated by two ARF GAP proteins with overlapping function

Area of Science:

• Cell biology
• Molecular and cell biology
• Protein function

Background:

• ADP-ribosylation factor (ARF) proteins are crucial for vesicular transport but require GTPase-activating proteins (GAPs) for function.
• The yeast Gcs1 protein stimulates ARF GTPase activity, but its dispensable nature suggests the existence of other ARF GAPs.
• Previous studies implicated Gcs1 in vesicular transport, but its precise role and the necessity of other GAPs remained unclear.

Purpose of the Study:

• To identify and characterize additional ARF GAPs in yeast.
• To elucidate the specific roles of Glo3 and Gcs1 in vesicular transport.
• To investigate the functional relationship between Glo3 and Gcs1 in the endoplasmic reticulum (ER)-Golgi pathway.

Main Methods:

• In vitro GTPase assays to measure ARF GAP activity of Glo3.
• Genetic interaction studies using yeast mutants deficient in Glo3 and Gcs1.
• Analysis of protein transport defects in ER-Golgi vesicular transport mutants.
• In vitro assays to assess Golgi-to-ER retrieval.

Main Results:

• Glo3, like Gcs1, exhibits ARF GAP activity and is dispensable on its own.
• Glo3 and Gcs1 share an essential function in ER-Golgi vesicular transport, with their combined absence leading to cell death.
• Mutants lacking both Glo3 and Gcs1 show severe ER accumulation and impaired protein transport between the ER and Golgi.
• Genetic interactions with COPI and v-SNARE components highlight the involvement of Glo3 and Gcs1 in retrograde transport.

Conclusions:

• Glo3 and Gcs1 are essential ARF GAPs with overlapping functions in yeast vesicular transport.
• These two GAPs play a critical role in mediating retrograde transport from the Golgi to the ER.
• The combined deficiency of Glo3 and Gcs1 disrupts ER-Golgi protein trafficking and is incompatible with cell proliferation.