Plant farnesyltransferase can restore yeast Ras signaling and mating

Area of Science:

• Molecular Biology
• Biochemistry
• Yeast Genetics

Background:

• Farnesyltransferase (FTase) is a crucial enzyme for protein modification in eukaryotes.
• FTase plays a vital role in cell growth and mating by farnesylating proteins like Ras and alpha-factor.
• Understanding FTase function across different species can reveal conserved biological mechanisms.

Purpose of the Study:

• To investigate the functional conservation of plant FTase subunits in a heterologous yeast system.
• To assess the ability of tomato FTase subunits to farnesylate Ras and alpha-factor proteins in yeast.
• To identify key domains within the plant FTase beta subunit essential for its function.

Main Methods:

• Cloning and expression of tomato FTase alpha (LeFTA) and beta (LeFTB) subunits in Saccharomyces cerevisiae.
• Complementation analysis of yeast ram1 delta mutant strains lacking endogenous FTase beta.
• Assessment of Ras membrane association and mating capabilities in complemented yeast strains.
• In vitro farnesylation assays using CaaX-motif-containing peptides.

Main Results:

• The tomato FTase beta subunit (LeFTB) alone could not rescue the growth defect of ram1 delta yeast.
• Co-expression of LeFTA and LeFTB restored normal yeast growth, Ras membrane association, and mating.
• A novel 66-amino-acid domain in LeFTB was found to be important for efficient complementation.
• The co-expressed plant FTase enzyme correctly farnesylated target peptides in vitro.

Conclusions:

• Plant FTase subunits are functionally conserved and can perform essential farnesylation in yeast.
• The study highlights the evolutionary conservation of FTase enzymatic activity and substrate specificity between plants and yeast.
• This research provides insights into the structural requirements of plant FTase for its biological function.