The Plastidial Protein Acetyltransferase GNAT1 Forms a Complex With GNAT2, yet Their Interaction Is Dispensable for...

Area of Science:

• Plant molecular biology
• Chloroplast biology
• Post-translational modifications

Background:

• Protein N-acetylation is a key co- and post-translational modification in eukaryotes, found in plant chloroplasts.
• A novel family of eight plastidial acetyltransferases with dual activity was recently identified in Arabidopsis.
• The NAA90 subgroup, including GNAT1, GNAT2, and GNAT3, are phylogenetically related acetyltransferases.

Purpose of the Study:

• To characterize the function of GNAT1, a chloroplast acetyltransferase closely related to GNAT2.
• To investigate the in vivo and in vitro N-terminus acetylation activity of GNAT1.
• To explore the interactions between GNAT1, GNAT2, and GNAT3 within the chloroplast.

Main Methods:

• Phylogenetic analysis to identify the NAA90 subgroup.
• Phenotypic analysis of gnat1 and gnat2 mutants under high light conditions.
• In vitro acetylation assays using recombinant GNAT1.
• In vivo N-terminal acetylome profiling of gnat1 knockout lines.
• Co-immunoprecipitation coupled with mass spectrometry to identify protein interactions.

Main Results:

• GNAT1 is not essential for state transitions but affects thylakoid membrane packing in gnat1 mutants.
• GNAT1 exhibits robust N-terminus acetylation activity in vitro and in vivo, targeting plastidial proteins.
• GNAT1's substrate spectrum is a subset of GNAT2's.
• GNAT1 interacts with GNAT2, and GNAT2 interacts with GNAT3, suggesting complex formation.
• Two distinct acetyltransferase complexes are identified in chloroplasts.

Conclusions:

• GNAT1 plays a role in chloroplast acetylation, distinct from GNAT2's role in state transitions.
• Complex formation of NAA90 acetyltransferases (GNAT1, GNAT2, GNAT3) is crucial for regulating acetylation activities.
• These findings reveal a novel regulatory mechanism for acetylation-dependent adjustments in plastidial metabolism.