Variable Effects of C-Terminal Fusions on FLS2 Function: Not All Epitope Tags Are Created Equal

Area of Science:

• Plant Molecular Biology
• Plant Signaling
• Biochemistry

Background:

• Receptor-like kinases (RLKs) are crucial plant proteins sensing external stimuli for growth and defense.
• Limited antibody availability necessitates using epitope tags for RLK research, often assumed to be non-disruptive.

Purpose of the Study:

• To investigate the functional impact of C-terminal epitope tags on the plant receptor FLAGELLIN SENSITIVE2 (FLS2).
• To assess the reliability of epitope-tagged RLKs in biochemical and functional analyses.

Main Methods:

• Complementation analysis of Arabidopsis *fls2* mutant plants expressing various FLS2 C-terminal epitope fusions.
• Evaluation of FLS2-mediated signaling outputs and growth inhibition in response to flg22 treatment.

Main Results:

• Fls2 epitope fusions exhibited highly variable and unpredictable signaling outputs.
• Only one of four FLS2 epitope fusions restored wild-type-like growth inhibition upon flg22 treatment.

Conclusions:

• C-terminal epitope tagging of RLKs, including FLS2, can significantly and unpredictably affect protein function and downstream signaling.
• The widespread use of RLK epitope tag fusions may lead to misinterpretation of functional studies, potentially overlooking critical biological insights.