Microcompartmentation of cytosolic aldolase by interaction with the actin cytoskeleton in Arabidopsis

Garagounis, Constantine; Kostaki, Kalliopi-Ioanna; Hawkins, Tim J.; Cummins, Ian; Fricker, Mark D.; Hussey, Patrick J.; Hetherington, Alistair M.; Sweetlove, Lee J.

Abstract

Evidence is accumulating for molecular microcompartments formed when proteins interact in localized domains with the cytoskeleton, organelle surfaces, and intracellular membranes. To understand the potential functional significance of protein microcompartmentation in plants, we studied the interaction of the glycolytic enzyme fructose bisphosphate aldolase with actin in Arabidopsis thaliana. Homology modelling of a major cytosolic isozyme of aldolase, FBA8, suggested that the tetrameric holoenzyme has two actin binding sites and could therefore act as an actin-bundling protein, as was reported for animal aldolases. This was confirmed by in vitro measurements of an increase in viscosity of F-actin polymerized in the presence of recombinant FBA8. Simultaneously, interaction with F-actin caused non-competitive inhibition of aldolase activity. We did not detect co-localization of an FBA8–RFP fusion protein, expressed in an fba8-knockout background, with the actin cytoskeleton using confocal laser-scanning microscopy. However, we did find evidence for a low level of interaction using FRET-FLIM analysis of FBA8–RFP co-expressed with the actin-binding protein GFP–Lifeact. Furthermore, knockout of FBA8 caused minor alterations of guard cell actin cytoskeleton morphology and resulted in a reduced rate of stomatal closure in response to decreased humidity. We conclude that cytosolic aldolase can be microcompartmented in vivo by interaction with the actin cytoskeleton and may subtly modulate guard cell behaviour as a result.

Citation

Garagounis, C., Kostaki, K., Hawkins, T. J., Cummins, I., Fricker, M. D., Hussey, P. J., …Sweetlove, L. J. (2017). Microcompartmentation of cytosolic aldolase by interaction with the actin cytoskeleton in Arabidopsis. Journal of Experimental Botany, 68(5), 885-898. https://doi.org/10.1093/jxb/erx015

• Journal Article Type: Article
• Acceptance Date: Jan 7, 2017
• Online Publication Date: Feb 17, 2017
• Publication Date: Feb 17, 2017
• Deposit Date: Feb 16, 2017
• Publicly Available Date: Feb 17, 2018
• Journal: Journal of Experimental Botany
• Print ISSN: 0022-0957
• Electronic ISSN: 1460-2431
• Publisher: Oxford University Press
• Peer Reviewed: Peer Reviewed
• Volume: 68
• Issue: 5
• Pages: 885-898
• DOI: https://doi.org/10.1093/jxb/erx015
• Public URL: https://durham-repository.worktribe.com/output/1394324