Abstract Detail

Cell biology

Eng, Ryan [1], Wasteneys, Geoffrey [2].

Dissecting the interaction between NEK6 and ARK1 in regulating microtubule dynamics and cell elongation in Arabidopsis thaliana.

Microtubules are dynamic polymers that are precisely regulated by associated proteins or post-translational modifications in order for normal plant cell growth to occur. In Arabidopsis thaliana, the ARMADILLO-REPEAT KINESIN 1 (ARK1) is known to promote the depolymerization of microtubules while the NIMA-related Kinase 6 (NEK6) has been shown to phosphorylate the β-tubulin subunit of microtubules, possibly as a way to increase microtubule dynamics. NEK6 is thought to suppress ectopic outgrowth of epidermal cells while promoting proper cell elongation. Previous data has shown a physical interaction between NEK6 and the Armadillo-domain (ARM) of ARK1, although the exact function and importance of this interaction is unknown. To elucidate this interaction, ark1 nek6 double mutants were created to observe potential changes in the number of ectopic protrusions in hypocotyl and petiole epidermal cells that are phenotypically associated with nek6 mutants. The number of ectopic protrusions in ark1 nek6 double mutants was not significantly different from nek6 mutants, suggesting that NEK6 does not require ARK1 for proper regulation of microtubule dynamics and cell elongation. In order to test if both proteins are mutually dependent on each other for localization and function, NEK6-GFP (green fluorescent protein) and ARK1-GFP fusion-protein constructs were transformed into the ark1 and nek6 mutant backgrounds, respectively, to observe any potential changes in protein localization; however, much like in the wild-type background, both proteins’ localization to microtubules remained unchanged relative to the wild-type background. Previous studies determined that the ectopic protrusions of nek6 were suppressed or enhanced when exposed to the plant hormones, ACC (a precursor to ethylene) and gibberellic acid (GA), respectively. To test for potential hormone-specific ARK1-NEK6 interactions at the cellular level, plants expressing ARK1-tagRFP and NEK6-GFP were exposed to exogenous ACC and GA but no changes in localization were noted relative to mock treatments. Over-expression of an ARM-RFP construct in NEK6-GFP transgenic lines led to complete inhibition of NEK6-GFP transcription. This observation was confirmed through confocal microscopy in which plants over-expressing ARM-RFP lacked GFP signal. The data suggests that ARK1 and NEK6 function independently from each other to regulate microtubule dynamics. However, ARK1 may play an important regulatory role in NEK6 expression.

1 - University of British Columbia, Botany, 6270 University Blvd, Vancouver, BC, V6T 1Z4, Canada
2 - University of British Columbia, 6270 University blvd, Vancouver, BC, V6T 1Z4, Canada

Keywords:
Kinesin
Development
Cell Biology
Kinase
Phosphorylation
microtubule
Ethylene
Giberellic Acid.

Presentation Type: Oral Paper:Papers for Topics
Session: 58
Location: Salon 17/The Shaw Conference Centre
Date: Wednesday, July 29th, 2015
Time: 9:00 AM
Number: 58005
Abstract ID:1092
Candidate for Awards:CSPB President's Award for Best Student Presentation