Association between elongation factor-1alpha and microtubules in vivo is domain dependent and conditional.
Although the precise definition for a microtubule-associated protein (MAP) has been the subject of debate, elongation factor-1alpha (EF-1alpha) fits the most basic criteria for a MAP [Durso and Cyr, 1994a]. It binds, bundles, stabilizes, and promotes the assembly of microtubules in vitro, and localizes to plant microtubule arrays in situ. In this study, the in vitro and in vivo association of EF-1alpha with microtubules was further investigated. Analysis of the in vitro binding data for EF-1alpha and microtubules indicates that EF-1alpha binds cooperatively to the microtubule lattice. In order to investigate the interaction of EF-1alpha with microtubules in vivo, GFP fusions to EF-1alpha or to EF-1alpha truncates were transiently expressed in living plant cells. Using this method, two putative microtubule-binding domains on EF-1alpha were identified: one in the N-terminal domain I and one in the C-terminal domain III. The binding of domain I to microtubules in vivo, like the binding of full-length EF-1alpha, is conditional, and requires incubation in weak, lipophilic organic acids. The binding of domain III to microtubules in vivo, however, is not conditional, and occurs under normal cellular regimes. Furthermore, domain III stabilizes cortical microtubules as determined by their resistance to the anti-microtubule herbicide, oryzalin. Because the accumulation of EF-1alpha onto microtubules is unconditional in the absence of domain I, we hypothesize that domain I negatively regulates the accumulation of EF-1alpha onto microtubules in vivo. This hypothesis is discussed in terms of possible regulatory mechanisms that could affect the accumulation of EF-1alpha onto microtubules within living cells.
Moore
R C
RC
Department of Biology and Intercollege Program in Plant Physiology, The Pennsylvania State University, University Park 16802, USA.
Cyr
R J
RJ
D003599:Cytoskeleton / Q000378:metabolism; D018552:Daucus carota / Q000378:metabolism; D007887:Fabaceae / Q000378:metabolism; D049452:Green Fluorescent Proteins; D007700:Kinetics; D008164:Luminescent Proteins; D018613:Microscopy, Confocal; D008870:Microtubules / Q000378:metabolism*; D017239:Paclitaxel / Q000494:pharmacology; D020648:Peptide Elongation Factor 1 / Q000235:genetics / Q000378:metabolism*; D010940:Plant Proteins / Q000378:metabolism*; D010946:Plants, Medicinal; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000378:metabolism
D008164:Luminescent Proteins; D020648:Peptide Elongation Factor 1; D010940:Plant Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D017239:Paclitaxel