Unique functional characteristics of the polymerization and MAP binding regulatory domains of plant tubulin.

Abstract

An understanding of the regulation of microtubule polymerization and dynamics in plant cells requires biochemical information on the structures, functions, and molecular interactions of plant tubulin and microtubule-associated proteins (MAPs) that regulate microtubule function. We have probed the regulatory domain and polymerization domain of purified maize tubulin using MAP2, an extensively characterized mammalian neumnal MAP. MAP2 bound to the surface of preformed, taxol-stabilized maize microtubules, with binding saturation occurring with one MAP2 molecule per five to six tubulin dimers, as it does with mammalian microtubules. MAP2 binding and dissociation analyses revealed two affinity classes of binding sites on maize microtubules: a high-affinity site 12 dimers apart that may be homologous to the cognate mammalian MAP2 binding site and an additional low-affinity site also 12 dimers apart that may be homologous to the mammalian tau binding site. MAPS corrected in vitro folding errors in taxol-stabilized maize microtubules and reduced the critical concentration of maize tubulin polymerization eightfold, from 8.3 to 1.0 pM. However, MAPS dissociated much more readily from maize microtubules than from mammalian microtubules and induced the assembly of maize tubulin into aberrant helical ribbon polymers that remained stable for prolonged periods. Our results indicated that MAPS binds to maize tubulin via a partially specific, low-fidelity interaction that reflects unique structural and functional properties of the polymerization and regulatory domains of plant tubulin and possibly of the tubulin binding domains of undocumented MAPs that regulate microtubule function in plant cells.

Disruption of Microtubule Organization and Centrosome Function by Expression ofTobacco Mosaic VirusMovement Protein

“…This observation is in agreement with our previous observation that MPassociated microtubules isolated from TMV-infected plant protoplasts are resistant to disruption by cold and millimolar amounts of salt (10). Obviously, MP forms specialized complexes with microtubules that differ from the salt-sensitive complexes formed by other MAPs (36,81). Exceptional MAPs that are similar to MP wt with respect to providing outstanding stability to microtubules have been described (8).…”

Identification and Characterization of a Novel Microtubule-Based Motor Associated with Membranous Organelles in Tobacco Pollen Tubes

“…Bovine brain kinesin was purified by cosedimentation with microtubules, followed by anion exchange chromatography on a Mono-Q column and gel filtration chromatography on a Sephacryl S-300 column (Amersham Pharmacia) (Kuznetsov et al, 1988). MAP2 was isolated from bovine brain by sedimentation with microtubules and heat denaturation (Hugdahl et al, 1993). Recombinant kinesin was obtained from Cytoskeleton, Inc. (Denver, CO).…”

α-Tubulin Mutations Alter Oryzalin Affinity and Microtubule Assembly Properties To Confer Dinitroaniline Resistance

“…This is within the range of values that have been observed for tubulins isolated from other eukaryotes. Although tubulins isolated from sea urchin (Strongylocentrotus purpuratus), budding yeast, and the protozoan parasite Leishmania amazonensis have C c values in the range of 0.1 to 0.2 mg/ml, tubulin from bovine brain has a C c of 0.56 mg/ml and tubulin from corn (Zea mays) has a C c of 0.83 mg/ml (8,10,27,34,35,71). It is important to note that buffer ionic strength and additives such as Taxol or DMSO influence relative C c values, so that a close comparison of values from different studies is not informative.…”