Bachelor of Science, National Taiwan University (2003)
Doctor of Philosophy, University of California Davis (2011)
Dominique Bergmann, Postdoctoral Faculty Sponsor
Plant cytokinesis is brought about by the phragmoplast, which contains an antiparallel microtubule (MT) array. The MT-associated protein MAP65-3 acts as an MT-bundling factor that specifically cross-links antiparallel MTs near their plus ends. MAP65 family proteins contain an N-terminal dimerization domain and C-terminal MT interaction domain. Compared with other MAP65 isoforms, MAP65-3 contains an extended C terminus. A MT binding site was discovered in the region between amino acids 496 and 588 and found to be essential for the organization of phragmoplast MTs. The frequent cytokinetic failure caused by loss of MAP65-3 was not rescued by ectopic expression of MAP65-1 under the control of the MAP65-3 promoter, indicating nonoverlapping functions between the two isoforms. In the presence of MAP65-3, however, ectopic MAP65-1 appeared in the phragmoplast midline. We show that MAP65-1 could acquire the function of MAP65-3 when the C terminus of MAP65-3, which contains the MT binding site, was grafted to it. Our results also show that MAP65-1 and MAP65-3 may share redundant functions in MT stabilization. Such a stabilization effect was likely brought about by MT binding and bundling. We conclude that MAP65-3 contains a distinct C-terminal MT binding site with a specific role in cross-linking antiparallel MTs toward their plus ends in the phragmoplast.
View details for DOI 10.1105/tpc.111.092569
View details for Web of Science ID 000306105400029
View details for PubMedID 22570443
Plant cells assemble the bipolar spindle and phragmoplast microtubule (MT) arrays in the absence of the centrosome structure. Our recent findings in Arabidopsis thaliana indicated that AUGMIN subunit3 (AUG3), a homolog of animal dim γ-tubulin 3, plays a critical role in γ-tubulin-dependent MT nucleation and amplification during mitosis. Here, we report the isolation of the entire plant augmin complex that contains eight subunits. Among them, AUG1 to AUG6 share low sequence similarity with their animal counterparts, but AUG7 and AUG8 share homology only with proteins of plant origin. Genetic analyses indicate that the AUG1, AUG2, AUG4, and AUG5 genes are essential, as stable mutations in these genes could only be transmitted to heterozygous plants. The sterile aug7-1 homozygous mutant in which AUG7 expression is significantly reduced exhibited pleiotropic phenotypes of seriously retarded vegetative and reproductive growth. The aug7-1 mutation caused delocalization of γ-tubulin in the mitotic spindle and phragmoplast. Consequently, spindles were abnormally elongated, and their poles failed to converge, as MTs were splayed to discrete positions rendering deformed arrays. In addition, the mutant phragmoplasts often had disorganized MT bundles with uneven edges. We conclude that assembly of MT arrays during plant mitosis depends on the augmin complex, which includes two plant-specific subunits.
View details for DOI 10.1105/tpc.112.096610
View details for PubMedID 22505726
In plant cells, microtubules (MTs) in the cytokinetic apparatus phragmoplast exhibit an antiparallel array and transport Golgi-derived vesicles toward MT plus ends located at or near the division site. By transmission electron microscopy, we observed that certain antiparallel phragmoplast MTs overlapped and were bridged by electron-dense materials in Arabidopsis thaliana. Robust MT polymerization, reported by fluorescently tagged End Binding1c (EB1c), took place in the phragmoplast midline. The engagement of antiparallel MTs in the central spindle and phragmoplast was largely abolished in mutant cells lacking the MT-associated protein, MAP65-3. We found that endogenous MAP65-3 was selectively detected on the middle segments of the central spindle MTs at late anaphase. When MTs exhibited a bipolar appearance with their plus ends placed in the middle, MAP65-3 exclusively decorated the phragmoplast midline. A bacterially expressed MAP65-3 protein was able to establish the interdigitation of MTs in vitro. MAP65-3 interacted with antiparallel microtubules before motor Kinesin-12 did during the establishment of the phragmoplast MT array. Thus, MAP65-3 selectively cross-linked interdigitating MTs (IMTs) to allow antiparallel MTs to be closely engaged in the phragmoplast. Although the presence of IMTs was not essential for vesicle trafficking, they were required for the phragmoplast-specific motors Kinesin-12 and Phragmoplast-Associated Kinesin-Related Protein2 to interact with MT plus ends. In conclusion, we suggest that the phragmoplast contains IMTs and highly dynamic noninterdigitating MTs, which work in concert to bring about cytokinesis in plant cells.
View details for DOI 10.1105/tpc.110.078204
View details for Web of Science ID 000295254700012
View details for PubMedID 21873565
In higher plant cells, microtubules (MTs) are nucleated and organized in a centrosome-independent manner. It is unclear whether augmin-dependent mechanisms underlie spindle MT organization in plant cells as they do in animal cells. When AUGMIN subunit3 (AUG3), which encodes a homolog of animal dim γ-tubulin 3/human augmin-like complex, subunit 3, was disrupted in Arabidopsis thaliana, gametogenesis frequently failed due to defects in cell division. Compared with the control microspores, which formed bipolar spindles at the cell periphery, the mutant cells often formed peripheral half spindles that only attached to condensed chromosomes or formed elongated spindles with unfocused interior poles. In addition, defective cells exhibited disorganized phragmoplast MT arrays, which caused aborted cytokinesis. The resulting pollen grains were either shrunken or contained two nuclei in an undivided cytoplasm. AUG3 was localized along MTs in the spindle and phragmoplast, and its signal was pronounced in anaphase spindle poles. An AUG3-green fluorescent protein fusion exhibited a dynamic distribution pattern, similar to that of the γ-tubulin complex protein2. When AUG3 was enriched from seedlings by affinity chromatography, AUG1 was detected by immunoblotting, suggesting an augmin-like complex was present in vivo. We conclude that augmin plays a critical role in MT organization during plant cell division.
View details for DOI 10.1105/tpc.111.086892
View details for Web of Science ID 000294164300016
View details for PubMedID 21750235