CLASP localizes in two discrete patterns on cortical microtubules and is required for cell morphogenesis and cell division in Arabidopsis

Abstract

In animals and yeast, CLASP proteins are microtubule plus-end tracking proteins (+TIPS) involved in the regulation of microtubule plus-end dynamics and stabilization. Here we show that mutations in the Arabidopsis CLASP homolog result in various plant growth reductions, cell form defects and reduced mitotic activity. Analysis of Arabidopsis plants that carry a YFP:AtCLASP fusion construct regulated by the AtCLASP native promoter showed similarities to the described localization of the animal CLASP proteins, but also prominent differences including punctate and preferential localization along cortical microtubules. Colocalization studies of YFP:AtCLASP and CFP:EB1b also showed that AtCLASP is enriched at the plus ends of microtubules where it localizes behind the AtEB1b protein. Moreover, AtCLASP overexpression causes abnormal cortical microtubule bundling and array organization. Cortical microtubule arrays have evolved to be prominent in plants, and our findings suggest that plant CLASP proteins may have adopted specific functions in regulating cortical microtubule properties and cell growth.

The IQD Family of Calmodulin-Binding Proteins Links Calcium Signaling to Microtubules, Membrane Subdomains, and the Nucleus

“…Interestingly, we noticed that most GFP-IQD proteins uniformly decorated MT arrays, similar to members of the MAP65 family (Lucas et al, 2011), which function mainly in MT bundling . In contrast, most other MAPs either accumulate at the plus end of MTs, such as CLIP-ASSOCIATED PROTEIN (CLASP) (Kirik et al, 2007), END BINDING PROTEIN EB1 (Bisgrove et al, 2008), or SPIRAL1 (SPR1) (Sedbrook et al, 2004), where they regulate the dynamics of MT growth and shrinkage, or associate in punctate to patchy patterns preferably at newly formed MT crossover sites, as described for KATANIN 1(KTN1) (Lindeboom et al, 2013) and SPR2 (Shoji et al, 2004). Evenly distributed MT association suggests that IQD proteins may influence MT bundling.…”

POM-POM2/CELLULOSE SYNTHASE INTERACTING1 Is Essential for the Functional Association of Cellulose Synthase and Microtubules inArabidopsis

“…In addition, twisting was also observed in soil-grown rosette leaves (left-handed; Figures 1A and 1B). Twisting or spiraling of organs occurs in mutants affecting cytoskeleton organization, for example, in the microtubuleassociated clasp, lefty1 and 2, and sku6/spr1 (Thitamadee et al, 2002;Sedbrook et al, 2004;Ambrose et al, 2007;Kirik et al, 2007). To investigate whether the microtubule organization was disturbed in pom2, we crossed pom2-4 to the microtubule marker lines Microtubule-Associated Protein4 (MAP4):green fluorescent protein (GFP) or mCherry:Tubulin alpha-5 (TUA5) (Marc et al, 1998;Gutierrez et al, 2009).…”

“…Apart from altered microtubule organization, mutations in several microtubule-associated genes also lead to reduced mitotic activity (Ambrose et al, 2007;Kirik et al, 2007;Sunohara et al, 2009). To test whether the pom2 mutants also hold fewer cells that actively divide, we crossed lines expressing the mitotic marker cycB1;1:Cyclin Destruction Box (CDB):b-glucuronidase (GUS) (Hauser and Bauer, 2000) with several pom2 mutant alleles.…”

The Microtubule Plus-End Tracking Proteins SPR1 and EB1b Interact to Maintain Polar Cell Elongation and Directional Organ Growth in Arabidopsis

“…However, despite showing a clear comet-like distribution at MT growing ends, the SPR1:EGFP signal was too weak and sparse to permit fast FRAP analysis of small regions just behind the MT tips, as was performed previously for EB1 (Dragestein et al, 2008). To overcome this problem, we took advantage of the fact that, relative to 35S-expressed mCherry: EB1b (Supplemental Figure 5; Chan et al, 2003;Mathur et al, 2003;Kirik et al, 2007), SPR1:EGFP is localized in long MTassociated tails in Arabidopsis cells (Sedbrook et al, 2004). These longer tails enabled us to photobleach larger regions and to determine if these longer tails recovered rapidly, consistent with rapid lateral exchange of SPR1 with the MT lattice, or if these tails were relatively slow to recover or did not recover at all.…”