Systematic Localization of the Arabidopsis Core Cell Cycle Proteins Reveals Novel Cell Division Complexes    

Abstract

Cell division depends on the correct localization of the cyclin-dependent kinases that are regulated by phosphorylation, cyclin proteolysis, and protein-protein interactions. Although immunological assays can define cell cycle protein abundance and localization, they are not suitable for detecting the dynamic rearrangements of molecular components during cell division.Here, we applied an in vivo approach to trace the subcellular localization of 60 Arabidopsis (Arabidopsis thaliana) core cell cycle proteins fused to green fluorescent proteins during cell division in tobacco (Nicotiana tabacum) and Arabidopsis. Several cell cycle proteins showed a dynamic association with mitotic structures, such as condensed chromosomes and the preprophase band in both species, suggesting a strong conservation of targeting mechanisms. Furthermore, colocalized proteins were shown to bind in vivo, strengthening their localization-function connection. Thus, we identified unknown spatiotemporal territories where functional cell cycle protein interactions are most likely to occur.

A protein phosphatase 2A complex spatially controls plant cell division

“…Several kinases may act to counteract the TTP phosphatase activity since reversible protein phosphorylation/dephosphorylation has an important role in cortical microtubular organization 37,38 . One of these could be the cyclin-dependent kinase CDKA;1, previously shown to decorate the late PPB and to induce its disassembly [39][40][41][42] , and which co-purified with TON1 in TAP experiments 43 . Aurora kinases, which in animals are functional antagonists of PP2A at the centrosome and microtubule plus ends [44][45][46] , are also putative candidates as suggested by recent results pointing to a function for Aurora kinases in premitotic division plane orientation and cortical cytoskeleton organization 47,48 .…”

The RING E3 Ligase KEEP ON GOING Modulates JASMONATE ZIM-DOMAIN12 Stability

“…Constructs were transiently expressed by A. tumefaciens-mediated transient transformation of lower epidermal leaf cells as described previously (Boruc et al, 2010) using a modified buffer (10 mM MgCl 2 [1 M stock solution; Merck], 10 mM MES [0.5 M stock solution; Duchefa], and 100 mM acetosyringone [100 mM stock solution; SigmaAldrich]) and the addition of a P19-expressing A. tumefaciens strain to boost protein expression (Voinnet et al, 2003). All A. tumefaciens strains were grown for 2 d, diluted to an optical density of 1 in infiltration buffer, and incubated for 2 h at room temperature before mixing in a 1:1 ratio with other strains and injecting.…”

“…The PCR product, to which attB sites were added, was recombined with pDONR221 (Invitrogen) and then pFAST-R02 (Shimada et al, 2010) as the destination vector. BiFC constructs for Pro-35S:ORF-tag or Pro-35S:tag-ORF using the N-and C-terminal halves of enhanced GFP were constructed by multisite Gateway reactions using pK7m34GW, pH7m34GW (Karimi et al, 2005), or pH7m24GW as described (Boruc et al, 2010) combined with pDONR207 and pDONR221 entry clones of JAZ12, NINJA, KEG, and KEG(AA) (with or without stop codon).…”

Enhanced levels of plant cell cycle inhibitors hamper root-knot nematode-induced feeding site development

“…These assumptions are supported by the different localization of these proteins in interphase nuclei or along mitosis. 25,26 KRP1 and KRP4 proteins present both nuclear and sub-nuclear localization and co-localize with chromosomes during mitosis. 16,26 KRP2 is evenly distributed in the interphase nucleus and apparently degraded during mitosis.…”