Microtubules may respond to the changes in tensile stress directly, but most likely multiple other proteins are required. Future research is needed to test whether microtubules, their regulators, or other factors are involved in sensing maximal tension. Actin has vital roles in cell elongation and vesicle trafficking in tip growing cells such as root hairs and pollen tubes, beautifully reviewed here Szymanski and Staiger, The importance of actin during diffuse growth and phragmoplast assembly is clear, but the underlying mechanisms are still unknown.
Actin filament alignment becomes more parallel and ordered in elongating Arabidopsis Arabidopsis thaliana root cells, but the significance is thus far unknown Dyachok et al. Actin may also mediate delivery of noncellulosic polysaccharides due to cell adhesion defects seen in actin and actin regulatory mutants and alteration in wall components by latrunculin B treatment El-Din El-Assal et al.
Actin plays different roles during different stages of phragmoplast assembly. Phragmoplast assembly was delayed when the actin-monomer stabilizing protein profilin was microinjected Valster et al. Actin motor proteins, myosins, also influence diffuse and polarized growth. There are 13 myosin XIs in Arabidopsis with often redundant functions.
Myosin double, triple, and quadruple mutants have reduced cell expansion as do cells treated with a myosin ATPase inhibitor Samaj et al. Myosin contribution to diffuse growth may be due to its role in Golgi movement within the cell, leading to well-distributed cellulose synthase deposition.
Actin regulatory proteins called formins have demonstrated roles in polarized and diffuse growth e. VidaLi et al. Rice Oryza sativa FH5 and Arabidopsis FH4 bind both microtubules and actin, indicating crosstalk between microtubules and the actin cytoskeleton Deeks et al. Lesions in rice FH5 result in compromised longitudinal actin cables but they do not affect the organization of microtubules. However, the short and swollen internode cells of the fh5 bui1 mutant indicate defects in diffuse growth Yang et al.
Several valuable approaches targeting groups of formins have also been used to clarify their functions. When all of the class 1 formins were silenced with RNAi in moss, tip growth was unaffected but plants were smaller with fewer cells, suggesting delayed cytokinesis.
No stubs or multinucleate cells were observed VidaLi et al. Despite often non-obvious roles in cytokinesis, formins sometimes localize to the phragmoplast or phragmoplast midline e. Li et al. The small molecule formin inhibitor SMIFH2, which sidesteps potentially redundant formin function, was used to understand how formins contribute to mitosis and cytokinesis Zhang et al.
SMIFH2 treatment reduces formin recruitment to the phragmoplast, slows telophase progression, and generates wavy cell plates. This powerful tool might also inhibit myosins in plants as it does in other organisms Nishimura et al. Combinations of loss-of-function formin mutants may provide conclusive evidence about their roles in growth and cytokinesis. During telophase, but before the phragmoplast reaches the cell cortex, actin filaments and cortical telophase microtubules accumulate at the cell cortex Figure 1, A ; Schmit and Lambert, ; Liu and Palevitz, ; Panteris et al.
Cortical telophase microtubules interact with TANGLED1 TAN1 and other division site-localized proteins through transient microtubule plus-end stabilization, leading to their perpendicular orientation, similar to the orientation of the phragmoplast itself Bellinger et al. Microtubule plus-end capture is consistent with the in vitro role of TAN1 angle-independent microtubule crosslinking and bundling Martinez et al.
Cortical telophase microtubules are added into the phragmoplast as it approaches the cell cortex. Further, if many cortical telophase microtubules accumulate on one side of the phragmoplast, they incorporate by parallel bundling into phragmoplast and then phragmoplast expansion along the cortex moves toward the accumulated cortical telophase microtubules.
Therefore, cortical telophase microtubules fine tune the position of the phragmoplast to the division site, but it is still unknown how microtubule-binding proteins at the division site other than TAN1 interact with cortical telophase microtubules Bellinger et al. Model of land-plant cytokinesis. A, The cell cortex underlying the plasma membrane before it comes into contact with the phragmoplast. B, Phragmoplast, membrane, and cell wall structures that form the cell plate from left to right: vesicle accumulation, vesicle fusion to form a tubulo-vesicular network, a tubular network, and fenestrated sheet.
Proteins that accumulate at the division site before the phragmoplast contacts the cell cortex often have essential roles in cytokinesis or phragmoplast positioning. TAN1 is critical for phragmoplast positioning in maize, and bundles microtubules in vitro Cleary and Smith, ; Walker et al. In Arabidopsis, tan1 mutants do not have aberrant phenotypes Walker et al. In contrast, tan1 air9 double mutants have division plane defects and short roots Mir et al.
Mitotic expression of TAN1 restores wild-type growth to the tan1 air9 double mutant Mills et al. The AIR9 protein localizes to the phragmoplast and accumulates at the division site after the phragmoplast contacts the cortex Buschmann et al.
Assessing localization of these division site proteins in various mutants will provide useful information about how distinct factors establish and maintain the division site until cytokinesis is complete.
Several other proteins also localize to the division site and are vital for cytokinesis but their functions in division plane positioning are unclear. When the phragmoplast reaches the cortex, interactions between the phragmoplast and cortex-localized microtubules and actin filaments guide the phragmoplast precisely to the division site.
F-actin, Myosin, and cell cortex interactions play an important role in the last slow step of phragmoplast expansion to meet with the mother cell plasma membrane: disruption leads to defects in final phragmoplast-mediated contact with the cell cortex Chua et al.
Interestingly, when the phragmoplast is displaced by centrifugation, microns long actin filaments contact the division site. The phragmoplast is subsequently recruited back to the division site, suggesting long distance actindivision site interactions Arima et al. Much remains to be learned about how actin and microtubules, and the proteins that crosslink them, function together to guide the phragmoplast to the division site and to complete cytokinesis.
Microtubule regulators such as nucleators, severing proteins, end-binding proteins, and cross-linkers are vital for proper cellular elongation and cytokinesis Hamada, Their roles can often be clarified using genetic and biochemical analyses. While many microtubule-associated proteins are essential for both diffuse growth and cytokinesis, others have more specific roles.
Microtubule nucleators such as gamma tubulin, and the gamma-tubulin ring complex are essential for diffuse growth and cytokinesis Liu et al. Unlike animal cells, microtubules nucleate from the plasma membrane, the nucleus, or from preexisting microtubules Murata et al.
During cytokinesis, the phragmoplast expands by addition of new microtubules nucleating on the phragmoplast leading edge Figure 1, B. Therefore, microtubule-dependent nucleation is critical for phragmoplast expansion Smertenko et al. Phragmoplasts in ede1 mutants are misformed, but no cytokinetic defects are observed Lee et al.
Gamma-tubulin ring complexes and AUGMIN complex proteins play both conserved and specific roles in microtubule nucleation during diffuse growth and cytokinesis. It will be interesting to identify how specificity is determined during specific cell-cycle stages. Microtubule severing plays critical roles in microtubule array reorientation in response to intrinsic and extrinsic cues, including cell elongation and phragmoplast assembly and dynamics.
Although mutants are viable, cell elongation and phragmoplast expansion rates are reduced in ktn1 mutants. KTN1 is required for microtubule reorientation in response to mechanical force, suggesting that severing promotes microtubule reorientation Bouquin et al. KTN1 is also required for proper phragmoplast expansion, positioning, and morphology Panteris et al. In the ktn1 mutant, cell shape, microtubule orientation, and reorganization in response to mechanical cues are disrupted Komis et al.
Circumferential microtubule orientation propagated across several cell files around the ablation location in wild-type shoot apical meristem cells while in ktn1 mutants circumferential orientation was limited to a single cell file Uyttewaal et al.
KTN1 is required for blue light-induced reorientation Lindeboom et al. Various locations of SPR2 on microtubule-minus ends and microtubule crossovers may allow them to have opposite functions. Similarly, the microtubule bundling performed by MAP also prevents KTN1-mediated severing in vitro Burkart and Dixit, , within a time frame similar to induced cortical tension change Colin et al. However, the mechanisms for mechanical force-dependent and blue light-induced microtubule reorientation are likely different.
MOR1 promotes microtubule bundling and polymerization and localizes to cortical microtubules, and mitotic microtubule arrays Twell et al.
The temperature-sensitive mor1 mutants have been used to clarify roles in regulating microtubule dynamics and aberrant cell plate formation Eleftheriou et al. Interestingly despite the loss of parallel cortical microtubules and reduced microtubule polymer mass in the mor1 mutant at restrictive temperature, cell wall crystallinity remained high, suggesting that microtubule mass is inversely related to cellulose crystallinity Fujita et al.
CLASP, which preferentially localizes to cell edges, promotes microtubule passage across cell edges by preventing edge-induced microtubule depolymerization Ambrose et al. MAP65 family members typically bundle microtubules Smertenko et al.
MAP is required for axial growth in dark-grown hypocotyls: map mutants have short hypocotyls. Generation of a double mutant with the related map strongly reduces both light- and dark-grown hypocotyl elongation. The synthetic phenotype of the map map double mutant indicates that these two proteins have overlapping roles in cell elongation Lucas et al. A few MAP65 family members play unique roles in phragmoplast organization during cytokinesis.
MAP localizes to the phragmoplast midline and crosslinks antiparallel microtubules through its unique C-terminal domain. MAP, through multiple protein interactions, acts as a hub during cytokinesis. Double bub3;1 bub3;2 mutants grow normally. However, when bub3;1 bub3;2 mutants are treated with caffeine, a drug known to disrupt phragmoplast attachment and maturation, MAP localization was abolished at the midline. Subsequent defects include phragmoplast expansion defects, short roots, and altered root cell morphology Zhang et al.
Interaction between MAP and its partners is required for their midline accumulation Herrmann et al. The localization and turnover of MAP from the phragmoplast midline in phosphoinositide kinase double mutants pi4k-beta1 pi4k-beta2 is altered.
This is consistent with the overstabilization of the phragmoplast, possibly due to an endocytosis defect Lin et al. Restriction of the phragmoplast midline mediated by MAP65s in moss promotes timely vesicle fusion Kosetsu et al. Overall, MAP performs critical functions during cytokinesis by crosslinking antiparallel microtubules, recruiting or retaining proteins at the phragmoplast midline often through direct interaction, and promoting vesicle fusion. Interestingly, MAP localizes to the very leading edge of the phragmoplast, where it crosslinks antiparallel microtubules Murata et al.
Similarly, single map mutants do not have an aberrant phenotype, but when combined with map mutants, significant cytokinesis defects cause gametophyte lethality Li et al. MAP faintly localizes to the phragmoplast or phragmoplast midline Smertenko et al. In the map mutant, MAP localizes more prominently in the phragmoplast midline to partially replace MAP antiparallel bundling function Li et al.
MAP bundles parallel and antiparallel microtubules, and promotes rescue in vitro Fache et al. MAP also localizes to the division site but its function there is unknown Li et al.
Further combinatorial mutant analysis of this important family of microtubule-bundling proteins will clarify unique and synergistic functions. Kinesins are mostly microtubule and sometimes also actin binding motor proteins that transport cargo including vesicles, organelles, and sometimes actin filaments or microtubules. Kinesins in plants either move toward the dynamic plus-end of microtubules or move toward the less dynamic minus-end of microtubules.
The phragmoplast directs the movement of Golgi-derived vesicles, likely transported by plus-end directed kinesins, toward the newly developing cell plate.
Indeed, even synthetic vesicles are efficiently transported to the newly developing cell plate Esseling-Ozdoba et al. Although phragmoplast microtubules with attached vesicles have been observed by electron microscopy Otegui et al. Many plus-end directed kinesins eventually accumulate in the phragmoplast midline, possibly reflecting the sheer abundance of microtubule plus-ends. It is likely that many plus-end directed kinesins transport vesicles containing both cell wall enzymes and precursors as well as proteins essential for regulating microtubule dynamics and vesicle fusion.
FRA1 is a highly processive plus-end directed kinesin Zhu and Dixit, Reduced mechanical strength in fra1 was attributed to defects in cellulose microfibril patterning Zhong et al. While FRA1 does not affect cellulose synthesis directly, it associates with cellulose synthase microtubule-uncoupling proteins CMUs , proteins that bind microtubules, localize to the cell plate, and prevent cell twisting Zhu et al. Microtubule detachment from the cortex may affect the delivery of polysaccharides to the apoplast, contributing to the cell wall defects in fra Zhu and Dixit, ; Zhu et al.
Indeed, fra has reduced secretion of fucose-alkyne-labeled pectin Zhu et al. However, fra does not have any cytokinesis defects Zhu et al. Mutants have reduced cell number but lack obvious microtubule structural defects during cytokinesis Zhang et al. KINc suppresses microtubule plus-end growth in vitro, so KIN4s may restrict microtubule overlap by inhibiting microtubule growth. Increased microtubule overlap regions accumulate more vesicles generating thick cell plates. Fascinatingly, the kina kinc double mutant phragmoplast does not reorient, indicating division plane defects de Keijzer et al.
In the male microspore, callose accumulates in failed cell plates in the pakrp1 pakrp1l double mutants. However, vegetative cytokinesis is normal, indicating that there are additional, or different, proteins required for vegetative phragmoplast assembly Lee et al.
These Kinesin proteins are discussed here because their roles in specific divisions in the microspore and their interaction with TIO suggest that other plus-end directed kinesins may functionally rescue double mutants during vegetative growth, highlighting the need to generate higher-order mutant combinations or otherwise identify proteins that likely traffic vesicles essential for cytokinesis.
POK2 is required for fast phragmoplast expansion Herrmann et al. Several kinesin-7 plus-end directed kinesins are essential for cytokinesis. NACKs are processive plus-end directed kinesins, suggesting that they may transport cell plate building material Naito and Goshima, Although another group of kinesin-7s are not essential for cytokinesis, they form a complex that promotes microtubule polymerization with a caspase-like protein essential for cytokinesis called Extra Spindle Poles1 Moschou et al.
PAKRP2 is a processive plus-end-directed kinesin that moves with an unusual combination of small step size and frequent steps to nearby filaments Gicking et al. Processive movement and BFA sensitivity suggest it may transport Golgi-derived vesicles on the phragmoplast to the cell plate Lee et al. The double mutants also have chloroplast positioning defects, which may interfere with phragmoplast expansion and lead to cytokinesis defects Hiwatashi et al. The minus-end-directed Kinesin-5 family plays a vital role in cytokinesis in moss, tobacco, and Arabidopsis but it may not participate directly in diffuse growth.
A tobacco Kinesin-5, Kinesin-Related Protein NtKRP , localizes to microtubules, and translocates antiparallel phragmoplast microtubules to minimize their overlap Asada et al. The related Kinesin-5 AtKRPc has a temperature-sensitive mutant, radially swollen7 rsw7 , with spindle collapse, defective phragmoplast assembly, and cytokinesis defects.
Although rsw7 mutants have swollen cells suggesting defects in diffuse growth, and AtKRPc localizes to cortical microtubules, the mutant phenotypes observed in mitosis and cytokinesis may account for apparent defects in cell shape Bannigan et al. In moss, RNAi knockdown of all Kinesin-5 a—d homologs result in similar cytokinesis defects. Counterintuitively, Kinesin-5s are depleted from the phragmoplast midline via minus-end-directed movement Miki et al.
It is unclear how Kinesin-5 crosslinks or translocates antiparallel microtubules when it is excluded from antiparallel microtubule accumulation at the midline. Several Kinesin family minus-end-directed kinesins participate in cytokinesis. Kinesins have diverse roles in spindle formation, chloroplast, and nuclear movement Gicking et al.
This suggests other related proteins may functionally compensate during vegetative cytokinesis. Another Kinesin, kinesin with calponin homology KCH, calponin homology domains suggest actin binding , binds both actin and microtubules, and localizes to mitotic microtubule structures Xu et al. Mutants have tip growth defects in moss Yamada and Goshima, and cell elongation defects in rice Frey et al.
However, these proteins do have an obvious role in cytokinesis. Microinjection of a calmodulin-domain-specific antibody that is thought to constitutively activate KCBP during anaphase causes aberrant phragmoplast positioning and assembly and subsequent cytokinesis defects Vos et al. Cellulose is a major load-bearing component in the primary cell wall. It is composed of long chains of beta-1,4-linked glucans. Cellulose is made at the plasma membrane by a large complex named the CSC.
The CSC was originally observed at the plasma membrane underlying microtubules using freeze fracture as a hexameric rosette structure Mueller and Brown, Mutants in cellulose synthase subunits produce tiny plants with cell elongation and sometimes cytokinesis defects.
The cesA1 mutant has defects in cytokinesis Beeckman et al. Isoxaben is an herbicide that disrupts cellulose synthesis. However, the exact inhibition mechanism is unknown Scheible et al. Other chemicals, such as the cellulose biosynthesis inhibitor Endosidin 20, are useful tools to understand how CESA reaches the plasma membrane.
Multiple ces6 alleles were identified in a screen for mutants resistant to Endosidin20 Huang et al. Transgenic lines carrying mutations in catalytic residues of cellulose synthase fail to localize to the plasma membrane, phenocopied by Endosidin20 treatment, highlighting Endosidin20 as a powerful tool to study CESA localization to the plasma membrane.
This suggests an intimate connection between the cell wall and cell-wall integrity pathways to limit growth under conditions disrupting the cell wall Wolf et al. Altogether, herbicides that disrupt cellulose synthesis provide valuable tools for understanding assembly and trafficking of CESAs. It was long proposed that the CSC interacts with microtubules at the plasma membrane by an elusive linker protein Heath, CESA particles fail to track with microtubules in the csi1 mutant, and their velocity is reduced Lei et al.
Two homologs have different expression levels: CSI3 is expressed in meristematic zones while CSI2 expression is not detected in all tissues. Single csi3 mutants have no obvious aberrant phenotype but csi1 csi3 double mutants have slower CESA particle movement, reduced cellulose content, and smaller dark-grown hypocotyls Lei et al. It is currently unclear whether CSI1 or its homologs are essential during cytokinesis.
Generating a triple mutant would be required to clarify whether they are required for cytokinesis. Model of trafficking of non-cellulosic polysaccharides and cellulose in the primary cell wall. The synthesis of non-cellulosic polysaccharides occurs in the Golgi. Non-cellulosic polysaccharides include xyloglucan and pectins. They are exported along actin filaments or microtubules in vesicles from the Golgi to the plasma membrane and incorporated into the apoplast.
Cellulose is synthesized exclusively at the plasma membrane. The components involved in cellulose synthesis and its regulation are highlighted in the box on the right. The accumulation of CSCs at the plasma membrane is influenced by exocytosis and endocytosis. Microtubules and actin filaments play important roles in the trafficking of CSCs to and from the membrane. There is mounting evidence that the regulation of cell growth is represented by a triangle that connects the cytoskeleton, cell wall, and cell shape.
Microtubules guide the deposition of the cell wall and therefore determine the final shape of the cell. Cell geometry feedback influences the orientation of microtubules. CSCs are physically linked to microtubules via CSI1, highlighting the importance of the discovery of the key molecular linker underlying the parallel relationship between cellulose microfibrils and microtubules.
However, despite the observation that CSC trajectories were uncoupled from the cortical microtubules in csi1 null mutants Li et al. The transverse orientation of cellulose microfibrils is independent of CSC-microtubule linkage. However, the crossed-polylamellate wall architecture was lost in csi1. The loss of crossed-polylamellate wall was phenocopied by removing microtubules with the microtubule-depolymerizing drug oryzalin Xin et al.
These results are consistent with an earlier study showing that perturbation of microtubule rotation abolished the crossed-polylamellate wall texture Chan et al.
The crossed-polylamellate wall is important for restricting cell expansion laterally and promoting longitudinal expansion as loss of crossed-polylamellate wall coincides with loss of auxin-induced elongation Xin et al. Later, a stronger mutant allele was identified that affected cytokinesis: KOR1 is also essential for cell plate formation Lane et al.
The transport of KOR1 to the cell plate requires two short amino acid motifs but these motifs are not required for localization at the plasma membrane Zuo et al. Although the exact role of KOR1 is unknown, glucanase activity is required for efficient cellulose biosynthesis Nicol et al.
CC1 and 2 are two related plant-specific transmembrane domain-containing proteins that together prevent growth cessation after exposure to salt stress. CC1 binds and bundles microtubules through four hydrophobic motifs that may promote microtubule reorganization after salt-stress Kesten et al. Additional proteins that are less well understood but important for cellulose synthesis and cell elongation include KOBITO, named for a Japanese word meaning small because the mutants are stunted, and COBRA COB , named because the mutant roots had a snake-like shape Schindelman et al.
COB is a glycosyl—phosphatidyl inositol-anchored protein. COB localizes to the Golgi and cell wall Roudier et al. In maize, root-hairless3 , a monocot-specific COB homolog, promotes root hair growth and higher grain yield Hochholdinger et al. BC1 binds crystalline cellulose in vitro through an N-terminal domain and localizes to vesicles at the plasma membrane and in the cell wall Liu et al. It is hypothesized that COB modulates cellulose assembly by its direct interaction with crystalline cellulose Liu et al.
The rosette CSCs were observed in the Golgi by electron microscopy, suggesting they are assembled there Haigler and Brown, Tracking fluorescent protein-tagged CESA movement revealed that CSC delivery coincides with Golgi pausing directly beneath the insertion site near microtubules Crowell et al. Prior to membrane fusion, a third type of CSC-containing vesicle co-localizes with the exocyst complex, an evolutionarily conserved vesicle tethering complex that regulates multiple membrane trafficking processes.
CSI1 is a central hub for delivery of CSCs by regulating the tethering of CSC-containing vesicles along microtubules, by defining the domain in the plasma membrane for delivery, possibly by directly interacting with plasma membrane lipids via its C2 domain Gu and Somerville, , and by bridging multiple cellular components including the exocyst complex, PATROL1 PTL1 and microtubules. Exocyst subunit mutants are defective in delivering CSCs to the plasma membrane Zhu et al. PTL1 also accumulates at the early cell plate, but what it does there is not yet known Maeda et al.
CSC transport to the cell plate likely occurs through three different mechanisms. Cellulose is preferentially labeled with the Scarlet Pontamine dye Anderson et al. Cellulose microfibrils are also observed either during or after cell plate fusion with the plasma membrane using field emission scanning electron microscopy Fujita and Wasteneys, Second, when the phragmoplast disassembles in the middle of the cell, motile BFA-sensitive organelles, perhaps Golgi, transport CESA to the cell plate.
Finally, when the cell plate fuses with the plasma membrane, CESAs move from the plasma membrane into the cell plate Miart et al. Much remains to be discovered about how and when CESAs accumulate at the cell plate, and how or when they might interact with microtubules to initiate cellulose synthesis.
Combined light and electron microscopy techniques will be essential for resolving these questions, discussed beautifully here Chen et al. While actin is not essential for formation or insertion of CSCs near microtubules, actin is required for the cell-wide distribution and movement of CSC-containing Golgi bodies and rapid CSC insertion Gutierrez et al.
A direct connection between the actin cytoskeleton and exocyst complex was recently identified in moss with the exocyst protein SEC10 containing a formin domain van Gisbergen et al. CSC secretion is affected in 7tm1 7tm5 double mutants under normal conditions even though mutants do not show any phenotype without isoxaben treatment. The increased density of CSCs in shou4 shou4l double mutant epidermal cells presumably reflects increased rates of exocytotic CSC delivery.
An elevated level of amorphous cellulose was detected within mutant inflorescence stems Polko et al. The CSC is a large plasma membrane-localized complex that is predicted to be regulated via endocytosis Lei et al. During cytokinesis, clathrin-mediated endocytosis removes CESA particles both from the middle of the cell plate and the plasma membrane Miart et al.
The dynamin-like protein, DRP1A is required for clathrin-mediated endocytosis and cytokinesis, and colocalizes with clathrin light chain Konopka et al.
A mutant locus in rice, brittle culm 3 , mapped to a dynamin-related gene, OsDRP2B , which is likely required for clathrin-mediated endocytosis. In ap2m knockout mutants, CESA particle density increases as a result of disrupted clathrin-mediated endocytosis Bashline et al.
Recent generation of a temperature sensitive tplate mutant will be a powerful tool to clarify its role in both diffuse growth and cytokinesis Wang et al. Unlike cellulose, which is synthesized at the plasma membrane and in the late cell plate in the late stages of development, the synthesis of non-cellulosic polysaccharides occurs primarily in the Golgi Camirand et al.
Major non-cellulosic polysaccharides of the primary cell wall including xyloglucan and pectin are discussed here. One important resource in identifying specific features of carbohydrates comes from a suite of well-characterized monoclonal antibodies Pattathil et al. Xyloglucans are beta-1,4-linked glucans, similar to cellulose, with additional sugar modifications on branched side chains.
Modification of xyloglucans, based on immunolocalization of terminal fucose antigens, likely occurs within the trans cisternae of the Golgi and the TGN Zhang and Staehelin, Consistent with this hypothesis, oligosaccharide-mass-profiling of Golgi-enriched fractions revealed less substituted xyloglucans compared with the apoplastic cell wall fractions Obel et al.
In phosphatidylinositol 4-kinase b1 and b2 pi4kb1 pi4kb2 mutants, because the vesicles were unusually large, xyloglucan transport is observed within secretory vesicles from trans Golgi to TGN and to the cell surface Kang et al. However, xyloglucan-synthesizing enzymes such as xylosyltransferase XT1 and galactosyltransferase MUR3 are detected in cis and medial cisternae of Golgi using immunogold labeling Chevalier et al.
While it is possible that XT1 and MUR3 are mislocalized in cis and medial Golgi due to overexpression, further investigation will clarify whether the initiation of xyloglucan side chains occurs in early Golgi compartments.
In addition to accumulating at the plasma membrane through the Golgi, xyloglucan accumulates in cell plates. Xyloglucan cell plate accumulation occurs in many species and cell types, including Arabidopsis endosperm, maize root cells, and tobacco cultured cells Moore and Staehelin, ; Otegui and Staehelin, ; Sonobe et al. Xyloglucan is trafficked through peripheral Golgi, to vesicles that are likely transported on microtubules to the cell plate Moore and Staehelin, Consistently, xyloglucan accumulates faintly on BY-2 phragmoplasts Sonobe et al.
The endoxyloglucan transferase protein, which modifies xyloglucans, co-localizes with the ER, the phragmoplast, and the cell plate Yokoyama and Nishitani, SYP61 is a syntaxin that co-localizes with a subset of the TGN, and plays a key role in trafficking to the plasma membrane Sanderfoot et al.
SYP61 vesicles contain both terminal fucose and galactosylated xyloglucans as well as pectins described in more detail below. Interestingly, the galactosylated xyloglucan is also present in cell wall fractions, indicating that substituted xyloglucan may already be in its final form in SYP61 TGN-derived vesicles Wilkop et al.
It is unlikely that SYPderived vesicles carry all non-cellulosic polysaccharides from the Golgi to the apoplast. Adaptation of this approach using different GFP-tagged proteins in wild-type and mutant lines will aid the characterization of polysaccharide transport. Pectin synthesis likely begins in the cis Golgi cisternae.
The addition of side chains and subsequent modification occur in late Golgi compartments. Antibodies recognizing pectin-specific epitopes show that rhamnogalacturonan-I RG-I modification occurs in trans Golgi and TGN in sycamore maple Acer pseudoplatanus cells and alfalfa Medicago sativa root border cells Zhang and Staehelin, ; Wang et al.
A comprehensive study using 39 monoclonal antibodies against pectin epitopes revealed that various degrees of substitution of pectin are enriched in SYPderived vesicles Wilkop et al. We are far from a complete picture of transport of non-cellulosic polysaccharides from specific Golgi compartments to the apoplast.
For example, little is known about the localization and assembly of RG-II. Developing techniques that distinguish transport of complex non-cellulosic polysaccharides through specific routes remains a major challenge. Mutants have defects in cell wall expansion and also produce cell wall stubs indicative of cytokinesis defects Li et al. CSLD5 localizes to punctate structures and accumulates in the cell plate Gu et al. The cell wall material generated by this enzyme is unknown, although it may be mannans or xylans Liepman et al.
Interestingly, the Arabidopsis clsd5 cytokinesis defect is enhanced by combining csld5 together with either the csld2 or the clsd3 mutant, even though CSLD2 and CSLD3 typically regulate root-hair growth Yin et al. Unambiguously identifying which cell wall material is generated by these cellulose-synthase like proteins will provide valuable insight into their roles.
Vesicles arrive between interdigitated phragmoplast microtubules de Keijzer et al. RAB-A5c localizes to the cell plate and is essential for cytokinesis and also diffuse growth Kirchhelle et al. RAB-A1a-c may function redundantly: triple mutants are hypersensitive to the chemical Endosidin1 during cytokinesis Qi and Zheng, Multiple subunits localized to the early and post-cytokinetic cell plate. Exocyst component mutants have defects in initial cell plate assembly Fendrych et al.
The fused vesicles form dumbbell-shaped structures that then coalesce into a tubular—vesicular network Samuels et al. Dynamin-related proteins DRPs , large GTPases essential for both cell plate formation and endocytosis, encircle membranes at the cell plate Gu and Verma, ; Kang et al.
In Arabidopsis, overexpression of the essential cytokinetic dynamin, phragmoplastin, causes overaccumulation of callose, possibly due to phragmoplastin-callose synthase CALS1 interaction Hong et al.
Mutants that disrupt sterol formation or modification often have defective cytokinesis. DRP1A and sterols co-localize in the cell plate: removal of either disrupts localization and cell plate formation, suggesting their mutual requirement for high-lipid order in the cell plate Frescatada-Rosa et al. The smt2 smt3 double mutants have misoriented phragmoplasts and cell wall stubs, which are partially rescued by exogenous application of sterols Nakamoto et al.
Other mutants with defects in sterol synthesis or modification also have defects in cytokinesis, likely due to defective endocytosis, independent of a role in steroid hormone brassinosteroid biosynthesis Willemsen et al.
Other lipids are required for proper cytokinesis, including very long chain fatty acids, sphingolipids, diacylglycerol, and phosphoinositides Bach et al. Callose, a prominent component in the cell plate Drakakaki, , is rarely represented in primary cell walls, except in plasmodesmata Wu et al.
Callose is composed of beta-1,3-linked glucans unlike the beta-1,4-linked glucan chains found in xyloglucans and cellulose. Multiple callose synthases have both discrete and overlapping functions in symplastic transport through plasmodesmata Wu et al.
Mixtures of cellulose with high amounts of callose provide more elasticity in vitro Abou-Saleh et al. Biophysical modeling suggests that callose polymerization may provide the spreading force required to promote flattening of the tubulo-vesicular network into the fenestrated sheet Jawaid et al. Callose is synthesized by large protein complexes containing callose synthase Glucan-synthase-like integral membrane proteins Schneider et al.
A clever approach to circumvent death of callose deposition defective mutants uses a chemical, Endosidin7 ES7 , that prevents callose accumulation Drakakaki et al. ES7 prevents callose synthesis in vitro by partially blocking UDP-glucose incorporation into 1,3-linked glucans from Arabidopsis cell membrane extracts.
Surprisingly, ES7 does not alter wound- or stress-induced callose accumulation Park et al. The specific target of ES7 is unknown, but blocking callose synthesis with ES7 affects cytokinesis in both land plants and algae, suggesting that the target of ES7 is highly conserved Park et al. Callose synthase activity depends on the presence of calcium Kauss, ; Amor et al.
The ER may be a calcium source for callose synthases during cell plate formation: ER—cell plate association occurs in diverse plant lineages Porter and Machado, ; Schmiedel et al. Intense calcium accumulation in the cell plate has been observed via staining Wick and Hepler, ; Schmiedel et al. The P. One tempting hypothesis is that the slow callose accumulation in the sabre mutant may be due to aberrant calcium gradients near the cell plate due to altered ER connections.
Cell wall modification and construction are dynamic processes during which secretion and endocytosis are tightly controlled. There are many open questions remaining such as how cellulose and non-cellulosic polymer insertion is coordinated, how non-cellulosic polymers are trafficked between the Golgi and plasma membrane and on which cytoskeletal tracks, and how the cytoskeleton and CSC activity are influenced by mechanical and environmental cues.
Provided in Supplemental Data Set S1. Supplemental Data Set S1. Summary of the cytoskeletal and cell wall proteins required for diffuse growth and cytokinesis. Purple text indicates that proteins are involved in both diffuse growth and cytokinesis. Red text indicates that proteins are likely involved in only diffuse growth.
Blue text indicates that proteins are likely involved only in cytokinesis. Rasmussen crasmu ucr. We gratefully acknowledge figure design assistance from Donghui Wei and Dr.
Morgane Gilliard. We apologize to colleagues whose relevant work was not discussed due to length limitations. Nat Commun 9 : 1 — Google Scholar. J Exp Bot 69 : — Plant Cell 29 : — Nat Commun 2 : Ambrose C , Wasteneys GO Microtubule initiation from the nuclear surface controls cortical microtubule growth polarity and orientation in Arabidopsis thaliana. Plant Cell Physiol 55 : — Plant Cell 19 : — J Exp Bot 67 : — Plant Physiol : — New Phytol : — J Plant Res : — J Cell Sci Pt 2 : — J Cell Biol : — Plant J 93 : — J Cell Sci : — Protoplasma : — Int J Mol Sci 20 : Role of cortical microtubules and cellulose microfibrils.
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When a ribosome begins making proteins, the translocon opens enough for the newly created protein to feed into the pore of the endoplasmic reticulum. The new protein passes into the pore in a linear or helical form, because the pore is too small to allow a folded protein to pass within. The translocon pore only opens if it recognizes a special sequence of amino acids that ribosomes use to start a newly created protein.
The translocon controls whether the new protein will be incorporated into the plasma membrane or will be stored in soluble form within the ER.
The proteins that enter the tight confines of the ER membranes get bent and folded into their characteristic final shapes. These shapes result in part from atomic bonds between different portions of the protein molecule. The ER performs "quality control" by transporting abnormal or misshaped proteins back into the cell body where they are recycled.
Stored proteins travel into another cell organelle, called the Golgi apparatus, and eventually exit the cell via a vesicle.
When the ribosome finishes synthesizing a protein, the translocon ejects the ribosome and plugs up the pore until another protein needs to be synthesized. He holds an M. You can see samples of his work at ericbank. Importance of Free Ribosomes. Steps of DNA Transcription. How Cell Organelles Work Together. The Location of Ribosomes in a Cell. What Are the Biomolecules of Ribosomes?
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