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Anisotropic cell expansion is crucial for the morphogenesis of land plants, as cell migration is restricted by the rigid cell wall. The anisotropy of cell expansion is regulated by mechanisms acting on the deposition or modification of cell wall polysaccharides. Besides the polysaccharide components in the cell wall, a layer of hydrophobic cuticle covers the outer cell wall and is subjected to tensile stress that mechanically restricts cell expansion. However, the molecular machinery that deposits cuticle materials in the appropriate spatiotemporal manner to accommodate cell and tissue expansion remains elusive. Here, we report that PpABCB14, an ATP-binding cassette transporter in the moss Physcomitrium patens, regulates the anisotropy of cell expansion. PpABCB14 localized to expanding regions of leaf cells. Deletion of PpABCB14 resulted in impaired anisotropic cell expansion. Unexpectedly, the cuticle proper was reduced in the mutants, and the cuticular lipid components decreased. Moreover, induced PpABCB14 expression resulted in deformed leaf cells with increased cuticle lipid accumulation on the cell surface. Taken together, PpABCB14 regulates the anisotropy of cell expansion via cuticle deposition, revealing a regulatory mechanism for cell expansion in addition to the mechanisms acting on cell wall polysaccharides., (© 2023 The Authors New Phytologist © 2023 New Phytologist Foundation.)

Plant cells are surrounded by a cell wall and do not migrate, which makes the regulation of cell division orientation crucial for development. Regulatory mechanisms controlling cell division orientation may have contributed to the evolution of body organization in land plants. The GRAS family of transcription factors was transferred horizontally from soil bacteria to an algal common ancestor of land plants. SHORTROOT ( SHR ) and SCARECROW ( SCR ) genes in this family regulate formative periclinal cell divisions in the roots of flowering plants, but their roles in nonflowering plants and their evolution have not been studied in relation to body organization. Here, we show that SHR cell autonomously inhibits formative periclinal cell divisions indispensable for leaf vein formation in the moss Physcomitrium patens , and SHR expression is positively and negatively regulated by SCR and the GRAS member LATERAL SUPPRESSOR , respectively. While precursor cells of a leaf vein lacking SHR usually follow the geometry rule of dividing along the division plane with the minimum surface area, SHR overrides this rule and forces cells to divide nonpericlinally. Together, these results imply that these bacterially derived GRAS transcription factors were involved in the establishment of the genetic regulatory networks modulating cell division orientation in the common ancestor of land plants and were later adapted to function in flowering plant and moss lineages for their specific body organizations.

Proper orientation of the cell division axis is critical for asymmetric cell divisions that underpin cell differentiation. In animals, centrosomes are the dominant microtubule organizing centers (MTOC) and play a pivotal role in axis determination by orienting the mitotic spindle. In land plants that lack centrosomes, a critical role of a microtubular ring structure, the preprophase band (PPB), has been observed in this process; the PPB is required for orienting (before prophase) and guiding (in telophase) the mitotic apparatus. However, plants must possess additional mechanisms to control the division axis, as certain cell types or mutants do not form PPBs. Here, using live imaging of the gametophore of the moss Physcomitrella patens , we identified acentrosomal MTOCs, which we termed "gametosomes," appearing de novo and transiently in the prophase cytoplasm independent of PPB formation. We show that gametosomes are dispensable for spindle formation but required for metaphase spindle orientation. In some cells, gametosomes appeared reminiscent of the bipolar MT "polar cap" structure that forms transiently around the prophase nucleus in angiosperms. Specific disruption of the polar caps in tobacco cells misoriented the metaphase spindles and frequently altered the final division plane, indicating that they are functionally analogous to the gametosomes. These results suggest a broad use of transient MTOC structures as the spindle orientation machinery in plants, compensating for the evolutionary loss of centrosomes, to secure the initial orientation of the spindle in a spatial window that allows subsequent fine-tuning of the division plane axis by the guidance machinery., Competing Interests: The authors declare no conflict of interest., (Copyright © 2017 the Author(s). Published by PNAS.)

Aurora kinases are key effectors of mitosis. Plant Auroras are functionally divided into two clades. The alpha Auroras (Aurora1 and Aurora2) associate with the spindle and the cell plate and are implicated in controlling formative divisions throughout plant development. The beta Aurora (Aurora3) localizes to centromeres and likely functions in chromosome separation. In contrast to the wealth of data available on the role of Aurora in other kingdoms, knowledge on their function in plants is merely emerging. This is exemplified by the fact that only histone H3 and the plant homolog of TPX2 have been identified as Aurora substrates in plants. Here we provide biochemical, genetic, and cell biological evidence that the microtubule-bundling protein MAP65-1-a member of the MAP65/Ase1/PRC1 protein family, implicated in central spindle formation and cytokinesis in animals, yeasts, and plants-is a genuine substrate of alpha Aurora kinases. MAP65-1 interacts with Aurora1 in vivo and is phosphorylated on two residues at its unfolded tail domain. Its overexpression and down-regulation antagonistically affect the alpha Aurora double mutant phenotypes. Phospho-mutant analysis shows that Aurora contributes to the microtubule bundling capacity of MAP65-1 in concert with other mitotic kinases., (© 2017 American Society of Plant Biologists. All Rights Reserved.)

The phragmoplast, a plant-specific apparatus that mediates cytokinesis, mainly consists of microtubules (MTs) arranged in a bipolar fashion, such that their plus ends interdigitate at the equator. Membrane vesicles are thought to move along the MTs toward the equator and fuse to form the cell plate. Although several genes required for phragmoplast MT organization have been identified, the mechanisms that maintain the bipolarity of phragmoplasts remain poorly understood. Here, we show that engaging phragmoplast MTs in a bipolar fashion in protonemal cells of the moss Physcomitrella patens requires the conserved MT cross-linking protein MICROTUBULE-ASSOCIATED PROTEIN65 (MAP65). Simultaneous knockdown of the three MAP65s expressed in those cells severely compromised MT interdigitation at the phragmoplast equator after anaphase onset, resulting in the collapse of the phragmoplast in telophase. Cytokinetic vesicles initially localized to the anaphase midzone as normal but failed to further accumulate in the next several minutes, although the bipolarity of the MT array was preserved. Our data indicate that the presence of bipolar MT arrays is insufficient for vesicle accumulation at the equator and further suggest that MAP65-mediated MT interdigitation is a prerequisite for maintenance of bipolarity of the phragmoplast and accumulation and/or fusion of cell plate-destined vesicles at the equatorial plane.

Plant cytokinesis occurs by the growth of cell plates from the interior to the periphery of the cell. These dynamic events in cytokinesis are mediated by a plant-specific microtubule (MT) array called the phragmoplast, which consists of bundled antiparallel MTs between the two daughter nuclei. The NACK-PQR pathway, a NACK1 kinesin-like protein and mitogen activated protein kinase (MAPK) cascade, is a key regulator of plant cytokinesis through the regulation of phragmoplast MTs. The MT-associated protein MAP65 has been identified as one of the structural components of MT assays involved in cell division, and we recently showed that Arabidopsis AtMAP65-3/PLEIADE (PLE) is a substrate of MPK4 that is a component of the NACK-PQR pathway in Arabidopsis. Here we show that AtMAP65-1 and AtMAP65-2 are also phosphorylated by MPK4. AtMAP65-1 and AtMAP65-2 that localize to the phragmoplast were phosphorylated by MPK4 in vitro. Although mutants of the Arabidopsis AtMAP65-1 and AtMAP65-2 genes exhibited a wild-type phenotype, double mutations of AtMAP65-3 and AtMAP65-1 or AtMAP65-2 caused more severe growth and cytokinetic defects than the single atmap65-3/ple mutation. These results suggest that AtMAP65-1 and AtMAP65-2 also function in cytokinesis downstream of MPK4.

Cytokinesis in plants is achieved by the formation of the cell plate. A pathway that includes mitogen-activated protein (MAP) kinase kinase kinase and MAP kinase kinase (MAPKK) plays a key role in the control of plant cytokinesis. We show here that a MAP kinase, MPK4, is required for the formation of the cell plate in Arabidopsis thaliana. Single mutations in MPK4 caused dwarfism and characteristic defects in cytokinesis, such as immature cell plates, which became much more prominent upon introduction of a mutation in MKK6/ANQ, the MAPKK for cytokinesis, into mpk4. MKK6/ANQ strongly activated MPK4 in protoplasts, and kinase activity of MPK4 was detected in wild-type tissues that contained dividing cells but not in mkk6/anq mutants. Fluorescent protein-fused MPK4 localized to the expanding cell plates in cells of root tips. Expansion of the cell plates in mpk4 root tips appeared to be retarded. The level of MPK11 transcripts was markedly elevated in mpk4 plants, and defects in the mpk4 mpk11 double mutant with respect to growth and cytokinesis were more severe than in the corresponding single mutants. These results indicate that MPK4 is the downstream target of MKK6/ANQ in the regulation of cytokinesis in Arabidopsis and that MPK11 is also involved in cytokinesis.

Cytokinesis is regulated to ensure the precise partitioning of cytoplasm and duplicated chromosomes to daughter cells. The NACK-PQR pathway, which includes NACK1 kinesin-like protein (KLP) and a mitogen-activated protein kinase (MAPK) cascade, plays a key role in cytokinesis in tobacco cells. Although HINKEL/AtNACK1 (HIK) KLP, ANP MAP kinase kinase kinases (MAPKKKs) and MKK6/ ANQ MAP kinase kinase (MAPKK) have been identified independently as regulators of cytokinesis in Arabidopsis thaliana, the involvement of HIK, ANPs and MKK6/ANQ in a regulatory cascade remains to be demonstrated. Here we provide details of the protein kinase pathway that controls cytokinesis in A. thaliana. Analysis of the subcellular distribution of six MAPKKs of A. thaliana that had been fused to green fluorescent protein revealed that only MKK6/ANQ protein was concentrated at the equatorial plane of the phragmoplast, at the site of localization of HIK. Expression of MKK6/ANQ in yeast cells replaced the growth-control function of the MAPKK encoded by yeast PBS2, provided that both ANP1 MAPKKK and HIK [or TETRASPORE/AtNACK2 (TES)] KLP were coexpressed, suggesting that ANP1 activates MKK6/ANQ in the presence of HIK (or TES). Coexpression of HIK and ANP3 (another member of the ANP MAPKKK family) weakly activated MKK6/ANQ but that of TES and ANP3 did not. MKK6/ANQ phosphorylated MPK4 MAPK in vitro to activate the latter kinase. Thus cytokinesis in A. thaliana is controlled by a pathway that consists of ANP MAPKKKs that can be activated by HIK and MKK6/ANQ MAPKK, with MPK4 MAPK being a probable target of MKK6/ANQ.

The purpose of this study was to clarify which direction of the condylar path is advantageous for releasing TMJ intermittent lock. The subjects were ten patients with temporomandibular disorders (TMD) and intermittent lock caused by anterior displacement of disks without obvious medial or lateral displacement. The patients could not open their mouths fully in habitual opening but could open fully in an intentional winding opening with reduction of the anteriorly displaced disks (winding opening). The two kinds of movement at the kinematic condylar point were measured in each subject and compared. The length of the condylar path in winding opening was significantly larger than that in habitual opening. The affected side condyle of winding opening traced medio-inferior paths in the early part of the condylar translation in comparison with habitual opening. From the viewpoint of the condylar path, the medio-inferior direction of condylar translation is thought to be advantageous for releasing intermittent lock.

The purpose of this study was to clarify which direction of jaw movement is advantageous for releasing TMJ intermittent lock. The subjects included ten patients with TMJ intermittent lock who could not open their mouths fully in habitual opening paths (habitual opening), but could open fully in intentionally winding opening paths (winding opening). The following movements were analyzed: habitual opening, winding opening, lateral border opening to the contralateral side (contralateral opening), lateral border opening to the affected side and anterior border opening. The incisal points in winding opening tended to trace antero-lateral paths that were shifted toward the side opposite the affected joints in comparison with habitual opening. In addition to winding opening, the success rate in releasing locking of contralateral opening was significantly greater than that of habitual opening. The findings demonstrated that contralateral movements directed toward the nonaffected side are advantageous for releasing the intermittent lock. The patients will have a better chance of releasing the intermittent lock by themselves, when opening path shifted to the nonaffected side is recommended as the first choice.

This study investigated the energy metabolism of masseter muscles by 31P-Magnetic Resonance Spectroscopy (MRS) during increased blood flow induced by hot pack application to clarify the influence of changes in blood flow on muscle fatigue. Twelve healthy subjects with no history of muscle pain in the masticatory system participated in this study. The 31P-MRS measurements were performed before and after hot pack application and the ratio of phosphocreatine (PCr) acting as the energy source to reproduce ATP to beta-ATP, the PCr/beta-ATP ratio, was analyzed. Results showed that PCr/beta-ATP ratios increased significantly by an average of 22.4% after the hot pack application. The results suggest that changes in blood flow volume influence the energy metabolism in masseter muscles and that blood flow increases due to the hot pack cause higher energy levels in masseter muscles and offer an advantageous condition for preventing and relieving muscle fatigue.

Laterotrusive training is often used together with mouth-opening training in order to achieve adequate translation of the affected condyle in treatment of TMJ closed lock. This training is usually performed by voluntary laterotrusive movement (VLM) or by laterotrusive movement using only the fingers (FLM). However, satisfactory results are often not obtained by using these methods. To resolve this problem, we devised a new laterotrusive training device (LT device). In this paper, we describe the method of the training of laterotrusive movement using the LT device (LLM) and present a comparison of the results obtained by using LLM with those obtained by using VLM and FLM. The subjects were ten patients with TMJ closed lock. The following results were obtained: 1. the range of LLM was significantly larger than those of VLM and FLM; and 2. all of the patients reported that LLM could be performed more easily than VLM and FLM. In conclusion, the LT device is thought to be useful for laterotrusive training in TMJ closed lock.

Contracture of the jaw-closing muscles is one of the causes of limitation of jaw opening. This study examined whether there is any difference between the EMG activities of jaw-closing muscles during jaw opening in healthy people and in patients with masseter muscle contracture (MMC), who do not have a history of trauma or infection. The patient group consisted of eleven females, 18 to 62 years old with no history of trauma or infections, with limited mouth opening due to MMC. The control group included eleven healthy females, 23 to 50 years old. The EMG activity was recorded bilaterally in the central portion of masseter muscles (Mm), the anterior portion of temporal muscles (Tm), and the anterior belly of digastric muscles (Dm). Nine out of the patient group showed obvious EMG activity in Mm during jaw opening which was different from typical EMG patterns during jaw opening of up to 40 mm in the control group. Among the nine patients, eight showed antagonistic contraction in Tm as well as Mm. The mean integral value of Mm and Tm during jaw opening in the patient group was significantly higher than in the control group (P<0.01). These results suggest that EMG activity during jaw opening in MMC patients with no history of trauma or infections is different from that in healthy people.