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Objective: To investigate the time delay effect from initial diagnosis to endoscopic submucosal dissection on superficial esophageal squamous cell carcinoma curability, considering the preoperative invasion depth., Methods: This study included superficial esophageal squamous cell carcinoma diagnosed as T1a-epithelial/lamina propria mucosa cancer (cEP/LPM; cancer invading up to the lamina propria mucosa) or cT1a-muscularis mucosa (MM)/T1b-submucosal cancer (cMM/SM1; cancer invading up to 200 µm into the submucosa) and treated using endoscopic submucosal dissection from January 2017 to December 2021. We compared curability in lesions treated within three months (early treatment group) versus those treated ≥7 months post-diagnosis (delayed treatment group). Curative resection criteria included lesions confined within the muscularis mucosae, with negative vertical margins, and with absence of lymphovascular invasion. Non-curative resection included all other cases., Results: Among the 231 and 75 lesions in the early and delayed treatment groups, respectively, no significant difference was observed in non-curative resections for all lesions and cEP/LPM lesions (early: 194, delayed: 70). Conversely, the proportions were significantly higher in the delayed treatment group than in the early treatment group for cMM/SM1 lesions (early: 37, delayed: 5; p = 0.018)., Conclusions: This study suggests that delayed endoscopic submucosal dissection does not significantly affect cEP/LPM lesions curability but is associated with reduced cMM/SM1 lesions curability. Prompt treatment is important for cMM/SM1, whereas delayed treatment may be acceptable for cEP/LPM., Competing Interests: None., (© 2024 The Author(s). DEN Open published by John Wiley & Sons Australia, Ltd on behalf of Japan Gastroenterological Endoscopy Society.)

Gall-forming insects induce various types of galls on their host plants by altering gene expression in host plant organs, and recent studies have been conducted for gene expression in galls. However, the evolutionary trajectories of gene expression patterns and the resulting phenotypes have not yet been studied using multiple related species. We investigated the speciation and the diversification process of galls induced by four closely related aphid species (Hormaphidini) on a host plant species (Hamamelis japonica) by examining the phylogenetic congruence between the geographical divergences of aphids and the host plant, and by comparing their gene expression patterns and resulting phenotypes. Phylogenetic analysis of aphids and the host plant showed that geographical isolation among host plant populations has interrupted gene flow in aphids and accelerated the speciation process. The concentration of phenolics and the complexity of the internal structure of galls were correlated with the expression levels of genes for the biosynthesis of phenolics and morphogenesis respectively. These results suggest that the expression levels of genes for the biosynthesis of phenolics and morphogenesis have evolutionarily increased in galls accelerated by the speciation process of aphids due to the distribution change of the host plant, leading to the related phenotypic evolution. Our study showed the evolutionary process of phenotypic traits in galls in the wild from both gene expression and actual phenotype levels., (© 2024 The Author(s). Molecular Ecology published by John Wiley & Sons Ltd.)

Objectives: The resection of vertical margin-negative submucosally invasive colorectal cancer (CRC) relies on the pathological risk assessment of lymph node metastasis. However, no large-scale study has clarified the endoscopic resection (ER) outcome for submucosally invasive CRC, focusing on the vertical margin status. This retrospective study aimed to examine vertical margin involvement in ER for submucosally invasive CRC and explore the treatment consequences associated with vertical margin status., Methods: We analyzed 395 submucosally invasive CRC cases in 389 patients who underwent ER at our hospital between 2008 and 2020. The presence of residual tumors and simultaneous lymph node metastasis in patients who underwent additional surgery was assessed and compared between the vertical incomplete ER and the vertical margin-negative groups., Results: Among the patients, 270 were men, with a median age of 69 years. The vertical incomplete ER rate was 21.5%, with positive vertical margins and unclear vertical margins identified in 12.2% and 9.3% of the cases, respectively. Among 154 patients who underwent additional surgery after ER, the vertical incomplete ER group had a significantly higher residual tumor rate than the vertical margin-negative group (P = 0.001). The vertical incomplete ER group had a significantly higher lymph node metastasis rate than the vertical margin-negative group (P = 0.029)., Conclusion: This study clarified the substantial risk of vertical incomplete ER in submucosally invasive CRC and revealed the high risk of residual tumor and lymph node metastasis in vertical incomplete ER for submucosal CRC., (© 2023 Japan Gastroenterological Endoscopy Society.)

Arabidopsis ASYMMETRIC LEAVES2 (AS2) plays a key role in the formation of flat symmetric leaves. AS2 represses the expression of the abaxial gene ETTIN / AUXIN RESPONSE FACTOR3 ( ETT / ARF3 ). AS2 interacts in vitro with the CGCCGC sequence in ETT / ARF3 exon 1. In cells of leaf primordia, AS2 localizes at peripheral regions of the nucleolus as two AS2 bodies, which are partially overlapped with chromocenters that contain condensed 45S ribosomal DNA repeats. AS2 contains the AS2/LOB domain, which consists of three sequences conserved in the AS2/LOB family: the zinc finger (ZF) motif, the ICG sequence including the conserved glycine residue, and the LZL motif. AS2 and the genes NUCLEOLIN1 ( NUC1 ), RNA HELICASE10 ( RH10 ), and ROOT INITIATION DEFECTIVE2 ( RID2 ) that encode nucleolar proteins coordinately act as repressors against the expression of ETT / ARF3. Here, we examined the formation and patterning of AS2 bodies made from as2 mutants with amino acid substitutions in the ZF motif and the ICG sequence in cells of cotyledons and leaf primordia. Our results showed that the amino acid residues next to the cysteine residues in the ZF motif were essential for both the formation of AS2 bodies and the interaction with ETT / ARF3 DNA. The conserved glycine residue in the ICG sequence was required for the formation of AS2 bodies, but not for the DNA interaction. We also examined the effects of nuc1 , rh10 , and rid2 mutations, which alter the metabolism of rRNA intermediates and the morphology of the nucleolus, and showed that more than two AS2 bodies were observed in the nucleolus and at its periphery. These results suggested that the patterning of AS2 bodies is tightly linked to the morphology and functions of the nucleolus and the development of flat symmetric leaves in plants.

Background: Precise area diagnosis of early gastric cancer (EGC) is critical for reliable endoscopic resection. Computer-aided diagnosis (CAD) shows strong potential for detecting EGC and reducing cancer-care disparities caused by differences in endoscopists' skills. To be used in clinical practice, CAD should enable both the detection and the demarcation of lesions. This study proposes a scheme for the detection and delineation of EGC under white-light endoscopy and validates its performance using 1-year consecutive cases., Methods: Only 300 endoscopic images randomly selected from 68 consecutive cases were used for training a convolutional neural network. All cases were treated with endoscopic submucosal dissection, enabling the accumulation of a training dataset in which the extent of lesions was precisely determined. For validation, 462 cancer images and 396 normal images from 137 consecutive cases were used. From the validation results, 38 randomly selected images were compared with those delineated by six endoscopists., Results: Successful detections of EGC in 387 cancer images (83.8%) and the absence of lesions in 307 normal images (77.5%) were achieved. Positive and negative predictive values were 81.3% and 80.4%, respectively. Successful detection was achieved in 130 cases (94.9%). We achieved precise demarcation of EGC with a mean intersection over union of 66.5%, showing the extent of lesions with a smooth boundary; the results were comparable to those achieved by specialists., Conclusions: Our scheme, validated using 1-year consecutive cases, shows potential for demarcating EGC. Its performance matched that of specialists; it might therefore be suitable for clinical use in the future., (© 2023. Japanese Society of Gastroenterology.)

Purpose: The pathological diagnosis of surgically resected gastric cancer involves both a macroscopic diagnosis by gross observation and a microscopic diagnosis by microscopy. Macroscopic diagnosis determines the location and stage of the disease and the involvement of other organs and surgical margin. Lesion recognition is, thus, an important diagnostic step that requires a skilled pathologist. Nonetheless, artificial intelligence (AI) technologies could allow even inexperienced doctors and laboratory technicians to examine surgically resected specimens without the need for pathologists. However, organ imaging conditions vary across hospitals, and an AI algorithm created in one setting may not work properly in another. Thus, we identified and standardized factors affecting the quality of pathological macroscopic images, which could further affect lesion identification using AI., Methods: We examined necessary image standardization for developing cancer detection AI for surgically resected gastric cancer by changing the following imaging conditions: focus, resolution, brightness, and contrast., Results: Regarding focus, brightness, and contrast, the farther away the test data were from the training macro-image, the less likely the inference was to be correct. Little change was observed for resolution, even with differing conditions for the training and test data. Regarding focus, brightness, and contrast, there were conditions appropriate for AI. Contrast, in particular, was far from the conditions appropriate for humans., Conclusion: Standardizing focus, brightness, and contrast is important in the development of AI methodologies for lesion detection in surgically resected gastric cancer. This standardization is essential for AI to be implemented across hospitals., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Background: Endoscopic resection (ER) with bipolar snare, in which the electric current only passes through the tissue between the device's two electrodes, is a prominent method used to prevent perforation due to electricity potentially. ER using bipolar snare with or without submucosal injection enabled safe resection of colorectal lesions measuring 10-15 mm in an ex vivo porcine model. ER with bipolar snare is expected to have good treatment outcomes in 10-15 mm colorectal lesions, with high safety even without submucosal injection. However, no clinical reports have compared treatment outcomes with and without submucosal injection., Aim: To compare the treatment outcomes of bipolar polypectomy with hot snare polypectomy (HSP) to those with endoscopic mucosal resection (EMR)., Methods: In this single-centre retrospective study, we enrolled 10-15 mm nonpedunculated colorectal lesions (565 Lesions in 463 patients) diagnosed as type 2A based on the Japan Narrow-band Imaging Expert Team classification, resected by either HSP or EMR between January 2018 and June 2021 at the National Cancer Center Hospital East. Lesions were divided into HSP and EMR groups, and propensity score matching was performed. In the matched cohort, en bloc and R0 resection rates and adverse events were compared between the two groups., Results: Of the 565 lesions in 463 patients, 117 lesions each in the HSP and EMR groups were selected after propensity score matching. In the original cohort, there was a significant difference in antithrombotic drug use ( P < 0.05), lesion size ( P < 0.01), location ( P < 0.01), and macroscopic type ( P < 0.05) between the HSP and EMR groups. In the matched cohort, the en bloc resection rates were comparable between both groups [93.2% (109/117) vs 92.3% (108/117), P = 0.81], and there was no significant difference in the R0 resection rate [77.8% (91/117) vs 80.3% (94/117), P = 0.64]. The incidence of delayed bleeding was similar in both groups [1.7% (2/117)]. Perforation occurred in the EMR group [0.9% (1/117)] but not in the HSP group., Conclusion: Using bipolar snare, ER of nonpedunculated 10-15 mm colorectal lesions may be performed safely and effectively, even without submucosal injection., Competing Interests: Conflict-of-interest statement: All the authors report no relevant conflicts of interest for this article., (©The Author(s) 2023. Published by Baishideng Publishing Group Inc. All rights reserved.)

Objectives: We aimed to evaluate the efficacy of texture and color enhancement imaging (TXI), which allows the acquisition of brighter images with enhanced color and surface structure in colorectal polyp detection compared to white light imaging., Methods: Patients who underwent colonoscopy with repeated ascending colon observation using TXI and white light imaging between August 2020 and January 2021 were identified in three institutions. The outcomes included the mean number of adenomas detected per procedure (MAP), adenoma detection rate (ADR), and ascending colonic adenoma miss rate (Ac-AMR). Logistic regression was used to determine the effects of the variables on the outcomes., Results: We included 1043 lesions from 470 patients in the analysis. The MAP, ADR, flat polyp detection rate, and Ac-AMR in TXI and white light imaging were 1.5% (95% confidence interval 1.3-1.6%) vs. 1.0% (0.9-1.1%), 58.2% (51.7-64.6%) vs. 46.8% (40.2-53.4%), 66.2% (59.8-72.2%) vs. 49.8% (43.2-56.4%), and 17.9% (12.1-25.2%) vs. 28.2% (20.0-37.6%), respectively. TXI, age, withdrawal time, and endoscopy type were identified as significant factors affecting the MAP and the ADR using multivariate regression analysis., Conclusions: Our study indicates that TXI improve the detection of colorectal neoplastic lesions. However, prospective randomized trials are required to confirm these findings., (© 2022 The Authors. Digestive Endoscopy published by John Wiley & Sons Australia, Ltd on behalf of Japan Gastroenterological Endoscopy Society.)

The ASYMMETRIC LEAVES2 (AS2) gene in Arabidopsis thaliana is responsible for the development of flat, symmetric, and extended leaf laminae and their vein systems. AS2 protein is a member of the plant-specific AS2/LOB protein family, which includes 42 members comprising the conserved amino-terminal domain referred to as the AS2/LOB domain, and the variable carboxyl-terminal region. Among the members, AS2 has been most intensively investigated on both genetic and molecular levels. AS2 forms a complex with the myb protein AS1, and is involved in epigenetic repression of the abaxial genes ETTIN/AUXIN RESPONSE FACTOR3 (ETT/ARF3), ARF4, and class 1 KNOX homeobox genes. The repressed expression of these genes by AS2 is markedly enhanced by the cooperative action of various modifier genes, some of which encode nucleolar proteins. Further downstream, progression of the cell division cycle in the developing organs is stimulated; meristematic states are suppressed in determinate leaf primordia; and the extension of leaf primordia is induced. AS2 binds the specific sequence in exon 1 of ETT/ARF3 and maintains methylated CpGs in several exons of ETT/ARF3. AS2 forms bodies (designated as AS2 bodies) at nucleolar peripheries. AS2 bodies partially overlap chromocenters, including inactive 45S ribosomal DNA repeats, suggesting the presence of molecular and functional links among AS2, the 45S rDNAs, and the nucleolus to exert the repressive regulation of ETT/ARF3. The AS2/LOB domain is characterized by three subdomains, the zinc finger (ZF) motif, the internally conserved-glycine containing (ICG) region, and the leucine-zipper-like (LZL) region. Each of these subdomains is essential for the formation of AS2 bodies. ICG to LZL are required for nuclear localization, but ZF is not. LZL intrinsically has the potential to be exported to the cytoplasm. In addition to its nuclear function, it has been reported that AS2 plays a positive role in geminivirus infection: its protein BV1 stimulates the expression of AS2 and recruits AS2 to the cytoplasm, which enhances virus infectivity by suppression of cytoplasmic post transcriptional gene silencing., (© 2021. The Author(s).)

Unauthorized resource access represents a typical security threat in the Internet of Things (IoT), while distributed ledger technologies (e.g., blockchain and IOTA) hold great promise to address this threat. Although blockchain-based IoT access control schemes have been the most popular ones, they suffer from several significant limitations, such as high monetary cost and low throughput of processing access requests. To overcome these limitations, this paper proposes a novel IoT access control scheme by combining the fee-less IOTA technology and the Ciphertext-Policy Attribute-Based Encryption (CP-ABE) technology. To control the access to a resource, a token, which records access permissions to this resource, is encrypted by the CP-ABE technology and uploaded to the IOTA Tangle (i.e., the underlying database of IOTA). Any user can fetch the encrypted token from the Tangle, while only those who can decrypt this token are authorized to access the resource. In this way, the proposed scheme enables not only distributed, fee-less and scalable access control thanks to the IOTA but also fine-grained attribute-based access control thanks to the CP-ABE. We show the feasibility of our scheme by implementing a proof-of-concept prototype system using smart phones (Google Pixel 3XL) and a commercial IoT gateway (NEC EGW001). We also evaluate the performance of the proposed scheme in terms of access request processing throughput. The experimental results show that our scheme enables object owners to authorize access rights to a large number of subjects in a much (about 5 times) shorter time than the existing access control scheme called Decentralized Capability-based Access Control framework using IOTA (DCACI), significantly improving the access request processing throughput.

Aim: To determine the relationship between multiple medications and falls., Methods: This case-control and case-crossover study was carried out at Kudanzaka Hospital in Chiyoda, Tokyo, Japan. A total of 325 patients who experienced their first falls when hospitalized between January 2016 and November 2018, and 1285 controls matched by sex, age and clinical departments were included in this study. Hospitalization duration and fall risk score were adjusted for in the analyses., Results: In the case-control study, multivariable logistic regression showed that increasing the intake of oral medications was not significantly associated with the incidence of falls (odds ratio 1.02, 95% confidence interval 0.998-1.049). In contrast, drugs prescribed with special caution in accordance with the Elderly Oral Medication Guidelines were significantly associated with falls (odds ratio 1.17, 95% confidence interval 1.09-1.26). A similar pattern was observed in the case-crossover analysis. Among the drugs to be prescribed with special caution according to the guidelines, atypical antipsychotics, non-benzodiazepine hypnotics and magnesium oxide were significantly associated with the risk of falls., Conclusion: The drugs to be prescribed with special caution according to the guidelines were associated with an increased fall risk. The risk of falls in hospitalized older people due to multiple medications varies among medications. Geriatr Gerontol Int ••; ••: ••-•• Geriatr Gerontol Int 2021; ••: ••-••., (© 2021 Japan Geriatrics Society.)

The aim of this study was to examine the risk of falls associated with the use of non-gamma amino butyric acid (GABA) sleep medications, suvorexant and ramelteon. This case-control and case-crossover study was performed at the Kudanzaka Hospital, Chiyoda Ward, Tokyo. A total of 325 patients who had falls and 1295 controls matched by sex and age were included. The inclusion criteria for the case group were hospitalized patients who had their first fall and that for the control were patients who were hospitalized and did not have a fall, between January 2016 and November 2018. The internal sleep medications administered were classified as suvorexant, ramelteon, non-benzodiazepines, benzodiazepines, or kampo. In the case-control study, age, sex, clinical department, the fall down risk score, and hospitalized duration were adjusted in the logistic regression model. In the case-control study, multivariable logistic regression showed that the use of suvorexant (odds ratio [OR]: 2.61, 95% confidence interval [CI]: 1.29-5.28), nonbenzodiazepines (OR: 2.49, 95% CI: 1.73-3.59), and benzodiazepines (OR: 1.65, 95% CI: 1.16-2.34) was significantly associated with an increased OR of falls. However, the use of ramelteon (OR: 1.40, 95% CI: 0.60-3.16) and kampo (OR: 1.55, 95% CI: 0.75-3.19) was not significantly associated with an increased OR of falls. In the case-crossover study, the use of suvorexant (OR: 1.78, 95% CI: 1.05-3.00) and nonbenzodiazepines (OR: 1.63, 95% CI: 1.17-2.27) was significantly associated with an increased OR of falls. Similar patterns were observed in several sensitivity analyses. It was suggested that suvorexant increases the OR of falls. This result is robust in various analyses. This study showed that the risk of falls also exists for non-GABA sleep medication, suvorexant, and thus it is necessary to carefully prescribe hypnotic drugs under appropriate assessment., Competing Interests: Rie Nishitani, Ayano Takeuchi, Mamoru Touko, Takashi Kato, Satoshi Ohtani, Sahoko Chiba, Keiko Ashidate, Tomoyasu Ichijo, Takanori Shirai, Nobuo Ishiwata, Masataka Sasabe. declares that they have no conflicts of interest. Yoshiki Ishibashi, reports personal fees from Children and Future co., Ltd. Akiyoshi Shimura reports personal fees from Dainippon Sumitomo Pharma, Yoshitomiyakuhin, and Meiji Seika Pharma. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

Polar auxin transport is involved in multiple aspects of plant development, including root growth, lateral root branching, embryogenesis, and vasculature development. PIN-FORMED (PIN) auxin efflux proteins exhibit asymmetric distribution at the plasma membrane (PM) and collectively play pivotal roles in generating local auxin accumulation, which underlies various auxin-dependent developmental processes. In previous research, it has been revealed that endosomal trafficking components BEN1/BIG5 (ARF GEF) and BEN2/VPS45 (Sec1/Munc 18 protein) function in intracellular trafficking of PIN proteins in Arabidopsis. Mutations in both BEN1 and BEN2 resulted in defects in polar PIN localization, auxin response gradients, and in root architecture. In this study, we have attempted to gain insight into the developmental roles of these trafficking components. We showed that while genetic or pharmacological disturbances of auxin distribution reduced dividing cells in the root tips and resulted in reduced root growth, the same manipulations had only moderate impact on ben1; ben2 double mutants. In addition, we established transgenic lines in which BEN2/VPS45 is expressed under control of tissue-specific promoters and demonstrated that BEN2/VPS45 regulates the intracellular traffic of PIN proteins in cell-autonomous manner, at least in stele and epidermal cells. Furthermore, BEN2/VPS45 rescued the root architecture defects when expressed in internal tissues of ben1; ben2 double mutants. These results corroborate the roles of the endosomal trafficking component BEN2/VPS45 in regulation of auxin-dependent developmental processes, and suggest that BEN2/VPS45 is required for sustainable root growth, most likely through regulation of tip-ward auxin transport through the internal tissues of root., (Copyright © 2020 Matsuura, Fukasawa, Ogita, Sasabe, Kakimoto and Tanaka.)

Due to the rapid penetration of the Internet of Things (IoT) into human life, illegal access to IoT resources (e.g., data and actuators) has greatly threatened our safety. Access control, which specifies who (i.e., subjects) can access what resources (i.e., objects) under what conditions, has been recognized as an effective solution to address this issue. To cope with the distributed and trust-less nature of IoT systems, we propose a decentralized and trustworthy Capability-Based Access Control (CapBAC) scheme by using the Ethereum smart contract technology. In this scheme, a smart contract is created for each object to store and manage the capability tokens (i.e., data structures recording granted access rights) assigned to the related subjects, and also to verify the ownership and validity of the tokens for access control. Different from previous schemes which manage the tokens in units of subjects, i.e., one token per subject, our scheme manages the tokens in units of access rights or actions, i.e., one token per action. Such novel management achieves more fine-grained and flexible capability delegation and also ensures the consistency between the delegation information and the information stored in the tokens. We implemented the proposed CapBAC scheme in a locally constructed Ethereum blockchain network to demonstrate its feasibility. In addition, we measured the monetary cost of our scheme in terms of gas consumption to compare our scheme with the existing Blockchain-Enabled Decentralized Capability-Based Access Control (BlendCAC) scheme proposed by other researchers. The experimental results show that the proposed scheme outperforms the BlendCAC scheme in terms of the flexibility, granularity, and consistency of capability delegation at almost the same monetary cost.

In Arabidopsis, the ASYMMETRIC LEAVES2 (AS2) protein plays a key role in the formation of flat symmetric leaves via direct repression of the abaxial gene ETT/ARF3. AS2 encodes a plant-specific nuclear protein that contains the AS2/LOB domain, which includes a zinc-finger (ZF) motif that is conserved in the AS2/LOB family. We have shown that AS2 binds to the coding DNA of ETT/ARF3, which requires the ZF motif. AS2 is co-localized with AS1 in perinucleolar bodies (AS2 bodies). To identify the amino acid signals in AS2 required for formation of AS2 bodies and function(s) in leaf formation, we constructed recombinant DNAs that encoded mutant AS2 proteins fused to yellow fluorescent protein. We examined the subcellular localization of these proteins in cells of cotyledons and leaf primordia of transgenic plants and cultured cells. The amino acid signals essential for formation of AS2 bodies were located within and adjacent to the ZF motif. Mutant AS2 that failed to form AS2 bodies also failed to rescue the as2-1 mutation. Our results suggest the importance of the formation of AS2 bodies and the nature of interactions of AS2 with its target DNA and nucleolar factors including NUCLEOLIN1. The partial overlap of AS2 bodies with perinucleolar chromocenters with condensed ribosomal RNA genes implies a correlation between AS2 bodies and the chromatin state. Patterns of AS2 bodies in cells during interphase and mitosis in leaf primordia were distinct from those in cultured cells, suggesting that the formation and distribution of AS2 bodies are developmentally modulated in plants., (© 2019 The Authors. The Plant Journal published by Society for Experimental Biology and John Wiley & Sons Ltd.)

Plant growth and development relies on the accurate positioning of the cell plate between dividing cells during cytokinesis. The cell plate is synthetized by a specialized structure called the phragmoplast, which contains bipolar microtubules that polymerize to form a framework with the plus ends at or near the division site. This allows the transport of Golgi-derived vesicles toward the plus ends to form and expand the cell plate. Actin filaments play important roles in cell plate expansion and guidance in plant cytokinesis at the late phase, but whether they are involved at the early phase is unknown. To investigate this further, we disrupted the actin filaments in cell cycle-synchronized tobacco BY-2 cells with latrunculin B (LatB), an actin polymerization inhibitor. We observed the cells under a transmission electron microscope or a spinning-disk confocal laser scanning microscope. We found that disruption of actin filaments by LatB caused the membrane vesicles at the equatorial plane of the cell plate to be dispersed rather than form clusters as they did in the untreated cells. The midzone constriction of phragmoplast microtubules also was perturbed in LatB-treated cells. The live cell imaging and kymograph analysis showed that disruption of actin filaments also changed the accumulation timing of NACK1 kinesin, which plays a crucial role in cell plate expansion. This suggests that there are two functionally different types of microtubules in the phragmoplast. Together, our results show that actin filaments regulate phragmoplast microtubules at the initial phase of plant cytokinesis., (� The Author(s) 2020. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

The diversity of genital morphology among closely related animals with internal fertilization is well known, but the genetic backgrounds are unclear. Here, we show that, in Carabus ( Ohomopterus ) beetles showing correlated evolution of male and female genital parts, only a few major quantitative trait loci (QTLs) determine differences in genital dimensions between sister species, and sequence divergence is pronounced in the genomic regions containing genital QTLs. The major QTLs for male and female genital dimensions reside in different locations within the same linkage group, implying that coevolution between the sexes is only loosely constrained and can respond to sexually antagonistic selection. The same genomic regions containing the major QTLs show elevated divergence between three pairs of parapatric species with marked differences in genital parts. Our study demonstrates that species diversification can follow coevolution of genitalia between the sexes, even without tight linkage of loci affecting male and female genital dimensions.

TRYPTICHON (TRY) and ENHANCER OF TRY AND CPC2 (ETC2) encode R3-type MYB transcription factors that are involved in epidermal cell differentiation in Arabidopsis thaliana. TRY and ETC2 belong to the CPC-like MYB gene family, which includes seven homolog genes. Previously, we showed that among the CPC family members, TRY and ETC2 are characterized by rapid proteolysis compared with that of other members, and we demonstrated that this proteolysis is mediated by the proteasome-dependent pathway. In this study, we compared the functions of the wild-type TRY and ETC2 proteins and their amino acid-substituted versions. Our results showed that the substitution of amino acids in the C-terminal of TRY and ETC2 conferred them the ability to induce root hair formation. Furthermore, we confirmed that these mutations enhanced the stability of the TRY and ETC2 proteins. These results revealed that the amino acids, which are important for the functions of TRY and ETC2, mediate morphological pattern formation and can be useful in understanding the pathway determining the fate of root hair cells., Competing Interests: The authors have declared that no competing interests exist.

Arabidopsis ASYMMETRIC LEAVES2 (AS2) plays a critical role in leaf adaxial-abaxial partitioning by repressing expression of the abaxial-determining gene ETTIN/AUXIN RESPONSE FACTOR3 (ETT/ARF3). We previously reported that six CpG dinucleotides in its exon 6 are thoroughly methylated by METHYLTRASFERASE1, that CpG methylation levels are inversely correlated with ETT/ARF3 transcript levels and that methylation levels at three out of the six CpG dinucleotides are decreased in as2-1. All these imply that AS2 is involved in epigenetic repression of ETT/ARF3 by gene body DNA methylation. The mechanism of the epigenetic repression by AS2, however, is unknown. Here, we tested mutations of NUCLEOLIN1 (NUC1) and RNA HELICASE10 (RH10) encoding nucleolus-localized proteins for the methylation in exon 6 as these mutations enhance the level of ETT/ARF3 transcripts in as2-1. Methylation levels at three specific CpGs were decreased in rh10-1, and two of those three overlapped with those in as2-1. Methylation levels at two specific CpGs were decreased in nuc1-1, and one of those three overlapped with that in as2-1. No site was affected by both rh10-1 and nuc1-1. One specific CpG was unaffected by these mutations. These results imply that the way in which RH10, NUC1 and AS2 are involved in maintaining methylation at five CpGs in exon 6 might be through at least several independent pathways, which might interact with each other. Furthermore, we found that AS2 binds specifically the sequence containing CpGs in exon 1 of ETT/ARF3, and that the binding requires the zinc-finger-like motif in AS2 that is structurally similar to the zinc finger-CxxC domain in vertebrate DNA methyltransferase1.

The evolutionarily conserved MAP65 family proteins bundle anti-parallel microtubules (MTs). In Arabidopsis thaliana, mutations in the MAP65-3 gene lead to serious defects in MT organization in the phragmoplast and cause failures in cytokinesis. However, the functions of other ArabidopsisMAP65 isoforms are largely unknown. MAP65 functions were analyzed based on genetic interactions among different map65 mutations. Live-cell imaging and immunolocalization experiments revealed dynamic activities of two closely related MAP65 proteins in dividing cells. The map65-4 mutation caused synthetic lethality with map65-3 although map65-4 alone did not cause a noticeable phenotype. Furthermore, the introduction of an extra copy of the MAP65-4 gene significantly suppressed defects in cytokinesis and seedling growth caused by map65-3 because of restoring MT engagement in the spindle midzone. During mitosis, MAP65-4 first appeared at the preprophase band and persisted at the cortical division site afterwards. It was also concentrated on MTs in the spindle midzone and the phragmoplast. In the absence of MAP65-3, MAP65-4 exhibited greatly enhanced localization in the midzone of developing phragmoplast. Therefore, we have uncovered redundant but differential contributions of MAP65-3 and MAP65-4 to engaging and bundling anti-parallel MTs in the phragmoplast and disclosed a novel action of MAP65-4 at the cortical cell division site., (© 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.)

The phosphorylation of proteins by protein kinases controls many cellular and physiological processes, which include intracellular signal transduction. However, the underlying molecular mechanisms of such controls and numerous substrates of protein kinases remain to be characterized. The mitogen-activated protein kinase (MAPK) cascade is of particular importance in a variety of extracellular and intracellular signaling processes. In plant cells, the progression of cytokinesis is an excellent example of an intracellular phenomenon that requires the MAPK cascade. However, the way in which MAPKs control downstream processes during cytokinesis in plant cells remains to be fully determined. We show here that comparisons, by two-dimensional difference gel electrophoresis, of phosphorylated proteins from wild-type Arabidopsis thaliana and mutant plants defective in a MAPK cascade allow identification of substrates of a specific MAPK. Using this method, we identified the PATELLIN2 (PATL2) protein, which has a SEC14 domain, as a substrate of MPK4 MAP kinase. PATL2 was concentrated at the cell division plane, as is MPK4, and had binding affinity for phosphoinositides. This binding affinity was altered after phosphorylation of PATL2 by MPK4, suggesting a role for the MAPK cascade in the formation of cell plates via regeneration of membranes during cytokinesis., (© The Author 2016. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists.)

Plant cytokinesis is achieved by formation of cell plates in the phragmoplast, a plant-specific cytokinetic apparatus, which consists of microtubules (MTs) and microfilaments. During cytokinesis, the cell plate is expanded centrifugally outward from the inside of cells in a process that is supported by dynamic turnover of MTs. M-phase-specific kinesin NACK1, which comprises the motor domain at the amino-terminal half to move on MT bundles and the stalk region in the carboxyl-terminal half, is a key player in the process of MT turnover. That is, the specific region in the stalk binds the MAP kinase kinase kinase to activate the whole MAP kinase cascade, which stimulates depolymerization of MTs for the MT turnover. The stalk is also responsible for recruiting the activated kinase cascade to the mid-zone of the phragmoplast, which corresponds to the cell-plate formation site. It should be crucial to uncover roles of the NACK1 kinesin stalk as well as the motor domain in the formation of cell plates in order to understand the mechanisms of cell plate formation. Using dissected Arabidopsis NACK1 (AtNACK1/HINKEL) molecules and AtNACK1-fused GFP, we showed that the C-terminal tail of the stalk in addition to the motor domain is critical for its proper localization to the site of cell plate formation in the phragmoplast, probably by affecting its motility activity.

Appropriate timing of flowering is critical for reproductive success and necessarily involves complex genetic regulatory networks. A mobile floral signal, called florigen, is a key molecule in this process, and flowering locus T (FT) protein is its major component in Arabidopsis. FT is produced in leaves, but promotes the floral transition in the shoot apex, where it forms a complex with a basic region/leucine-zipper (bZIP) transcription factor, FD. Formation of the florigen complex depends on the supposed phosphorylation of FD; hitherto, however, the responsible protein kinase(s) have not been identified. In this study, we prepared protein extracts from shoot apices of plants around the floral transition, and detected a protein kinase activity that phosphorylates a threonine residue at position 282 of FD (FD T282), which is a crucial residue for the complex formation with FT via 14-3-3. The kinase activity was calcium-dependent. Subsequent biochemical, cellular, and genetic analyses showed that three calcium-dependent protein kinases (CDPKs) efficiently phosphorylate FD T282. Two of them (CPK6 and CPK33) are expressed in shoot apical meristem and directly interact with FD, suggesting they have redundant functions. The loss of function of one CDPK (CPK33) resulted in a weak but significant late-flowering phenotype.