Katanin-Dependent Microtubule Ordering in Association with ABA Is Important for Root Hydrotropism

Root hydrotropism refers to root directional growth toward soil moisture. Cortical microtubule arrays are essential for determining the growth axis of the elongating cells in plants. However, the role of microtubule reorganization in root hydrotropism remains elusive. Here, we demonstrate that the well-ordered microtubule arrays and the microtubule-severing protein KATANIN (KTN) play important roles in regulating root hydrotropism in Arabidopsis. We found that the root hydrotropic bending of the ktn1 mutant was severely attenuated but not root gravitropism. After hydrostimulation, cortical microtubule arrays in cells of the elongation zone of wild-type (WT) Col-0 roots were reoriented from transverse into an oblique array along the axis of cell elongation, whereas the microtubule arrays in the ktn1 mutant remained in disorder. Moreover, we revealed that abscisic acid (ABA) signaling enhanced the root hydrotropism of WT and partially rescued the oryzalin (a microtubule destabilizer) alterative root hydrotropism of WT but not ktn1 mutants. These results suggest that katanin-dependent microtubule ordering is required for root hydrotropism, which might work downstream of ABA signaling pathways for plant roots to search for water

Aneuploid Embryonic Stem Cells Drive Teratoma Metastasis

Aneuploidy, a deviation of the chromosome number from euploidy, is one of the hallmarks of cancer. High levels of aneuploidy are generally correlated with metastasis and poor prognosis in cancer patients. However, the causality of aneuploidy in cancer metastasis remains to be explored. Here we demonstrate that teratomas derived from aneuploid murine embryonic stem cells (ESCs), but not from isogenic diploid ESCs, disseminated to multiple organs, for which no additional copy number variations were required. Notably, no cancer driver gene mutations were identified in any metastases. Aneuploid circulating teratoma cells were successfully isolated from peripheral blood and showed high capacities for migration and organ colonization. Single-cell RNA sequencing of aneuploid primary teratomas and metastases identified a unique cell population with high stemness that was absent in diploid ESCs-derived teratomas. Further investigation revealed that aneuploid cells displayed decreased proteasome activity and overactivated endoplasmic reticulum (ER) stress during differentiation, thereby restricting the degradation of proteins produced from extra chromosomes in the ESC state and causing differentiation deficiencies. Noticeably, both proteasome activator Oleuropein and ER stress inhibitor 4-PBA can effectively inhibit aneuploid teratoma metastasis

A ROP GTPase-Dependent Auxin Signaling Pathway Regulates the Subcellular Distribution of PIN2 in Arabidopsis Roots

SummaryPIN-FORMED (PIN) protein-mediated auxin polar transport is critically important for development, pattern formation, and morphogenesis in plants. Auxin has been implicated in the regulation of polar auxin transport by inhibiting PIN endocytosis [1, 2], but how auxin regulates this process is poorly understood. Our genetic screen identified the Arabidopsis SPIKE1 (SPK1) gene whose loss-of-function mutations increased lateral root density and retarded gravitropic responses, as do pin2 knockout mutations [3]. SPK1 belongs to the conserved DHR2-Dock family of Rho guanine nucleotide exchange factors [4–6]. The spk1 mutations induced PIN2 internalization that was not suppressed by auxin, as did the loss-of-function mutations for Rho-like GTPase from Plants 6 (ROP6)-GTPase or its effector RIC1. Furthermore, SPK1 was required for auxin induction of ROP6 activation. Our results have established a Rho GTPase-based auxin signaling pathway that maintains PIN2 polar distribution to the plasma membrane via inhibition of its internalization in Arabidopsis roots. Our findings provide new insights into signaling mechanisms that underlie the regulation of the dynamic trafficking of PINs required for long-distance auxin transport and that link auxin signaling to PIN-mediated pattern formation and morphogenesis

Ischemia promotes hypertrophic nerve trunk formation and enteric neuron cell death in Hirschsprung's disease: Source data

<table> <tbody> <tr> <td>Items</td> <td>Annotation</td> </tr> <tr> <td>ST-cluster_distrition.csv</td> <td>A table containing the number of ST-clusters across samples</td> </tr> <tr> <td>DEGs_C10_v_restCMusc.csv</td> <td>A table containing the result of DEG analysis for the comparison between ST cluster C10 and other muscular ST clusters (C8 and C9)</td> </tr> <tr> <td>DEGs_C13_v_C12.csv</td> <td>A table containing the result of DEG analysis for the comparison between ST cluster C13 (HNTs) and C12 (ganglia).</td> </tr> <tr> <td>DEGs_ST-cluster_total_marker.csv</td> <td>A table containing the markers for each ST cluster.</td> </tr> <tr> <td>DEGs_iMusc_ganglia_v_nMusc_ganglia.csv</td> <td>A table containing the result of DEG analysis for the comparison between ST iMusc_ganglia and nMusc_ganglia.</td> </tr> <tr> <td>DEGs_sc_all_markers.csv</td> <td>A table containing the markers of single-cell populations.</td> </tr> <tr> <td>DEGs_pseudotime_CLDN1.csv</td> <td>A table containing the results of gene differentially expressed during the differentiation from GREM1+ FLCs to CLDN1+ FLCs.</td> </tr> <tr> <td>Gene_set_score_for_C12_sub.csv</td> <td>A table containing the results of gene set score in the sub-population of ganglia (C12).</td> </tr> <tr> <td>STM_WES_FASTQ_meta_data.xlsx</td> <td>A table containing the ST-seq, scRNA-seq, and WES FASTQ file metadata.</td> </tr> <tr> <td>scRNA-seq_reference_raw.RDS</td> <td>A Seurat object containing the raw count and meta matrix of the single-cell reference </td> </tr> <tr> <td>ST-seq_raw_count.txt.gz</td> <td>A raw count matrix of ST-seq data</td> </tr> <tr> <td>ST-spots_meta_info.txt</td> <td>A table containing the metainformation of ST-seq data</td> </tr> <tr> <td>Object_of_bulk_RNA-seq.RDS</td> <td>A Seurat object used to reproduce the results of gene set scores in bulk RNA-seq.</td> </tr> <tr> <td>ST-spots_cell_abundance.txt</td> <td>A table containing the abundance of single-cell populations in each ST-spot.</td> </tr> <tr> <td>Cell_trajectory_of_FLCs.RDS</td> <td>A Seurat object used to reproduce the results of the cell trajectory of fibroblast-like cells.</td> </tr> <tr> <td>p_value_scRNA-seq_abundance.xlsx</td> <td>A table containing the p-value of cell abundance across groups for single-cell populations</td> </tr> <tr> <td>CIBERSORT_CPM_count_matrix.txt</td> <td>A table containing the CMP matrix of bulk RNA-seq data for CIBERSORT</td> </tr> <tr> <td>CIBERSORT_ST_raw_matrix.txt</td> <td>A table containing the raw count matrix of ST-seq data for CIBERSORT</td> </tr> <tr> <td>scRNA-seq_distribution.csv</td> <td>A table containing the distribution of single-cell populations across the sample</td> </tr> <tr> <td>DEGs_FLC_APOD_across_group.csv</td> <td>A table containing the DEGs in the comparison among APOD+ FLCs in different groups</td> </tr> <tr> <td>Gene_set_score_for_sc_Pericyte.csv</td> <td>A table containing the results of gene set score in the single-cell population of pericytes</td> </tr> <tr> <td>Gene_set_score_for_sc_Endo_score.csv</td> <td>A table containing the results of gene set score in the single-cell population of endothelial cells</td> </tr> <tr> <td>ENC_FLC_vessel_interaction.RDS</td> <td>A Cellchat object used to reproduce the results of cell-cell interaction among ENCs, glial cells, APOD+ FLCs, CLDN1+ FLCs, perictyes, and vascular endothelial cells.</td> </tr> </tbody> </table> <p> </p> <p> </p&gt

Ischemia promotes hypertrophic nerve trunk formation and enteric neuron cell death in Hirschsprung's disease: Source data

<table> <tbody> <tr> <td>Items</td> <td>Annotation</td> </tr> <tr> <td>ST-cluster_distrition.csv</td> <td>A table containing the number of ST-clusters across samples</td> </tr> <tr> <td>DEGs_C10_v_restCMusc.csv</td> <td>A table containing the result of DEG analysis for the comparison between ST cluster C10 and other muscular ST clusters (C8 and C9)</td> </tr> <tr> <td>DEGs_C13_v_C12.csv</td> <td>A table containing the result of DEG analysis for the comparison between ST cluster C13 (HNTs) and C12 (ganglia).</td> </tr> <tr> <td>DEGs_ST-cluster_total_marker.csv</td> <td>A table containing the markers for each ST cluster.</td> </tr> <tr> <td>DEGs_iMusc_ganglia_v_nMusc_ganglia.csv</td> <td>A table containing the result of DEG analysis for the comparison between ST iMusc_ganglia and nMusc_ganglia.</td> </tr> <tr> <td>DEGs_sc_all_markers.csv</td> <td>A table containing the markers of single-cell populations.</td> </tr> <tr> <td>DEGs_pseudotime_CLDN1.csv</td> <td>A table containing the results of gene differentially expressed during the differentiation from GREM1+ FLCs to CLDN1+ FLCs.</td> </tr> <tr> <td>Gene_set_score_for_C12_sub.csv</td> <td>A table containing the results of gene set score in the sub-population of ganglia (C12).</td> </tr> <tr> <td>STM_WES_FASTQ_meta_data.xlsx</td> <td>A table containing the ST-seq, scRNA-seq, and WES FASTQ file metadata.</td> </tr> <tr> <td>scRNA-seq_reference_raw.RDS</td> <td>A Seurat object containing the raw count and meta matrix of the single-cell reference </td> </tr> <tr> <td>ST-seq_raw_count.txt.gz</td> <td>A raw count matrix of ST-seq data</td> </tr> <tr> <td>ST-spots_meta_info.txt</td> <td>A table containing the metainformation of ST-seq data</td> </tr> <tr> <td>Object_of_bulk_RNA-seq.RDS</td> <td>A Seurat object used to reproduce the results of gene set scores in bulk RNA-seq.</td> </tr> <tr> <td>ST-spots_cell_abundance.txt</td> <td>A table containing the abundance of single-cell populations in each ST-spot.</td> </tr> <tr> <td>Cell_trajectory_of_FLCs.RDS</td> <td>A Seurat object used to reproduce the results of the cell trajectory of fibroblast-like cells.</td> </tr> <tr> <td>p_value_scRNA-seq_abundance.xlsx</td> <td>A table containing the p-value of cell abundance across groups for single-cell populations</td> </tr> <tr> <td>CIBERSORT_CPM_count_matrix.txt</td> <td>A table containing the CMP matrix of bulk RNA-seq data for CIBERSORT</td> </tr> <tr> <td>CIBERSORT_ST_raw_matrix.txt</td> <td>A table containing the raw count matrix of ST-seq data for CIBERSORT</td> </tr> <tr> <td>scRNA-seq_distribution.csv</td> <td>A table containing the distribution of single-cell populations across the sample</td> </tr> <tr> <td>DEGs_FLC_APOD_across_group.csv</td> <td>A table containing the DEGs in the comparison among APOD+ FLCs in different groups</td> </tr> <tr> <td>Gene_set_score_for_sc_Pericyte.csv</td> <td>A table containing the results of gene set score in the single-cell population of pericytes</td> </tr> <tr> <td>Gene_set_score_for_sc_Endo_score.csv</td> <td>A table containing the results of gene set score in the single-cell population of endothelial cells</td> </tr> <tr> <td>ENC_FLC_vessel_interaction.RDS</td> <td>A Cellchat object used to reproduce the results of cell-cell interaction among ENCs, glial cells, APOD+ FLCs, CLDN1+ FLCs, perictyes, and vascular endothelial cells.</td> </tr> <tr> <td>Metainfor_FASTQ1.xlsx</td> <td>Metainformation of FASTQ files</td> </tr> <tr> <td>Metainfor_FASTQ2.xlsx</td> <td>Metainformation of FASTQ files</td> </tr> </tbody> </table> <p> </p> <p> </p&gt

ABP1 and ROP6 GTPase signaling regulate clathrin-mediated endocytosis in Arabidopsis roots

The dynamic spatial and temporal distribution of the crucial plant signaling molecule auxin is achieved by feedback coordination of auxin signaling and intercellular auxin transport pathways [1, 2]. Developmental roles of auxin have been attributed predominantly to its effect on transcription; however, an alternative pathway involving AUXIN BINDING PROTEIN1 (ABP1) has been proposed to regulate clathrin-mediated endocytosis in roots and Rho-like GTPase (ROP)-dependent pavement cell interdigitation in leaves [3, 4]. In this study, we show that ROP6 and its downstream effector RIC1 regulate clathrin association with the plasma membrane for clathrin-mediated endocytosis, as well as for its feedback regulation by auxin. Genetic analysis revealed that ROP6/RIC1 acts downstream of ABP1 to regulate endocytosis. This signaling circuit is also involved in the feedback regulation of PIN-FORMED 1 (PIN1) and PIN2 auxin transporters activity (via its constitutive endocytosis) and corresponding auxin transport-mediated processes, including root gravitropism and leave vascular tissue patterning. Our findings suggest that the signaling module auxin ABP1 ROP6/RIC1 clathrin PIN1/PIN2 is a shared component of the feedback regulation of auxin transport during both root and aerial development

ROP GTPase-Dependent Actin Microfilaments Promote PIN1 Polarization by Localized Inhibition of Clathrin-Dependent Endocytosis

Cell polarization via asymmetrical distribution of structures or molecules is essential for diverse cellular functions and development of organisms, but how polarity is developmentally controlled has been poorly understood. In plants, the asymmetrical distribution of the PIN-FORMED (PIN) proteins involved in the cellular efflux of the quintessential phytohormone auxin plays a central role in developmental patterning, morphogenesis, and differential growth. Recently we showed that auxin promotes cell interdigitation by activating the Rho family ROP GTPases in leaf epidermal pavement cells. Here we found that auxin activation of the ROP2 signaling pathway regulates the asymmetric distribution of PIN1 by inhibiting its endocytosis. ROP2 inhibits PIN1 endocytosis via the accumulation of cortical actin microfilaments induced by the ROP2 effector protein RIC4. Our findings suggest a link between the developmental auxin signal and polar PIN1 distribution via Rho-dependent cytoskeletal reorganization and reveal the conservation of a design principle for cell polarization that is based on Rho GTPase-mediated inhibition of endocytosis

Single-Cell Real-Time Visualization and Quantification of Perylene Bioaccumulation in Microorganisms

Bioaccumulation of perylene in <i>Escherichia coli</i> and <i>Staphylococcus aureus</i> was visualized and quantified in real time with high sensitivity at high temporal resolution. For the first time, single-molecule fluorescence microscopy (SMFM) with a microfluidic flow chamber and temperature control has enabled us to record the dynamic process of perylene bioaccumulation in single bacterial cells and examine the cell-to-cell heterogeneity. Although with identical genomes, individual <i>E. coli</i> cells exhibited a high degree of heterogeneity in perylene accumulation dynamics, as shown by the high coefficient of variation (C.V = 1.40). This remarkable heterogeneity was exhibited only in live <i>E. coli</i> cells. However, the bioaccumulation of perylene in live and dead <i>S. aureus</i> cells showed similar patterns with a low degree of heterogeneity (C.V = 0.36). We found that the efflux systems associated with Tol C played an essential role in perylene bioaccumulation in <i>E. coli</i>, which caused a significantly lower accumulation and a high cell-to-cell heterogeneity. In comparison with <i>E. coli</i>, the Gram-positive bacteria <i>S. aureus</i> lacked an efficient efflux system against perylene. Therefore, perylene bioaccumulation in <i>S. aureus</i> was simply a passive diffusion process across the cell membrane