Rab6-Mediated Polarized Transport of Synaptic Vesicle Precursors Is Essential for the Establishment of Neuronal Polarity and Brain Formation.

Neurons are highly polarized cells that are composed of a single axon and multiple dendrites. Axon-dendrite polarity is essential for proper tissue formation and brain functions. Intracellular protein transport plays an important role in the establishment of neuronal polarity. However, the regulatory mechanism of polarized transport remains unclear. Here, we show that Rab6, a small GTPase that acts on the regulation of intracellular vesicular trafficking, plays key roles in neuronal polarization and brain development. Central nervous system-specific Rab6a/b double knock-out (Rab6 DKO) mice of both sexes exhibit severe dysplasia of the neocortex and the cerebellum. In the Rab6 DKO neocortex, impaired axonal extension of neurons results in hypoplasia of the intermediate zone. In vitro, deletion of Rab6a and Rab6b in cultured neurons from both sexes causes the abnormal accumulation of synaptic vesicle precursors (SVPs) adjacent to the Golgi apparatus, which leads to defects in axonal extension and the loss of axon-dendrite polarity. Moreover, Rab6 DKO causes significant expansion of lysosomes in the soma in neurons. Overall, our results reveal that Rab6-mediated polarized transport of SVPs is crucial for neuronal polarization and subsequent brain formation.

Dynamic movement of the Golgi unit and its glycosylation enzyme zones.

Knowledge on the distribution and dynamics of glycosylation enzymes in the Golgi is essential for better understanding this modification. Here, using a combination of CRISPR/Cas9 knockin technology and super-resolution microscopy, we show that the Golgi complex is assembled by a number of small 'Golgi units' that have 1-3 µm in diameter. Each Golgi unit contains small domains of glycosylation enzymes which we call 'zones'. The zones of N- and O-glycosylation enzymes are colocalised. However, they are less colocalised with the zones of a glycosaminoglycan synthesizing enzyme. Golgi units change shapes dynamically and the zones of glycosylation enzymes rapidly move near the rim of the unit. Photobleaching analysis indicates that a glycosaminoglycan synthesizing enzyme moves between units. Depletion of giantin dissociates units and prevents the movement of glycosaminoglycan synthesizing enzymes, which leads to insufficient glycosaminoglycan synthesis. Thus, we show the structure-function relationship of the Golgi and its implications in human pathogenesis.

EHBP1L1, an apicobasal polarity regulator, is critical for nuclear polarization during enucleation of erythroblasts.

Cell polarity, the asymmetric distribution of proteins and organelles, is permanently or transiently established in various cell types and plays an important role in many physiological events. epidermal growth factor receptor substrate 15 homology domain-binding protein 1-like 1 (EHBP1L1) is an adapter protein that is localized on recycling endosomes and regulates apical-directed transport in polarized epithelial cells. However, the role of EHBP1L1 in nonepithelial cells, remains unknown. Here, Ehbp1l1-/- mice showed impaired erythroblast enucleation. Further analyses showed that nuclear polarization before enucleation was impaired in Ehbp1l1-/- erythroblasts. It was also revealed that EHBP1L1 interactors Rab10, Bin1, and dynamin were involved in erythroblast enucleation. In addition, Ehbp1l1-/- erythrocytes exhibited stomatocytic morphology and dehydration. These defects in erythroid cells culminated in early postnatal anemic lethality in Ehbp1l1-/- mice. Moreover, we found the mislocalization of nuclei and mitochondria in the skeletal muscle cells of Ehbp1l1-/- mice, as observed in patients with centronuclear myopathy with genetic mutations in Bin1 or dynamin 2. Taken together, our findings indicate that the Rab8/10-EHBP1L1-Bin1-dynamin axis plays an important role in multiple cell polarity systems in epithelial and nonepithelial cells.

Fezf2-positive fork cell-like neurons in the mouse insular cortex.

The fork cell and von Economo neuron, which are found in the insular cortex and/or the anterior cingulate cortex, are defined by their unique morphologies. Their shapes are not pyramidal; the fork cell has two primary apical dendrites and the von Economo neurons are spindle-shaped (bipolar). Presence of such neurons are reported only in the higher animals, especially in human and great ape, indicating that they are specific for most evolved species. Although it is likely that these neurons are involved in higher brain function, lack of results with experimental animals makes further investigation difficult. We here ask whether equivalent neurons exist in the mouse insular cortex. In human, Fezf2 has been reported to be highly expressed in these morphologically distinctive neurons and thus, we examined the detailed morphology of Fezf2-positive neurons in the mouse brain. Although von Economo-like neurons were not identified, Fezf2-positive fork cell-like neurons with two characteristic apical dendrites, were discovered. Examination with electron microscope indicated that these neurons did not embrace capillaries, rather they held another cell. We here term such neurons as holding neurons. We further observed several molecules, including neuromedin B (NMB) and gastrin releasing peptide (GRP) that are known to be localized in the fork cells and/or von Economo cells in human, were localized in the mouse insular cortex. Based on these observations, it is likely that an equivalent of the fork cell is present in the mouse.

Cytoneme-like protrusion formation induced by LAR is promoted by receptor dimerization.

Actin-based protrusions called cytonemes are reported to function in cell communication by supporting events such as morphogen gradient establishment and pattern formation. Despite the crucial roles of cytonemes in cell signaling, the molecular mechanism for cytoneme establishment remains elusive. In this study, we showed that the leukocyte common antigen-related (LAR) receptor protein tyrosine phosphatase plays an important role in cytoneme-like protrusion formation. Overexpression of LAR in HEK293T cells induced the formation of actin-based protrusions, some of which exceeded 200 µm in length and displayed a complex morphology with branches. Upon focusing on the regulation of LAR dimerization or clustering and the resulting regulatory effects on LAR phosphatase activity, we found that longer and more branched protrusions were formed when LAR dimerization was artificially induced and when heparan sulfate was applied. Interestingly, although the truncated form of LAR lacking phosphatase-related domains promoted protrusion formation, the phosphatase-inactive forms did not show clear changes, suggesting that LAR dimerization triggers the formation of cytoneme-like protrusions in a phosphatase-independent manner. Our results thus emphasize the importance of LAR and its dimerization in cell signaling. This article has an associated First Person interview with the first author of the paper.

Transient expansion of the expression region of Hsd11b1, encoding 11ß-hydroxysteroid dehydrogenase type 1, in the developing mouse neocortex.

Corticosteroids are stress-related hormones that maintain homeostasis. The most effective corticosteroids are corticosterone (CORT) in rodents and cortisol in primates. 11ß-Hydroxysteroid dehydrogenase type 1 (11ß-HSD1; EC 1.1.1.146), encoded by Hsd11b1, is a key regulator of the local concentration of CORT/cortisol. Hsd11b1 expression in layer 5 of the primary somatosensory cortex has been shown in adult mice. However, its localization in the entire neocortex, especially during development, has not been fully addressed. Here, we established robust and dynamic expression profiles of Hsd11b1 in the developing mouse neocortex. Hsd11b1 was found mostly in pyramidal neurons. By retrograde tracing, we observed that some Hsd11b1-positive cells were projection neurons, indicating that at least some were excitatory. At postnatal day 0 (P0), Hsd11b1 was expressed in the deep layer of the somatosensory cortex. Then, from P3 to P8, the expression area expanded broadly; it was observed in layers 4 and 5, spanning the whole neocortex, including the primary motor cortex (M1) and the primary visual cortex (V1). The positive region gradually narrowed from P14 onwards and was ultimately limited to layer 5 of the somatosensory cortex at P26 and later. Furthermore, we administered CORT to nursing dams to increase the systemic CORT level of their pups. Here, we observed a reduced number of Hsd11b1-positive cells in the neocortex of these pups. Our observation suggests that Hsd11b1 expression in the developing neocortex is affected by systemic CORT levels. It is possible that stress on mothers influences the neocortical development of their children.

Interstitial Axon Collaterals of Callosal Neurons Form Association Projections from the Primary Somatosensory to Motor Cortex in Mice.

Association projections from cortical pyramidal neurons connect disparate intrahemispheric cortical areas, which are implicated in higher cortical functions. The underlying developmental processes of these association projections, especially the initial phase before reaching the target areas, remain unknown. To visualize developing axons of individual neurons with association projections in the mouse neocortex, we devised a sparse labeling method that combined in utero electroporation and confocal imaging of flattened and optically cleared cortices. Using the promoter of an established callosal neuron marker gene that was expressed in over 80% of L2/3 neurons in the primary somatosensory cortex (S1) that project to the primary motor cortex (M1), we found that an association projection of a single neuron was the longest among the interstitial collaterals that branched out in L5 from the earlier-extended callosal projection. Collaterals to M1 elongated primarily within the cortical gray matter with little branching before reaching the target. Our results suggest that dual-projection neurons in S1 make a significant fraction of the association projections to M1, supporting the directed guidance mechanism in long-range corticocortical circuit formation over random projections followed by specific pruning.

Mutually Repulsive EphA7-EfnA5 Organize Region-to-Region Corticopontine Projection by Inhibiting Collateral Extension.

Coordination of skilled movements and motor planning relies on the formation of regionally restricted brain circuits that connect cortex with subcortical areas during embryonic development. Layer 5 neurons that are distributed across most cortical areas innervate the pontine nuclei (basilar pons) by protrusion and extension of collateral branches interstitially along their corticospinal extending axons. Pons-derived chemotropic cues are known to attract extending axons, but molecules that regulate collateral extension to create regionally segregated targeting patterns have not been identified. Here, we discovered that EphA7 and EfnA5 are expressed in the cortex and the basilar pons in a region-specific and mutually exclusive manner, and that their repulsive activities are essential for segregating collateral extensions from corticospinal axonal tracts in mice. Specifically, EphA7 and EfnA5 forward and reverse inhibitory signals direct collateral extension such that EphA7-positive frontal and occipital cortical areas extend their axon collaterals into the EfnA5-negative rostral part of the basilar pons, whereas EfnA5-positive parietal cortical areas extend their collaterals into the EphA7-negative caudal part of the basilar pons. Together, our results provide a molecular basis that explains how the corticopontine projection connects multimodal cortical outputs to their subcortical targets.SIGNIFICANCE STATEMENT Our findings put forward a model in which region-to-region connections between cortex and subcortical areas are shaped by mutually exclusive molecules to ensure the fidelity of regionally restricted circuitry. This model is distinct from earlier work showing that neuronal circuits within individual cortical modalities form in a topographical manner controlled by a gradient of axon guidance molecules. The principle that a shared molecular program of mutually repulsive signaling instructs regional organization-both within each brain region and between connected brain regions-may well be applicable to other contexts in which information is sorted by converging and diverging neuronal circuits.

Loss of Rab6a in the small intestine causes lipid accumulation and epithelial cell death from lactation.

Intestinal epithelial cells (IECs) are not only responsible for the digestion and absorption of dietary substrates but also function as a first line of host defense against commensal and pathogenic luminal bacteria. Disruption of the epithelial layer causes malnutrition and enteritis. Rab6 is a small GTPase localized to the Golgi, where it regulates anterograde and retrograde transport by interacting with various effector proteins. Here, we generated mice with IEC-specific deletion of Rab6a (Rab6a∆IEC mice). While Rab6aΔIEC mice were born at the Mendelian ratio, they started to show IEC death, inflammation, and bleeding in the small intestine shortly after birth, and these changes culminated in early postnatal death. We further found massive lipid accumulation in the IECs of Rab6a∆IEC neonates. In contrast to Rab6a∆IEC neonates, knockout embryos did not show any of these abnormalities. Lipid accumulation and IEC death became evident when Rab6a∆IEC embryos were nursed by a foster mother, suggesting that dietary milk-derived lipids accumulated in Rab6a-deficient IECs and triggered IEC death. These results indicate that Rab6a plays a crucial role in regulating the lipid transport and maintaining tissue integrity.

Highly Deep Ultraviolet-Transparent h-BN Film Deposited on an Al0.7Ga0.3N Template by Low-Temperature Radio-Frequency Sputtering.

Hexagonal boron nitride (h-BN) is an attractive wide-bandgap material for application to emitters and detectors operating in the deep ultraviolet (DUV) spectral region. The optical transmittance of h-BN in the DUV region is particularly important for these devices. We report on the deposition of thick h-BN films (>200 nm) on Al0.7Ga0.3N templates via radio-frequency sputtering, along with the realization of ultrahigh transmittance in the DUV region. The fraction of the gas mixture (Ar/N2) was varied to investigate its effects on the optical transmittance of BN. DUV light transmittance of as high as 94% was achieved at 265 nm. This value could be further enhanced to exceed 98% by a post-annealing treatment at 800 °C in a N2 ambient for 20 min. The phase of the highly DUV-transparent BN film was determined to be a purely hexagonal structure via Raman spectra measurements. More importantly, these deposition processes were performed at a low temperature (300 °C), which can provide protection from device performance degradation when applied to actual devices.

Kcnab1 Is Expressed in Subplate Neurons With Unilateral Long-Range Inter-Areal Projections.

Subplate (SP) neurons are among the earliest-born neurons in the cerebral cortex and heterogeneous in terms of gene expression. SP neurons consist mainly of projection neurons, which begin to extend their axons to specific target areas very early during development. However, the relationships between axon projection and gene expression patterns of the SP neurons, and their remnant layer 6b (L6b) neurons, are largely unknown. In this study, we analyzed the corticocortical projections of L6b/SP neurons in the mouse cortex and searched for a marker gene expressed in L6b/SP neurons that have ipsilateral inter-areal projections. Retrograde tracing experiments demonstrated that L6b/SP neurons in the primary somatosensory cortex (S1) projected to the primary motor cortex (M1) within the same cortical hemisphere at postnatal day (PD) 2 but did not show any callosal projection. This unilateral projection pattern persisted into adulthood. Our microarray analysis identified the gene encoding a ß subunit of voltage-gated potassium channel (Kcnab1) as being expressed in L6b/SP. Double labeling with retrograde tracing and in situ hybridization demonstrated that Kcnab1 was expressed in the unilaterally-projecting neurons in L6b/SP. Embryonic expression was specifically detected in the SP as early as embryonic day (E) 14.5, shortly after the emergence of SP. Double immunostaining experiments revealed different degrees of co-expression of the protein product Kvß1 with L6b/SP markers Ctgf (88%), Cplx3 (79%), and Nurr1 (58%), suggesting molecular subdivision of unilaterally-projecting L6b/SP neurons. In addition to expression in L6b/SP, scattered expression of Kcnab1 was observed during postnatal stages without layer specificity. Among splicing variants with three alternative first exons, the variant 1.1 explained all the cortical expression mentioned in this study. Together, our data suggest that L6b/SP neurons have corticocortical projections and Kcnab1 expression defines a subpopulation of L6b/SP neurons with a unilateral inter-areal projection.

Fibroblast growth factor 12 is expressed in spiral and vestibular ganglia and necessary for auditory and equilibrium function.

We investigated fibroblast growth factor 12 (FGF12) as a transcript enriched in the inner ear by searching published cDNA library databases. FGF12 is a fibroblast growth factor homologous factor, a subset of the FGF superfamily. To date, its localisation and function in the inner ear have not been determined. Here, we show that FGF12 mRNA is localised in spiral ganglion neurons (SGNs) and the vestibular ganglion. We also show that FGF12 protein is localised in SGNs, the vestibular ganglion, and nerve fibres extending beneath hair cells. Moreover, we investigated FGF12 function in auditory and vestibular systems using Fgf12-knockout (FGF12-KO) mice generated with CRISPR/Cas9 technology. Our results show that the inner ear morphology of FGF12-KO mice is not significantly different compared with wild-type mice. However, FGF12-KO mice exhibited an increased hearing threshold, as measured by the auditory brainstem response, as well as deficits in rotarod and balance beam performance tests. These results suggest that FGF12 is necessary for normal auditory and equilibrium function.

Epiphycan is specifically expressed in cochlear supporting cells and is necessary for normal hearing.

The study of inner ear specific transcripts has revealed novel information about hereditary hearing loss and a mechanism of normal hearing. In this study, by analyzing a published cDNA library, we focused on Epiphycan (Epyc), a member of the small leucine-rich repeat proteoglycan family, whose transcript is enriched in the inner ear. Epyc mRNA was expressed abundantly and specifically in adult mice cochleae and was localized in supporting cells within the organ of Corti of both neonatal and adult mice. To examine the function of Epyc, we generated Epyc knockout (KO) mice using the CRISPR/Cas9 system. Epyc KO mice cochleae exhibited normal morphology. However, measurement of the auditory brain-stem response in Epyc KO mice revealed an elevated hearing threshold above 16 kHz frequency. This study suggests that Epyc is necessary for normal auditory function.

Echocardiographic Estimates of Left Ventricular Diastolic Dysfunction Do Not Predict the Clinical Course in Elderly Patients Undergoing Transcatheter Atrial Septal Defect Closure: Impact of Early Diastolic Mitral Annular Velocity.

OBJECTIVE: This study evaluated the effect of echocardiographic left ventricular (LV) diastolic dysfunction on acute congestive heart failure after transcatheter atrial septal defect (ASD) closure in elderly patients. BACKGROUND: Although there is concern that LV diastolic dysfunction develops acute congestive heart failure after ASD closure, limited information is available regarding the influence, especially in elderly patients with severe LV diastolic dysfunction. METHODS: Two hundred consecutive patients older than 60 years were divided into 3 groups according to echocardiographic LV diastolic dysfunction: severe (early diastolic mitral annular velocity [e'] <5.0 cm/s), mild (5.0≤ e' <8.0 cm/s), and normal (e' ≥ 8.0 cm/s). Changes in plasma B-type natriuretic peptide (BNP) levels were evaluated. RESULTS: No patients with severe LV diastolic dysfunction developed acute congestive heart failure immediately after the procedure. BNP levels unchanged after the procedure in patients with severe LV diastolic dysfunction (126 ± 181 to 131 ± 148 pg/ml, P = 0.885), and this increase in BNP levels was not different from that between the diagnosis of ASD and the procedure. The change in BNP levels in patients with severe LV diastolic dysfunction, who were frequently treated with diuretics before the procedure, was equivalent to that in patients with mild LV diastolic dysfunction and normal LV diastolic function (5 ± 119 vs. 16 ± 101 vs. 9 ± 131 pg/ml, P = 0.724). CONCLUSIONS: Our findings suggest that transcatheter ASD closure under volume management is safe and valuable in elderly patients with echocardiographic severe LV diastolic dysfunction.

Deficient Surrounding Rims in Patients Undergoing Transcatheter Atrial Septal Defect Closure.

BACKGROUND: The influence of deficient rims surrounding atrial septal defects (ASDs) in patients undergoing transcatheter closure has yet to be clarified. The aim of this study was to assess the influence of a deficient surrounding rim on the procedural success and clinical outcome of transcatheter ASD closure using an Amplatzer septal occluder. METHODS: A total of 474 patients (mean age, 46 ± 22 years) with ostium secundum ASDs measuring ≤40 mm in diameter who had undergone attempted transcatheter closure using Amplatzer septal occluders from September 2007 to August 2013 were assessed. A comprehensive transesophageal echocardiographic examination was done to assess the morphologic characteristics of the defects in all patients. Subjects were classified into three groups by the extent and location of rim deficiency (<5 mm): patients without deficient rims (sufficient group, n = 101), patients with single deficient rims, (single group, n = 338), and patients with multiple rim deficiencies (multiple group, n = 35). RESULTS: There was a significant difference in the maximal defect diameter among the sufficient, single, and multiple groups (15 ± 6, 18 ± 6, and 29 ± 7 mm, respectively, P < .001). Transcatheter closure was successfully accomplished in 463 patients (98%). The prevalence of procedural success differed significantly among the sufficient, single, and multiple groups (100%, 98%, and 86%, respectively, P < .001). There was no significant difference in the occurrence of cardiovascular events among the three groups during a mean follow-up period of 25 ± 19 months (P = .926, log-rank test). CONCLUSIONS: In patients with ASDs with multiple rim deficiencies as determined by transesophageal echocardiography, successful transcatheter ASD closure using Amplatzer septal occluders is more difficult to accomplish. However, if closure is successful, rim deficiencies rarely affect intermediate-term outcomes.

Association between chronic stress-induced structural abnormalities in Ranvier nodes and reduced oligodendrocyte activity in major depression.

Repeated stressful events are associated with the onset of major depressive disorder (MDD). We previously showed oligodendrocyte (OL)-specific activation of the serum/glucocorticoid-regulated kinase (SGK)1 cascade, increased expression of axon-myelin adhesion molecules, and elaboration of the oligodendrocytic arbor in the corpus callosum of chronically stressed mice. In the current study, we demonstrate that the nodes and paranodes of Ranvier in the corpus callosum were narrower in these mice. Chronic stress also led to diffuse redistribution of Caspr and Kv 1.1 and decreased the activity in white matter, suggesting a link between morphological changes in OLs and inhibition of axonal activity. OL primary cultures subjected to chronic stress resulted in SGK1 activation and translocation to the nucleus, where it inhibited the transcription of metabotropic glutamate receptors (mGluRs). Furthermore, the cAMP level and membrane potential of OLs were reduced by chronic stress exposure. We showed by diffusion tensor imaging that the corpus callosum of patients with MDD exhibited reduced fractional anisotropy, reflecting compromised white matter integrity possibly caused by axonal damage. Our findings suggest that chronic stress disrupts the organization of the nodes of Ranvier by suppressing mGluR activation in OLs, and that specific white matter abnormalities are closely associated with MDD onset.

Sgk1 regulates desmoglein 1 expression levels in oligodendrocytes in the mouse corpus callosum after chronic stress exposure.

Major depression, one of the most prevalent mental illnesses, is thought to be a multifactorial disease related to both genetic and environmental factors. However, the genes responsible for and the pathogenesis of major depression at the molecular level remain unclear. Recently, we reported that stressed mice with elevated plasma corticosterone levels show upregulation and activation of serum glucocorticoid-regulated kinase (Sgk1) in oligodendrocytes. Active Sgk1 causes phosphorylation of N-myc downstream-regulated gene 1 (Ndrg1), and phospho-Ndrg1 increases the expression of N-cadherin, α-catenin, and ß-catenin in oligodendrocytes. This activation of the Sgk1 cascade results in morphological changes in the oligodendrocytes of nerve fiber bundles, such as those present in the corpus callosum. However, little is known about the molecular functions of the traditional and/or desmosomal cadherin superfamily in oligodendrocytes. Therefore, in this study, we aimed to elucidate the functions of the desmosomal cadherin superfamily in oligodendrocytes. Desmoglein (Dsg) 1, Dsg2, and desmocollin 1 (Dsc1) were found to be expressed in the corpus callosum of mouse brain, and the expression of a subtype of Dsg1, Dsg1c, was upregulated in oligodendrocytes after chronic stress exposure. Furthermore, Dsg1 proteins were localized around the plasma membrane regions of oligodendrocytes. A study in primary oligodendrocyte cultures also revealed that chronic upregulation of Sgk1 by dexamethasone administration is involved in upregulation of Dsg1c mRNA. These results may indicate that chronic stress induced Sgk1 activation in oligodendrocytes, which increases Dsg1 expression near the plasma membrane. Thus, Dsg1 upregulation may be implicated in the molecular mechanisms underlying the morphological changes in oligodendrocytes in response to chronic stress exposure.

Fate of Mitral Regurgitation After Transcatheter Closure of Atrial Septal Defect in Adults.

Although the volume overload of pulmonary circulation improves after atrial septal defect (ASD) closure, the increasing left ventricular preload may contribute to mitral regurgitation (MR) deterioration. We aimed to evaluate the impact of MR after transcatheter ASD closure on clinical outcomes in adults. A total of 288 consecutive patients who underwent transcatheter ASD closure were enrolled. Changes in MR were assessed at 1 month after the procedure. The end point was defined as cardiovascular events. After the procedure, MR ameliorated in 3 patients and unchanged in 253, whereas MR deteriorated in 32. During a median follow-up of 24 months, patients with MR deterioration had no cardiovascular events, and the event-free survival rate was not different between patients with MR deterioration and those with MR amelioration or no-change (p = 0.355). Even in patients with MR deterioration, the New York Heart Association functional class improved after the procedure, with no cases of worsening functional class. Multivariate logistic regression analysis showed that MR deterioration was independently related to advanced age and female gender. The degree of enlargement of mitral valve annulus diameter after the procedure was greater in patients with MR deterioration than in those with MR amelioration or no-change, and it was correlated with the degree of MR deterioration. In conclusion, MR deterioration occurs in a minority of adult patients after transcatheter ASD closure; however, it is not linked with adverse outcomes. MR deterioration may be provoked by geometric changes in mitral valve annulus, especially in women with advanced age.

Physiological ER Stress Mediates the Differentiation of Fibroblasts.

Recently, accumulating reports have suggested the importance of endoplasmic reticulum (ER) stress signaling in the differentiation of several tissues and cells, including myoblasts and osteoblasts. Secretory cells are easily subjected to ER stress during maturation of their secreted proteins. Skin fibroblasts produce and release several proteins, such as collagens, matrix metalloproteinases (MMPs), the tissue inhibitors of metalloproteinases (TIMPs) and glycosaminoglycans (GAGs), and the production of these proteins is increased at wound sites. Differentiation of fibroblasts into myofibroblasts is one of the key factors for wound healing and that TGF-ß can induce fibroblast differentiation into myofibroblasts, which express α-smooth muscle actin. Well-differentiated myofibroblasts show increased production of collagen and TGF-ß, and bring about wound healing. In this study, we examined the effects of ER stress signaling on the differentiation of fibroblasts, which is required for wound healing, using constitutively ER stress-activated primary cultured fibroblasts. The cells expressed positive α-smooth muscle actin signals without TGF-ß stimulation compared with control fibroblasts. Gel-contraction assays suggested that ER stress-treated primary fibroblasts caused stronger shrinkage of collagen gels than control cells. These results suggest that ER stress signaling could accelerate the differentiation of fibroblasts to myofibroblasts at injured sites. The present findings may provide important insights for developing therapies to improve wound healing.

DBZ regulates cortical cell positioning and neurite development by sustaining the anterograde transport of Lis1 and DISC1 through control of Ndel1 dual-phosphorylation.

Cell positioning and neuronal network formation are crucial for proper brain function. Disrupted-in-Schizophrenia 1 (DISC1) is anterogradely transported to the neurite tips, together with Lis1, and functions in neurite extension via suppression of GSK3ß activity. Then, transported Lis1 is retrogradely transported and functions in cell migration. Here, we show that DISC1-binding zinc finger protein (DBZ), together with DISC1, regulates mouse cortical cell positioning and neurite development in vivo. DBZ hindered Ndel1 phosphorylation at threonine 219 and serine 251. DBZ depletion or expression of a double-phosphorylated mimetic form of Ndel1 impaired the transport of Lis1 and DISC1 to the neurite tips and hampered microtubule elongation. Moreover, application of DISC1 or a GSK3ß inhibitor rescued the impairments caused by DBZ insufficiency or double-phosphorylated Ndel1 expression. We concluded that DBZ controls cell positioning and neurite development by interfering with Ndel1 from disproportionate phosphorylation, which is critical for appropriate anterograde transport of the DISC1-complex.