bims-bicyki 2021-03-14 papers

bims-bicyki

Biomed News

on Bicaudal-C1 and interactors in cystic kidney disease

Issue of 2021–03–14
nineteen papers selected by
Céline Gagnieux, École Polytechnique Fédérale de Lausanne

The genetic background significantly impacts the severity of kidney cystic disease in the Pkd1RC/RC mouse model of autosomal dominant polycystic kidney disease.
The wind of change in the management of autosomal dominant polycystic kidney disease in childhood.
Assessing Risk of Rapid Progression in Autosomal Dominant Polycystic Kidney Disease and Special Considerations for Disease-Modifying Therapy.
Post-translational modification enzymes as key regulators of ciliary protein trafficking.
Changes of Motile Ciliary Phenotype in Patients with Primary Ciliopathies.
CiliaQ: a simple, open-source software for automated quantification of ciliary morphology and fluorescence in 2D, 3D, and 4D images.
Clinical Factors Associated with the Risk of Intracranial Aneurysm Rupture in Autosomal Dominant Polycystic Kidney Disease.
The OFD1 protein is a novel player in selective autophagy: another tile to the cilia/autophagy puzzle.
The multi-scale architecture of mammalian sperm flagella and implications for ciliary motility.
Mechanical stiffness augments ligand-dependent progesterone receptor B activation via MEK 1/2 and Rho/ROCK-dependent signaling pathways in uterine fibroid cells.
PKCα-mediated phosphorylation of the diacylglycerol kinase ζ MARCKS domain switches cell migration modes by regulating interactions with Rac1 and RhoA.
Primary cilia-dependent lipid raft/caveolin dynamics regulate adipogenesis.
Dual Inhibition of TGFβR and ROCK Reverses the Epithelial to Mesenchymal Transition in Collectively Migrating Zebrafish Keratocytes.
Primary cilia regulate gastric cancer-induced bone loss via cilia/Wnt/β-catenin signaling pathway.
Subcellular localization of the voltage-gated K+ channel EGL-36 , a member of the KV3 subfamily, in the ciliated sensory neurons in C. elegans.
Porcine Sapovirus-Induced Tight Junction Dissociation via Activation of RhoA/ROCK/MLC Signaling Pathway.
Integrin β1 coordinates survival and morphogenesis of the embryonic lineage upon implantation and pluripotency transition.
A novel JAK/ROCK inhibitor, CPL409116, demonstrates potent efficacy in the mouse model of systemic lupus erythematosus.
Early signaling pathways mediating dormant cyst formation in terrestrial unicellular eukaryote Colpoda.

Kidney Int. 2021 Mar 08. pii: S0085-2538(21)00259-3. [Epub ahead of print]

The genetic background significantly impacts the severity of kidney cystic disease in the Pkd1RC/RC mouse model of autosomal dominant polycystic kidney disease.

Jennifer Arroyo, Diana Escobar-Zarate, Harrison H Wells, Megan M Constans, Ka Thao, Jessica M Smith, Cynthia J Sieben, Madeline R Martell, Timothy L Kline, Maria V Irazabal, Vicente E Torres, Katharina Hopp, Peter C Harris.

  Autosomal dominant polycystic kidney disease (ADPKD), primarily due to PKD1 or PKD2 mutations, causes progressive kidney cyst development and kidney failure. There is significant intrafamilial variability likely due to the genetic background and environmental/lifestyle factors; variability that can be modeled in PKD mice. Here, we characterized mice homozygous for the PKD1 hypomorphic allele, p.Arg3277Cys (Pkd1RC/RC), inbred into the BalbC/cJ (BC) or the 129S6/SvEvTac (129) strains, plus F1 progeny bred with the previously characterized C57BL/6J (B6) model; F1(BC/B6) or F1(129/B6). By one-month cystic disease in both the BC and 129 Pkd1RC/RC mice was more severe than in B6 and continued with more rapid progression to six-nine months. Thereafter, the expansive disease stage plateaued/declined, coinciding with increased fibrosis and a clear decline in kidney function. Greater severity correlated with more inter-animal and inter-kidney disease variability, especially in the 129-line. Both F1 combinations had intermediate disease severity, more similar to B6 but progressive from one-month of age. Mild biliary dysgenesis, and an early switch from proximal tubule to collecting duct cysts, was seen in all backgrounds. Preclinical testing with a positive control, tolvaptan, employed the F1(129/B6)-Pkd1RC/RC line, which has moderately progressive disease and limited isogenic variability. Magnetic resonance imaging was utilized to randomize animals and provide total kidney volume endpoints; complementing more traditional data. Thus, we show how genetic background can tailor the Pkd1RC/RC model to address different aspects of pathogenesis and disease modification, and describe a possible standardized protocol for preclinical testing.

Keywords:  ADPKD; Animal Models; Disease Modifiers; PKD1; Preclinical Testing

DOI:  https://doi.org/10.1016/j.kint.2021.01.028
Pediatr Nephrol. 2021 Mar 07.

The wind of change in the management of autosomal dominant polycystic kidney disease in childhood.

Charlotte Gimpel, Carsten Bergmann, Djalila Mekahli.

  Significant progress has been made in understanding the genetic basis of autosomal dominant polycystic kidney disease (ADPKD), quantifying disease manifestations in children, exploring very-early onset ADPKD as well as pharmacological delay of disease progression in adults. At least 20% of children with ADPKD have relevant, yet mainly asymptomatic disease manifestations such as hypertension or proteinuria (in line with findings in adults with ADPKD, where hypertension and cardiovascular damage precede decline in kidney function). We propose an algorithm for work-up and management based on current recommendations that integrates the need to screen regularly for hypertension and proteinuria in offspring of affected parents with different options regarding diagnostic testing, which need to be discussed with the family with regard to ethical and practical aspects. Indications and scope of genetic testing are discussed. Pharmacological management includes renin-angiotensin system blockade as first-line therapy for hypertension and proteinuria. The vasopressin receptor antagonist tolvaptan is licensed for delaying disease progression in adults with ADPKD who are likely to experience kidney failure. A clinical trial in children is currently ongoing; however, valid prediction models to identify children likely to suffer kidney failure are lacking. Non-pharmacological interventions in this population also deserve further study.

Keywords:  Adolescent; Autosomal dominant polycystic kidney disease (ADPKD); Children; Hypertension; Neonate; Proteinuria; Tolvaptan

DOI:  https://doi.org/10.1007/s00467-021-04974-4
Am J Kidney Dis. 2021 Mar 08. pii: S0272-6386(21)00439-X. [Epub ahead of print]

Assessing Risk of Rapid Progression in Autosomal Dominant Polycystic Kidney Disease and Special Considerations for Disease-Modifying Therapy.

Fouad T Chebib, Vicente E Torres.

Autosomal Dominant Polycystic Kidney Disease is the most common inherited cause of kidney failure accounting for 5-10% of cases. Predicting which patient would progress rapidly to kidney failure is critical to assess the risk-benefit ratio of any intervention and consider early initiation of long-term kidney protective measures which will maximize the cumulative benefit of slowing down the disease progression. Surrogate prognostic biomarkers are required to predict future decline in kidney function. Clinical, genetic, environmental, epigenetic and radiological factors have been studied as predictors of progression to kidney failure in ADPKD. A complex interaction of these prognostic factors determines the number of kidney cysts and its growth rate which affect total kidney volume (TKV). Age-adjusted TKV, represented by the Mayo Imaging Classification, estimates each patient's unique rate of kidney growth and provides the most individualized approach available clinically so far. Tolvaptan has been approved to slow disease progression in patients at risk of rapidly progressive disease. Several other disease-modifying treatments are being studied in clinical trials. Selection criteria of rapid progression vary widely among countries, using a combination of age, baseline GFR, GFR slope, baseline TKV and TKV rate of growth. This review details the approach in assessing the risk of disease progression in ADPKD and identifying patients who would benefit from long-term therapy of disease-modifying agents.

DOI: https://doi.org/10.1053/j.ajkd.2020.12.020
J Biochem. 2021 Mar 03. pii: mvab024. [Epub ahead of print]

Post-translational modification enzymes as key regulators of ciliary protein trafficking.

Taro Chaya, Takahisa Furukawa.

  Primary cilia are evolutionarily conserved microtubule-based organelles that protrude from the surface of almost all cell types and decode a variety of extracellular stimuli. Ciliary dysfunction causes human diseases named ciliopathies, which span a wide range of symptoms, such as developmental and sensory abnormalities. The assembly, disassembly, maintenance, and function of cilia rely on protein transport systems including intraflagellar transport (IFT) and lipidated protein intraflagellar targeting (LIFT). IFT is coordinated by three multisubunit protein complexes with molecular motors along the ciliary axoneme, while LIFT is mediated by specific chaperones that directly recognize lipid chains. Recently, it has become clear that several post-translational modification enzymes play crucial roles in the regulation of IFT and LIFT. Here we review our current understanding of the roles of these post-translational modification enzymes in the regulation of ciliary protein trafficking as well as their regulatory mechanisms, physiological significance, and involvement in human diseases.

Keywords:  ICK/CILK1; MAK; kinase; retina; ubiquitin

DOI:  https://doi.org/10.1093/jb/mvab024
Adv Exp Med Biol. 2021 Mar 10.

Changes of Motile Ciliary Phenotype in Patients with Primary Ciliopathies.

Miroslava Brndiarova, Julia Kvassayova, Jarmila Vojtkova, Matus Igaz, Tomas Buday, Jana Plevkova.

  Primary ciliopathies are a group of disorders associated with abnormal formation and function of primary cilia. Many cilia-associated proteins found in primary cilia are also present in motile cilia. Such proteins are important for the ciliary base, such as the transition zone or basal bodies, and the intraflagellar transport. Their exact role in the respiratory motile cilia is unsettled. In this prospective clinical single-center study, we investigated the hypothesis that these proteins regulate the function of motile cilia. We addressed the issue by defining the motile cilia beat frequency in the respiratory tract of patients with primary ciliopathies accompanied by chronic kidney disease and comparing it in those without kidney involvement. Ciliary beat frequency in the nasal mucosa samples was evaluated by the ciliary analysis software LabVIEW. Both children and their parents with primary ciliopathies and kidney involvement had significantly lower median airway ciliary beat frequencies than those without kidney involvement who have normal ciliary motility. Further, the ciliary beat frequency is inversely associated with the serum creatinine level. These findings strongly suggest that kidney involvement in patients with primary ciliopathy may underlie the development of motile cilia dysfunction in the respiratory tract, potentially increasing respiratory morbidity.

Keywords:  Cilia; Ciliary beat frequency; Ciliopathy; Kidney disease; Respiratory morbidity

DOI:  https://doi.org/10.1007/5584_2021_617
Eur Phys J E Soft Matter. 2021 Mar 08. 44(2): 18

CiliaQ: a simple, open-source software for automated quantification of ciliary morphology and fluorescence in 2D, 3D, and 4D images.

Jan Niklas Hansen, Sebastian Rassmann, Birthe Stüven, Nathalie Jurisch-Yaksi, Dagmar Wachten.

Cilia are hair-like membrane protrusions that emanate from the surface of most vertebrate cells and are classified into motile and primary cilia. Motile cilia move fluid flow or propel cells, while also fulfill sensory functions. Primary cilia are immotile and act as a cellular antenna, translating environmental cues into cellular responses. Ciliary dysfunction leads to severe diseases, commonly termed ciliopathies. The molecular details underlying ciliopathies and ciliary function are, however, not well understood. Since cilia are small subcellular compartments, imaging-based approaches have been used to study them. However, tools to comprehensively analyze images are lacking. Automatic analysis approaches require commercial software and are limited to 2D analysis and only a few parameters. The widely used manual analysis approaches are time consuming, user-biased, and difficult to compare. Here, we present CiliaQ, a package of open-source, freely available, and easy-to-use ImageJ plugins. CiliaQ allows high-throughput analysis of 2D and 3D, static or time-lapse images from fluorescence microscopy of cilia in cell culture or tissues, and outputs a comprehensive list of parameters for ciliary morphology, length, bending, orientation, and fluorescence intensity, making it broadly applicable. We envision CiliaQ as a resource and platform for reproducible and comprehensive analysis of ciliary function in health and disease.

DOI: https://doi.org/10.1140/epje/s10189-021-00031-y
Cerebrovasc Dis. 2021 Mar 11. 1-8

Clinical Factors Associated with the Risk of Intracranial Aneurysm Rupture in Autosomal Dominant Polycystic Kidney Disease.

Chan-Hyuk Lee, Curie Ahn, Hyunjin Ryu, Hyun-Seung Kang, Seul-Ki Jeong, Keun-Hwa Jung.

   BACKGROUND: The occurrence of intracranial aneurysms is higher in patients with autosomal dominant polycystic kidney disease (ADPKD) than in the healthy population. However, research concerning the factors related to the risk of intracranial aneurysm rupture in patients with ADPKD is still insufficient.
OBJECTIVES: The aim of the study was to investigate the prevalence of intracranial aneurysms and aneurysmal subarachnoid hemorrhage (SAH) and to analyze the systemic factors associated with high-risk aneurysms in patients with ADPKD.
METHODS: We screened patients who underwent cerebral angiography between January 2007 and May 2017 in the ADPKD registry. Patients were examined for the presence of intracranial aneurysms and subsequently reclassified into 3 groups based on the risk of aneurysmal rupture: the aneurysm-negative (group 1), low-risk aneurysm (group 2), or high-risk aneurysm (group 3). Various systemic factors were compared, and independent factors associated with high-risk aneurysms were analyzed.
RESULTS: Among the 926 patients, 148 (16.0%) had intracranial aneurysms and 11 (1.2%) had previous aneurysmal SAH. Patients with intracranial aneurysms were further classified into group 2 (low-risk aneurysms, 15.5%) or group 3 (high-risk aneurysms, 84.5%). Age (odds ratio [OR] 1.03, 95% confidence interval [CI] 1.01-1.05, p = 0.004), female sex (OR 3.13, 95% CI 1.94-5.0 6, p < 0.001), dolichoectasia (OR 8.57, 95% CI 1.53-48.17, p = 0.015), and mitral inflow deceleration time (DT) (OR 1.01, 95% CI 1.00-1.01, p = 0.046) were independently associated with high-risk aneurysms, whereas hypercholesterolemia (OR 0.46, 95% CI 0.29-0.72, p = 0.001) was negatively associated.
CONCLUSION: In the present study among patients with ADPKD, the prevalence of intracranial aneurysms and aneurysmal SAH was 16 and 1.2%, respectively. Age, female sex, dolichoectasia, and mitral inflow DT were positively associated with high-risk aneurysms, whereas hypercholesterolemia was negatively associated. A subsequent large-scaled longitudinal study is needed to define the plausibility of the clinical parameters.

Keywords:  Aneurysmal rupture; Autosomal dominant polycystic kidney disease; Dolichoectasia; Hypercholesterolemia; Intracranial aneurysm; Mitral inflow deceleration time

DOI:  https://doi.org/10.1159/000513709
Cell Stress. 2021 Feb 17. 5(3): 33-36

The OFD1 protein is a novel player in selective autophagy: another tile to the cilia/autophagy puzzle.

Manuela Morleo, Brunella Franco.

  The autophagy-lysosomal pathway is one of the main degradative routes which cells use to balance sources of energy. A number of proteins orchestrate the formation of autophagosomes, membranous organelles instrumental in autophagy. Selective autophagy, involving the recognition and removal of specific targets, is mediated by autophagy receptors, which recognize cargos and the autophagosomal membrane protein LC3 for lysosomal degradation. Recently, bidirectional crosstalk has emerged between autophagy and primary cilia, microtubule-based sensory organelles extending from cells and anchored by the basal body, derived from the mother centriole of the centrosome. The molecular mechanisms underlying the direct role of autophagic proteins in cilia biology and, conversely, the impact of this organelle in autophagy remains elusive. Recently, we uncovered the molecular mechanism by which the centrosomal/basal body protein OFD1 controls the LC3-mediated autophagic cascade. In particular, we demonstrated that OFD1 acts as a selective autophagy receptor by regulating the turnover of unc-51-like kinase (ULK1) complex, which plays a crucial role in the initiation steps of autophagosome biogenesis. Moreover, we showed that patients with a genetic condition caused by mutations in OFD1 and associated with cilia dysfunction, display excessive autophagy and we demonstrated that autophagy inhibition significantly ameliorates the renal cystic phenotype in a conditional mouse model recapitulating the features of the disease (Morleo et al. 2020, EMBO J, doi: 10.15252/embj.2020105120). We speculate that abnormal autophagy may underlie some of the clinical manifestations observed in the disorders ascribed to cilia dysfunction.

Keywords:  Autophagy receptor; OFD1; Oral-Facial-Digital type I syndrome; Primary cilium; Renal cystic disease; Selective autophagy; ULK1 complex

DOI:  https://doi.org/10.15698/cst2021.03.244
EMBO J. 2021 Mar 10. e107410

The multi-scale architecture of mammalian sperm flagella and implications for ciliary motility.

Miguel Ricardo Leung, Marc C Roelofs, Ravi Teja Ravi, Paula Maitan, Heiko Henning, Min Zhang, Elizabeth G Bromfield, Stuart C Howes, Bart M Gadella, Hermes Bloomfield-Gadêlha, Tzviya Zeev-Ben-Mordehai.

  Motile cilia are molecular machines used by a myriad of eukaryotic cells to swim through fluid environments. However, available molecular structures represent only a handful of cell types, limiting our understanding of how cilia are modified to support motility in diverse media. Here, we use cryo-focused ion beam milling-enabled cryo-electron tomography to image sperm flagella from three mammalian species. We resolve in-cell structures of centrioles, axonemal doublets, central pair apparatus, and endpiece singlets, revealing novel protofilament-bridging microtubule inner proteins throughout the flagellum. We present native structures of the flagellar base, which is crucial for shaping the flagellar beat. We show that outer dense fibers are directly coupled to microtubule doublets in the principal piece but not in the midpiece. Thus, mammalian sperm flagella are ornamented across scales, from protofilament-bracing structures reinforcing microtubules at the nano-scale to accessory structures that impose micron-scale asymmetries on the entire assembly. Our structures provide vital foundations for linking molecular structure to ciliary motility and evolution.

Keywords:  centrioles; cryo-FIB milling; cryo-electron tomography; motile cilia; sperm

DOI:  https://doi.org/10.15252/embj.2020107410
Fertil Steril. 2021 Mar 03. pii: S0015-0282(20)32753-9. [Epub ahead of print]

Mechanical stiffness augments ligand-dependent progesterone receptor B activation via MEK 1/2 and Rho/ROCK-dependent signaling pathways in uterine fibroid cells.

Christina N Cordeiro Mitchell, Md Soriful Islam, Sadia Afrin, Joshua Brennan, Kevin J Psoter, James H Segars.

   OBJECTIVE: To test whether mechanical substrate stiffness would influence progesterone receptor B (PRB) signaling in fibroid cells. Uterine fibroids feature an excessive extracellular matrix, increased stiffness, and altered mechanical signaling. Fibroid growth is stimulated by progestins and opposed by anti-progestins, but a functional interaction between progesterone action and mechanical signaling has not been evaluated.
DESIGN: Laboratory studies.
SETTING: Translational science laboratory.
PATIENT(S)/ANIMAL(S): Human fibroid cell lines and patient-matched fibroid and myometrial cell lines.
INTERVENTION(S): Progesterone receptor B-dependent reporter assays and messenger RNA quantitation in cells cultured on stiff polystyrene plates (3GPa) or soft silicone plates (930KPa). Pharmacologic inhibitors of extracellular signal-related protein kinase (ERK) kinase 1/2 (MEK 1/2; PD98059), p38 mitogen-activated protein kinase (SB202190), receptor tyrosine kinases (RTKs; nintedanib), RhoA (A13), and Rho-associated coiled-coil kinase (ROCK; Y27632).
MAIN OUTCOME MEASURE(S): Progesterone-responsive reporter activation.
RESULT(S): Fibroid cells exhibited higher PRB-dependent reporter activity with progesterone (P4) in cells cultured on stiff vs. soft plates. Mechanically induced PRB activation with P4 was decreased 62% by PD98059, 78% by nintedanib, 38% by A13, and 50% by Y27632. Overexpression of the Rho-guanine nucleotide exchange factor (Rho-GEF), AKAP13, significantly increased PRB-dependent reporter activity. Collagen 1 messenger RNA levels were higher in fibroid cells grown on stiff vs. soft plates with P4.
CONCLUSION(S): Cells cultured on mechanically stiff substrates had enhanced PRB activation via a mechanism that required MEK 1/2 and AKAP13/RhoA/ROCK signaling pathways. These studies provide a framework to explore the mechanisms by which mechanical stiffness affects progesterone receptor activation.

Keywords:  Progesterone signaling; mechanotransduction; nonclassical signaling; progesterone receptor B; uterine leiomyoma

DOI:  https://doi.org/10.1016/j.fertnstert.2020.12.011
J Biol Chem. 2021 Mar 04. pii: S0021-9258(21)00292-1. [Epub ahead of print] 100516

PKCα-mediated phosphorylation of the diacylglycerol kinase ζ MARCKS domain switches cell migration modes by regulating interactions with Rac1 and RhoA.

Ryan Ard, Jean-Christian Maillet, Elias Daher, Michael Phan, Radoslav Zinoviev, Robin J Parks, Stephen H Gee.

  Cells can switch between Rac1 (lamellipodia-based) and RhoA (blebbing-based) migration modes but the molecular mechanisms regulating this shift are not fully understood. Diacylglycerol kinase ζ (DGKζ), which phosphorylates diacylglycerol to yield phosphatidic acid, forms independent complexes with Rac1 and RhoA, selectively dissociating each from their common inhibitor RhoGDI. DGKζ catalytic activity is required for Rac1 dissociation but is dispensable for RhoA dissociation; instead, DGKζ stimulates RhoA release via a kinase-independent scaffolding mechanism. The molecular determinants that mediate the selective targeting of DGKζ to Rac1 or RhoA signaling complexes are unknown. Here, we show that protein kinase Cα (PKCα)-mediated phosphorylation of the DGKζ MARCKS domain increased DGKζ association with RhoA and decreased its interaction with Rac1. The same modification also enhanced DGKζ interaction with the scaffold protein syntrophin. Expression of a phosphomimetic DGKζ mutant stimulated membrane blebbing in mouse embryonic fibroblasts and C2C12 myoblasts, which was augmented by inhibition of endogenous Rac1. DGKζ expression in differentiated C2 myotubes, which have low endogenous Rac1 levels, also induced substantial membrane blebbing via the RhoA-ROCK pathway. These events were independent of DGKζ catalytic activity, but dependent upon a functional C-terminal PDZ-binding motif. Rescue of RhoA activity in DGKζ-null cells also required the PDZ-binding motif, suggesting syntrophin interaction is necessary for optimal RhoA activation. Collectively, our results define a switch-like mechanism whereby DGKζ phosphorylation by PKCα plays a role in the interconversion between Rac1 and RhoA signaling pathways that underlie different cellular migration modes.

Keywords:  DAGK); PDZ domain; Ras homolog gene family; Ras‐related C3 botulinum toxin substrate 1 (Rac1); bleb; cell biology; cell migration; cell signaling; diacylglycerol; diacylglycerol kinase (DGK; fibroblast; lamellipodium; member A (RhoA); membrane; phosphatidic acid; phosphorylation; protein kinase C (PKC); scaffold protein; skeletal muscle; syntrophin

DOI:  https://doi.org/10.1016/j.jbc.2021.100516
Cell Rep. 2021 Mar 09. pii: S2211-1247(21)00131-5. [Epub ahead of print]34(10): 108817

Primary cilia-dependent lipid raft/caveolin dynamics regulate adipogenesis.

Daishi Yamakawa, Daisuke Katoh, Kousuke Kasahara, Takashi Shiromizu, Makoto Matsuyama, Chise Matsuda, Yumi Maeno, Masatoshi Watanabe, Yuhei Nishimura, Masaki Inagaki.

  Primary cilia play a pivotal role in signal transduction and development and are known to serve as signaling hubs. Recent studies have shown that primary cilium dysfunction influences adipogenesis, but the mechanisms are unclear. Here, we show that mesenchymal progenitors C3H10T1/2 depleted of trichoplein, a key regulator of cilium formation, have significantly longer cilia than control cells and fail to differentiate into adipocytes. Mechanistically, the elongated cilia prevent caveolin-1- and/or GM3-positive lipid rafts from being assembled around the ciliary base where insulin receptor proteins accumulate, thereby inhibiting the insulin-Akt signaling. We further generate trichoplein knockout mice, in which adipogenic progenitors display elongated cilia and impair the lipid raft dynamics. The knockout mice on an extended high-fat diet exhibit reduced body fat and smaller adipocytes than wild-type (WT) mice. Overall, our results suggest a role for primary cilia in regulating adipogenic signal transduction via control of the lipid raft dynamics around cilia.

Keywords:  Akt; adipogenesis; caveolae; ciliary base; insulin signaling; lipid rafts; obesity; primary cilia; trichoplein

DOI:  https://doi.org/10.1016/j.celrep.2021.108817
Cell Biol Int. 2021 Mar 12.

Dual Inhibition of TGFβR and ROCK Reverses the Epithelial to Mesenchymal Transition in Collectively Migrating Zebrafish Keratocytes.

Agnes S Pascual, Jose L Rapanan, Chandana K Uppalapati, Kimbal E Cooper, Kathryn J Leyva, Elizabeth E Hull.

  There is a growing controversy about the role of the epithelial to mesenchymal transition (EMT) in the fibrosis associated with chronic disease. Recent studies suggest that it is not the EMT transcriptional program but differentiation of progenitor cells, response to chronic inflammation, or some combination of both which cause the appearance of fibroblasts and the production of extracellular matrix. To address this issue, we study the EMT process in the zebrafish keratocytes which migrate from primary explants of epithelial tissue as these cells are both terminally differentiated and able to divide. To firmly place this EMT process in the context of other systems, we first demonstrate that the zebrafish keratocyte EMT process involves nuclear accumulation of twist and snail/slug transcription factors as part of a TGFβR-mediated EMT process. As assessed by expression and localization of EMT transcription factors, the zebrafish keratocyte EMT process is reversed by the addition of Rho activated kinase (ROCK) in combination with TGFβR inhibitors. The complete cycle of EMT to MET observed in this system links these in vitro results more closely to the process of wound healing in vivo. However, the absence of observable activation of EMT transcription factors when keratocytes are cultured on compliant substrata in a TGFβ1-containing medium suggests that ROCK signaling, initiated by tension within the sheet, is an essential contributor to the EMT process. Most importantly, the requirement for ROCK activation by culturing on non-compliant substrata suggests that EMT in these terminally differentiated cells would not occur in vivo. This article is protected by copyright. All rights reserved.

Keywords:  Actin; Protein kinases/phosphatases; Protein signalling modules/scaffolds; Transcription/transcription factors

DOI:  https://doi.org/10.1002/cbin.11587
Aging (Albany NY). 2021 Mar 09. 13

Primary cilia regulate gastric cancer-induced bone loss via cilia/Wnt/β-catenin signaling pathway.

Jie Xu, Xiaoyan Deng, Xiangmei Wu, Huifang Zhu, Yinghua Zhu, Jie Liu, Qian Chen, Chengfu Yuan, Geli Liu, Changdong Wang.

  Cancer-associated bone disease is a frequent occurrence in cancer patients and is associated with pain, bone fragility, loss, and fractures. However, whether primary or non-bone metastatic gastric cancer induces bone loss remains unclear. Here, we collected clinical evidence of bone loss by analyzing serum and X-rays of 25 non-bone metastatic gastric cancer patients. In addition, C57BL mice were injected with the human gastric cancer cell line HGC27 and its effect on bone mass was analyzed by Micro-CT, immunoblotting, and immunohistochemistry. Furthermore, the degree of the proliferation and osteogenic differentiation of mesenchymal stem cells (MSCs) co-cultured with HGC-27 or SGC-7901 cells was analyzed by colony-formation assay, alizarin red staining, immunofluorescence, qPCR, immunoblotting, and alkaline phosphatase activity assay. These indicated that gastric cancer could damage bone tissue before the occurrence of bone metastases. We also found that cilia formation of MSCs was increased in the presence of HGC27 cells, which was associated with abnormal activation of the Wnt/β-catenin pathway. Expression of DKK1 inhibited the Wnt/β-catenin signaling pathway and partially rescued osteogenic differentiation of MSCs. In summary, our results suggest that gastric cancer cells might cause bone damage prior to the occurrence of bone metastasis via cilia-dependent activation of the Wnt/β-catenin signaling pathway.

Keywords:  Wnt/β-catenin signaling pathway; bone loss; cilia; gastric cancer

DOI:  https://doi.org/10.18632/aging.202734
MicroPubl Biol. 2021 Mar 05. 2021

Subcellular localization of the voltage-gated K+ channel EGL-36 , a member of the KV3 subfamily, in the ciliated sensory neurons in C. elegans.

Sebiha Cevik, Oktay I Kaplan.

Delineated as the first cellular organelle in 1675 by Antonie van Leeuwenhoek, cilia did not receive much attention until the 2000s, when it became apparent that cilia played a key role in the development of embryos, a variety of signaling pathways. Therefore, collective efforts by many scientists have led to the identification of many novel ciliopathy and cilia genes, while we are still far from disclosing the complete components of cilia.Here we used the ciliated sensory neurons in C. elegans as a model system that revealed the voltage-gated K+ channel EGL-36 (a member of the Shaw subfamily) as a new component associated with cilia. The confocal microscopy examination of fluorescence tagged EGL-36 together with ciliary (IFT-140) or transition zone (MKS-6) markers reveal that EGL-36 is only expressed in subsets of the ciliated sensory neurons, where it partially overlaps with the basal body signals and predominantly localizes to the periciliary membrane compartment. This expression pattern along with studies of egl-36 gain-of-function variants indicates that egl-36 is not essential for ciliogenesis in C. elegans. Our data identify the voltage-gated K+ channel EGL-36 as a new cilia-associated protein, and future studies should reveal the functional significance of EGL-36 in cilia biogenesis.

DOI: https://doi.org/10.17912/micropub.biology.000367
J Virol. 2021 Mar 10. pii: JVI.00051-21. [Epub ahead of print]

Porcine Sapovirus-Induced Tight Junction Dissociation via Activation of RhoA/ROCK/MLC Signaling Pathway.

Muhammad Sharif, Yeong-Bin Baek, Ahsan Naveed, Nattan Stalin, Mun-Il Kang, Sang-Ik Park, Mahmoud Soliman, Kyoung-Oh Cho.

Tight junctions (TJs) are a major barrier and also an important portal of entry for different pathogens. Porcine sapovirus (PSaV) induces early disruption of the TJ integrity of polarized LLC-PK cells, allowing it to bind to the buried occludin co-receptors hidden beneath the TJs on the basolateral surface. However, the signaling pathways involved in the PSaV-induced TJ dissociation are not yet known. Here, we found that the RhoA/ROCK/MLC signaling pathway was activated in polarized LLC-PK cells during the early infection of PSaV Cowden strain in the presence of bile acid. Specific inhibitors of RhoA, ROCK, and MLC restored PSaV-induced reduction of transepithelial resistance, increase of paracellular flux, intracellular translocation of occludin, and lateral membrane lipid diffusion. Moreover, each inhibitor significantly reduced PSaV replication, as evidenced by a reduction in viral protein synthesis, genome copy number, and progeny viruses. The PKC/MLCK and RhoA/ROCK/MYPT signaling pathways, known to dissociate TJs, were not activated during early PSaV infection. Among the above signaling pathways, the RhoA/ROCK/MLC signaling pathway was only activated by PSaV in the absence of bile acid, and specific inhibitors of this signaling pathway restored early TJ dissociation. Our findings demonstrate that PSaV binding to cell surface receptors activates the RhoA/ROCK/MLC signaling pathway, which in turn disrupts TJ integrity via the contraction of the actomyosin ring. Our study contributes to understanding how PSaV enters the cells and will aid in developing efficient and affordable therapies against PSaV and other calicivirus infections.IMPORTANCEPorcine sapovirus (PSaV), one of the most important enteric pathogens, is known to disrupt tight junction (TJ) integrity to expose its buried co-receptor occludin in polarized LLC-PK cells. However, the cellular signaling pathways that facilitate TJ dissociation are not yet completely understood. Here, we demonstrate that early infection of PSaV in polarized LLC-PK cells in either the presence or absence of bile acids activates the RhoA/ROCK/MLC signaling pathway, whose inhibitors reverse the early PSaV infection-induced early dissociation of TJs and reduce PSaV replication. However, early PSaV infection did not activate the PKC/MLCK and RhoA/ROCK/MYPT signaling pathways, which are also known to dissociate TJs. This study provides a better understanding of the mechanism involved in early PSaV infection-induced disruption of TJs, which is important for controlling or preventing PSaV and other calicivirus infections.

DOI: https://doi.org/10.1128/JVI.00051-21
Cell Rep. 2021 Mar 09. pii: S2211-1247(21)00148-0. [Epub ahead of print]34(10): 108834

Integrin β1 coordinates survival and morphogenesis of the embryonic lineage upon implantation and pluripotency transition.

Matteo Amitaba Molè, Antonia Weberling, Reinhard Fässler, Alison Campbell, Simon Fishel, Magdalena Zernicka-Goetz.

  At implantation, the embryo establishes contacts with the maternal endometrium. This stage is associated with a high incidence of preclinical pregnancy losses. While the maternal factors underlying uterine receptivity have been investigated, the signals required by the embryo for successful peri-implantation development remain elusive. To explore these, we studied integrin β1 signaling, as embryos deficient for this receptor degenerate at implantation. We demonstrate that the coordinated action of pro-survival signals and localized actomyosin suppression via integrin β1 permits the development of the embryo beyond implantation. Failure of either process leads to developmental arrest and apoptosis. Pharmacological stimulation through fibroblast growth factor 2 (FGF2) and insulin-like growth factor 1 (IGF1), coupled with ROCK-mediated actomyosin inhibition, rescues the deficiency of integrin β1, promoting progression to post-implantation stages. Mutual exclusion between integrin β1 and actomyosin seems to be conserved in the human embryo, suggesting the possibility that these mechanisms could also underlie the transition of the human epiblast from pre- to post-implantation.

Keywords:  actomyosin; apoptosis; epiblast; human embryo; integrins; morphogenesis; mouse embryo; survival

DOI:  https://doi.org/10.1016/j.celrep.2021.108834
J Pharmacol Sci. 2021 Apr;pii: S1347-8613(21)00013-X. [Epub ahead of print]145(4): 340-348

A novel JAK/ROCK inhibitor, CPL409116, demonstrates potent efficacy in the mouse model of systemic lupus erythematosus.

Maria Dulak-Lis, Anna Bujak, Kamila Gala, Martyna Banach, Urszula Kędzierska, Joanna Miszkiel, Joanna Hucz-Kalitowska, Michał Mroczkiewicz, Bartosz Stypik, Krzysztof Szymczak, Paweł Gunerka, Krzysztof Dubiel, Beata M Zygmunt, Maciej Wieczorek, Jerzy S Pieczykolan.

  Systemic lupus erythematosus is a chronic inflammatory disease, in which treatment is still limited due to suboptimal efficacy and toxicities associated with the available therapies. JAK kinases are well known to play an important role in systemic lupus erythematous. There is growing evidence that ROCK kinases are also important in disease development. In this paper, we present the results of the development of CPL409116, a dual JAK and ROCK inhibitor. The studies we performed demonstrate that this molecule is an effective JAK and ROCK inhibitor which efficiently blocks disease progression in NZBWF1/J mouse models of systemic lupus erythematous.

Keywords:  JAK1; JAK3; Lupus; Nephritis; ROCK

DOI:  https://doi.org/10.1016/j.jphs.2021.02.002
FEMS Microbiol Lett. 2021 Mar 02. pii: fnab019. [Epub ahead of print]

Early signaling pathways mediating dormant cyst formation in terrestrial unicellular eukaryote Colpoda.

Tatsuomi Matsuoka.

  Dormant (resting) cyst formation (encystment) in unicellular eukaryotes is the process of a large-scale digestion of vegetative cell structures and reconstruction into the dormant form, which is performed by cell signaling pathways accompanied by up- or downregulation of protein expression, and by posttranslational modification such as phosphorylation. In this review, the author describes the morphogenetic events during encystment of Colpoda and the early molecular events in the Ca2+/calmodulin-triggered signaling pathways for encystment, based mainly on our research results of the past 10 years; especially, the author discusses the role of c-AMP dependently phosphorylated proteins (ribosomal P0 protein, ribosomal S5 protein, Rieske iron-sulfur protein, actin, histone H4) and encystment-dependently upregulated (EF-1α, HSP60, actin-related protein) and downregulated proteins (ATP synthase β chain). In addition, the roles of AMPK, a key molecule in the signaling pathways leading to Colpoda encystment, and differentially expressed genes and proteins during encystment of other ciliates are discussed.

Keywords:  Ca2+; cAMP; calmodulin; cell signaling; encystment; protein phosphorylation

DOI:  https://doi.org/10.1093/femsle/fnab019