scholarly journals STAM and Hrs Down-Regulate Ciliary TRP Receptors

2007 ◽  
Vol 18 (9) ◽  
pp. 3277-3289 ◽  
Author(s):  
Jinghua Hu ◽  
Samuel G. Wittekind ◽  
Maureen M. Barr
Keyword(s):  
Growth Factor ◽  
Mammalian Cells ◽  
Primary Cilia ◽  
Membrane Receptors ◽  
Sensory Transduction ◽  
Kinase Substrate ◽  
C Elegans ◽  
Early Endosomes ◽  
Transient Receptor ◽  
And Function

Cilia are endowed with membrane receptors, channels, and signaling components whose localization and function must be tightly controlled. In primary cilia of mammalian kidney epithelia and sensory cilia of Caenorhabditis elegans neurons, polycystin-1 (PC1) and transient receptor polycystin-2 channel (TRPP2 or PC2), function together as a mechanosensory receptor-channel complex. Despite the importance of the polycystins in sensory transduction, the mechanisms that regulate polycystin activity and localization, or ciliary membrane receptors in general, remain poorly understood. We demonstrate that signal transduction adaptor molecule STAM-1A interacts with C. elegans LOV-1 (PC1), and that STAM functions with hepatocyte growth factor–regulated tyrosine kinase substrate (Hrs) on early endosomes to direct the LOV-1-PKD-2 complex for lysosomal degradation. In a stam-1 mutant, both LOV-1 and PKD-2 improperly accumulate at the ciliary base. Conversely, overexpression of STAM or Hrs promotes the removal of PKD-2 from cilia, culminating in sensory behavioral defects. These data reveal that the STAM-Hrs complex, which down-regulates ligand-activated growth factor receptors from the cell surface of yeast and mammalian cells, also regulates the localization and signaling of a ciliary PC1 receptor-TRPP2 complex.

scholarly journals The retromer complex regulates C. elegans development and mammalian ciliogenesis

2021 ◽  
Author(s):  
Shuwei Xie ◽  
Ellie Smith ◽  
Carter Dierlam ◽  
Danita Mathew ◽  
Angelina Davis ◽  
...  
Keyword(s):  
Potential Function ◽  
Primary Cilium ◽  
Mammalian Cells ◽  
Primary Cilia ◽  
Vulval Development ◽  
Sorting Nexin ◽  
Capping Protein ◽  
C Elegans ◽  
Retromer Complex ◽  
Mother Centriole

The mammalian retromer is comprised of subunits VPS26, VPS29 and VPS35, and a more loosely-associated sorting nexin (SNX) heterodimer. Despite known roles for the retromer in multiple trafficking events in yeast and mammalian cells, its role in development is poorly understood, and its potential function in primary ciliogenesis remains unknown. Using CRISPR-Cas9 editing, we demonstrated that vps-26 homozygous knockout C. elegans have reduced brood sizes and impaired vulval development, as well as decreased body length which has been linked to defects in primary ciliogenesis. Since many endocytic proteins are implicated in the generation of primary cilia, we addressed whether the retromer regulates ciliogenesis in mammalian cells. We observed VPS35 localized to the primary cilium, and depletion of VPS26, VPS35 or SNX1/SNX5 led to decreased ciliogenesis. Retromer also coimmunoprecipitated with the capping protein, CP110, and was required for its removal from the mother centriole. Herein, we characterize new roles for the retromer in C. elegans development and in the regulation of ciliogenesis in mammalian cells, and suggest a novel role for the retromer in CP110 removal from the mother centriole.

scholarly journals Epidermal-growth-factor-stimulated phosphorylation of calpactin II in membrane vesicles shed from cultured A-431 cells

1989 ◽  
Vol 259 (2) ◽  
pp. 577-583 ◽  
Author(s):  
J Blay ◽  
K A Valentine-Braun ◽  
J K Northup ◽  
M D Hollenberg
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Egf Receptor ◽  
Membrane Vesicles ◽  
Receptor Internalization ◽  
Receptor Kinase ◽  
Kinase Substrate ◽  
Epidermal Growth ◽  
Shed Vesicles ◽  
And Function

Membrane vesicles shed from intact A-431 epidermoid carcinoma cells and harvested in the presence of Ca2+ contained epidermal-growth-factor (EGF) receptor/kinase substrates of apparent molecular masses 185, 85, 70, 55, 38 and 27 kDa. The 38 kDa substrate (p38) was recognized by an antibody that had been raised against the human placental EGF receptor/kinase substrate calpactin II (lipocortin I). The A-431 and placental substrates, isolated by immunoprecipitation after phosphorylation in situ, yielded identical phosphopeptide maps upon limited proteolytic digestion with each of five different enzymes. The A-431-cell vesicular p38 is therefore calpactin II. EGF treatment of the intact A-431 cells before inducing vesiculation was not necessary for the substrate to be present within the vesicles. Our data thus indicate that receptor internalization is not a prerequisite for receptor-mediated phosphorylation of calpactin II. The ability of the protein to function as a substrate for the receptor/kinase depended upon the continued presence of Ca2+ during the vesicle-isolation procedure. EGF-stimulated phosphorylation of calpactin II was much less pronounced in vesicles prepared from A-431 cells in the absence of Ca2+, although comparable amounts of the protein were detectable by immunoblotting. Calpactin II therefore appears to be sequestered in a Ca2+-modulated manner within shed vesicles, along with at least four other major targets for the EGF receptor/kinase. The vesicle preparation may be a useful model system in which to study the phosphorylation and function of potentially important membrane-associated substrates for the receptor.

Characterization, regulation, and function of specific cell membrane receptors for insulin-like growth factor I on bone endothelial cells

2009 ◽  
Vol 9 (3) ◽  
pp. 329-337 ◽  
Author(s):  
Gianna Fiorelli ◽  
Claudio Orlando ◽  
Susanna Benvenuti ◽  
Francesco Franceschelli ◽  
Sandro Bianchi ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Growth Factor ◽  
Cell Membrane ◽  
Membrane Receptors ◽  
Specific Cell ◽  
Insulin Like Growth Factor ◽  
Factor I ◽  
Growth Factor I ◽  
And Function

scholarly journals xbx-4, a homolog of the Joubert syndrome gene FAM149B1, acts via the CCRK and MAK kinase cascade to regulate cilia morphology

2021 ◽  
Author(s):  
Ashish K Maurya ◽  
Piali Sengupta
Keyword(s):  
Sensory Neuron ◽  
Mammalian Cells ◽  
Primary Cilia ◽  
Cell Types ◽  
Joubert Syndrome ◽  
C Elegans ◽  
Kinase Cascade ◽  
Multiple Cell ◽  
Kinase Pathway ◽  
Sensory Functions

Primary cilia are microtubule (MT)-based organelles that mediate sensory functions in multiple cell types. Disruption of cilia structure or function leads to a diverse collection of diseases termed ciliopathies. Mutations in the DUF3719 domain-containing protein FAM149B1 have recently been shown to elongate cilia via unknown mechanisms and result in the ciliopathy Joubert syndrome. The highly conserved CCRK and MAK/RCK kinases negatively regulate cilia length and structure in Chlamydomonas, C. elegans, and mammalian cells. How the activity of this kinase cascade is tuned to precisely regulate cilia architecture is unclear. Here we identify XBX-4, a DUF3719 domain-containing protein related to human FAM149B1, as a novel regulator of the DYF-18 CCRK and DYF-5 MAK kinase pathway in C. elegans. As in dyf-18 and dyf-5 mutants, sensory neuron cilia are elongated in xbx-4 mutants and exhibit altered axonemal MT stability. XBX-4 promotes DYF-18 CCRK activity to regulate DYF-5 MAK function and localization. We find that Joubert syndrome-associated mutations in the XBX-4 DUF3719 domain also elongate cilia in C. elegans. Our results identify a new metazoan-specific regulator of this highly conserved kinase pathway, and suggest that FAM149B1 may similarly act via the CCRK/MAK kinase pathway to regulate ciliary homeostasis in humans.

scholarly journals Polycystic Diseases in Visceral Organs

2011 ◽  
Vol 2011 ◽  
pp. 1-7 ◽  
Author(s):  
Shakila Abdul-Majeed ◽  
Surya M. Nauli
Keyword(s):  
Mammalian Cells ◽  
Primary Cilia ◽  
Polycystic Ovarian Syndrome ◽  
Cyst Formation ◽  
Apical Surface ◽  
Polycystic Kidney ◽  
Pancreas Disease ◽  
Polycystic Liver ◽  
And Function ◽  
Ovarian Syndrome

Primary cilia are nonmotile, microtubule-based, antenna-like organelles projecting from the apical surface of most mammalian cells. Elegant studies have established the importance of ciliary structure and function in signal transduction and the sensory roles of cilia in maintaining healthy cellular state. In particular, dysfunctional cilia have been implicated in a large number of diseases mainly characterized by the presence of fluid-filled cysts in various organs. Aside from polycystic kidney disease (PKD), however, the roles of cilia in polycystic liver disease (PLD), polycystic pancreas disease (PPD), and polycystic ovarian syndrome (PCOS) are still very vague. In addition, although gender and sex hormones are known to regulate cyst formation, their roles in regulating physiological functions of cilia need to be further explored.

scholarly journals Signaling compartment at the ciliary tip is formed and maintained by intraflagellar transport and functions as sensitive salt detector

2019 ◽  
Author(s):  
Servaas N. van der Burght ◽  
Suzanne Rademakers ◽  
Jacque-Lynne Johnson ◽  
Chunmei Li ◽  
Gert-Jan Kremers ◽  
...  
Keyword(s):  
Primary Cilia ◽  
Physiological Function ◽  
Intraflagellar Transport ◽  
Distal Region ◽  
Central Question ◽  
Membrane Diffusion ◽  
High Concentration ◽  
C Elegans ◽  
Cgmp Signaling ◽  
And Function

AbstractPrimary cilia are ubiquitous antenna-like organelles that mediate cellular signaling and represent hotspots for human diseases termed ciliopathies. How signaling subcompartments are established within the microtubule-based organelle, and for example support Hedgehog or cGMP signal transduction pathways, remains a central question. Here we show that a C. elegans salt-sensing receptor type guanylate cyclase, GCY-22, accumulates at a high concentration within the distal region of the cilium. This receptor uses DAF-25 (Ankmy2 in mammals) to cross the transition zone (TZ) membrane diffusion barrier in the proximal-most region of the ciliary axoneme. Targeting of GCY-22 to the ciliary tip is dynamic, requiring the cargo-mobilizing intraflagellar transport (IFT) system. Disruption of transit across the TZ barrier or IFT trafficking causes GCY-22 protein mislocalization and defects in the formation, maintenance, and function of the ciliary tip compartment required for chemotaxis to low NaCl concentrations. Together, our findings reveal how a previously undescribed cilium tip cGMP signaling compartment is established and contributes to the physiological function of a primary cilium.

scholarly journals A polarity pathway for exocyst-dependent intracellular tube extension

2020 ◽  
Author(s):  
Joshua Abrams ◽  
Jeremy Nance
Keyword(s):  
Mammalian Cells ◽  
Rho Gtpase ◽  
Tube Formation ◽  
Membrane Domain ◽  
Lumen Formation ◽  
Vesicle Tethering ◽  
C Elegans ◽  
And Function ◽  
Excretory Cell ◽  
Upstream Regulators

ABSTRACTLumen extension in intracellular tubes can occur by the directed fusion of vesicles with an invading apical membrane domain. Within the C. elegans excretory cell, which contains an intracellular tube, the exocyst vesicle-tethering complex is enriched at the lumenal membrane domain and is required for tube formation, suggesting that it targets vesicles needed for lumen extension. Here, we identify a polarity pathway that promotes intracellular tube formation by enriching the exocyst at the lumenal membrane. We show that the PAR polarity proteins PAR-6 and PKC-3/aPKC localize to the lumenal membrane domain and function within the excretory cell to promote lumen extension, similar to exocyst component SEC-5 and exocyst regulator RAL-1. Using acute protein depletion, we find that PAR-6 is required to recruit the exocyst to the lumenal membrane domain, whereas PAR-3, which functions as an exocyst receptor in mammalian cells, appears to be dispensable for exocyst localization and lumen extension. Finally, we show that the Rho GTPase CDC-42 and the RhoGEF EXC-5/FGD act as upstream regulators of lumen formation by recruiting PAR-6 and PKC-3 to the lumenal membrane. Our findings reveal a molecular pathway that connects Rho GTPase signaling, cell polarity, and vesicle-tethering proteins to promote lumen extension in intracellular tubes.

scholarly journals The WD40-repeat protein WDR-48 promotes the stability of the deubiquitinating enzyme USP-46 by inhibiting its ubiquitination and degradation

2020 ◽  
Vol 295 (33) ◽  
pp. 11776-11788
Author(s):  
Molly Hodul ◽  
Rakesh Ganji ◽  
Caroline L. Dahlberg ◽  
Malavika Raman ◽  
Peter Juo
Keyword(s):  
Mammalian Cells ◽  
Deubiquitinating Enzymes ◽  
Post Translational Modification ◽  
Wd40 Repeat ◽  
C Elegans ◽  
Protein Levels ◽  
Specific Protease ◽  
The Stability ◽  
And Function

Ubiquitination is a reversible post-translational modification that has emerged as a critical regulator of synapse development and function. However, the mechanisms that regulate the deubiquitinating enzymes (DUBs) responsible for the removal of ubiquitin from target proteins are poorly understood. We have previously shown that the DUB ubiquitin-specific protease 46 (USP-46) removes ubiquitin from the glutamate receptor GLR-1 and regulates its trafficking and degradation in Caenorhabditis elegans. We found that the WD40-repeat proteins WDR-20 and WDR-48 bind and stimulate the catalytic activity of USP-46. Here, we identified another mechanism by which WDR-48 regulates USP-46. We found that increased expression of WDR-48, but not WDR-20, promotes USP-46 abundance in mammalian cells in culture and in C. elegans neurons in vivo. Inhibition of the proteasome increased USP-46 abundance, and this effect was nonadditive with increased WDR-48 expression. We found that USP-46 is ubiquitinated and that expression of WDR-48 reduces the levels of ubiquitin–USP-46 conjugates and increases the t1/2 of USP-46. A point-mutated WDR-48 variant that disrupts binding to USP-46 was unable to promote USP-46 abundance in vivo. Finally, siRNA-mediated knockdown of wdr48 destabilizes USP46 in mammalian cells. Together, these results support a model in which WDR-48 binds and stabilizes USP-46 protein levels by preventing the ubiquitination and degradation of USP-46 in the proteasome. Given that a large number of USPs interact with WDR proteins, we propose that stabilization of DUBs by their interacting WDR proteins may be a conserved and widely used mechanism that controls DUB availability and function.

scholarly journals Interpreting the pathogenicity of Joubert Syndrome missense variants in Caenorhabditis elegans

2020 ◽  
pp. dmm.046631
Author(s):  
Karen I. Lange ◽  
Sofia Tsiropoulou ◽  
Katarzyna Kucharska ◽  
Oliver E. Blacque
Keyword(s):  
Caenorhabditis Elegans ◽  
Primary Cilia ◽  
Model Organism ◽  
Joubert Syndrome ◽  
Compound Heterozygous ◽  
Inherited Disorders ◽  
Missense Variants ◽  
C Elegans ◽  
Variant Alleles ◽  
And Function

Ciliopathies are inherited disorders caused by defects in motile and non-motile (primary) cilia. Ciliopathy syndromes and associated gene variants are often highly pleiotropic and represent exemplars for interrogating genotype-phenotype correlations. Towards understanding disease mechanisms in the context of ciliopathy mutations, we have employed a leading model organism for cilia and ciliopathy research, Caenorhabditis elegans, together with gene editing, to characterise two missense variants (P74S, G155S) in B9D2/mksr-2 associated with Joubert Syndrome (JBTS). B9D2 functions within the Meckel syndrome (MKS) module at the ciliary base transition zone (TZ) compartment, and regulates the cilium's molecular composition and sensory/signaling functions. Quantitative assays of cilium/TZ structure and function, together with knock-in reporters, confirm both variant alleles are pathogenic in worms. G155S causes a more severe overall phenotype and disrupts endogenous MKSR-2 organisation at the TZ. Recapitulation of the patient biallelic genotype shows that compound heterozygous worms phenocopy worms homozygous for P74S. The P74S and G155S alleles also reveal evidence of a very close functional association between the B9D2-associated B9 complex and TMEM216/MKS-2. Together, these data establish C. elegans as a paradigm for interpreting JBTS mutations, and provide further insight into MKS module organisation.

Hepatocyte growth factor regulated tyrosine kinase substrate in the peripheral development and function of B-cells

2014 ◽  
Vol 443 (2) ◽  
pp. 351-356 ◽  
Author(s):  
Takayuki Nagata ◽  
Kazuko Murata ◽  
Ryo Murata ◽  
Shu-lan Sun ◽  
Yutaro Saito ◽  
...  
Keyword(s):  
B Cells ◽  
Growth Factor ◽  
Hepatocyte Growth Factor ◽  
Tyrosine Kinase ◽  
Kinase Substrate ◽  
And Function ◽  
Hepatocyte Growth ◽  
Tyrosine Kinase Substrate
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