scholarly journals RABL2 interacts with the intraflagellar transport-B complex and CEP19 and participates in ciliary assembly

2017 ◽  
Vol 28 (12) ◽  
pp. 1652-1666 ◽  
Author(s):  
Yuya Nishijima ◽  
Yohei Hagiya ◽  
Tomohiro Kubo ◽  
Ryota Takei ◽  
Yohei Katoh ◽  
...  
Keyword(s):  
Basal Body ◽  
Intraflagellar Transport ◽  
Bound State ◽  
Dependent Manner ◽  
Centrosomal Protein ◽  
Mother Centriole ◽  
Flagellar Assembly ◽  
Exogenous Expression ◽  
And Function

Proteins localized to the basal body and the centrosome play crucial roles in ciliary assembly and function. Although RABL2 and CEP19 are conserved in ciliated organisms and have been implicated in ciliary/flagellar functions, their roles are poorly understood. Here we show that RABL2 interacts with CEP19 and is recruited to the mother centriole and basal body in a CEP19-dependent manner and that CEP19 is recruited to the centriole probably via its binding to the centrosomal protein FGFR1OP. Disruption of the RABL2 gene in Chlamydomonas reinhardtii results in the nonflagellated phenotype, suggesting a crucial role of RABL2 in ciliary/flagellar assembly. We also show that RABL2 interacts, in its GTP-bound state, with the intraflagellar transport (IFT)-B complex via the IFT74–IFT81 heterodimer and that the interaction is disrupted by a mutation found in male infertile mice (Mot mice) with a sperm flagella motility defect. Intriguingly, RABL2 binds to CEP19 and the IFT74–IFT81 heterodimer in a mutually exclusive manner. Furthermore, exogenous expression of the GDP-locked or Mot-type RABL2 mutant in human cells results in mild defects in ciliary assembly. These results indicate that RABL2 localized to the basal body plays crucial roles in ciliary/flagellar assembly via its interaction with the IFT-B complex.

scholarly journals The role of retrograde intraflagellar transport in flagellar assembly, maintenance, and function

2012 ◽  
Vol 199 (1) ◽  
pp. 151-167 ◽  
Author(s):  
Benjamin D. Engel ◽  
Hiroaki Ishikawa ◽  
Kimberly A. Wemmer ◽  
Stefan Geimer ◽  
Ken-ichi Wakabayashi ◽  
...  
Keyword(s):  
Intraflagellar Transport ◽  
Retrograde Transport ◽  
Full Length ◽  
Temperature Sensitive ◽  
Nonpermissive Temperature ◽  
Flagellar Beat ◽  
Flagellar Assembly ◽  
Ph Shock ◽  
And Function

The maintenance of flagellar length is believed to require both anterograde and retrograde intraflagellar transport (IFT). However, it is difficult to uncouple the functions of retrograde transport from anterograde, as null mutants in dynein heavy chain 1b (DHC1b) have stumpy flagella, demonstrating solely that retrograde IFT is required for flagellar assembly. We isolated a Chlamydomonas reinhardtii mutant (dhc1b-3) with a temperature-sensitive defect in DHC1b, enabling inducible inhibition of retrograde IFT in full-length flagella. Although dhc1b-3 flagella at the nonpermissive temperature (34°C) showed a dramatic reduction of retrograde IFT, they remained nearly full-length for many hours. However, dhc1b-3 cells at 34°C had strong defects in flagellar assembly after cell division or pH shock. Furthermore, dhc1b-3 cells displayed altered phototaxis and flagellar beat. Thus, robust retrograde IFT is required for flagellar assembly and function but is dispensable for the maintenance of flagellar length. Proteomic analysis of dhc1b-3 flagella revealed distinct classes of proteins that change in abundance when retrograde IFT is inhibited.

scholarly journals The role of the dynein light intermediate chain in retrograde IFT and flagellar function in Chlamydomonas

2016 ◽  
Vol 27 (15) ◽  
pp. 2404-2422 ◽  
Author(s):  
Jaimee Reck ◽  
Alexandria M. Schauer ◽  
Kristyn VanderWaal Mills ◽  
Raqual Bower ◽  
Douglas Tritschler ◽  
...  
Keyword(s):  
Intraflagellar Transport ◽  
Small Subset ◽  
Dependent Manner ◽  
Microtubule Motor ◽  
Flagellar Assembly ◽  
Cilia And Flagella ◽  
The Stability ◽  
Mutant Phenotypes ◽  
Intermediate Chain

The assembly of cilia and flagella depends on the activity of two microtubule motor complexes, kinesin-2 and dynein-2/1b, but the specific functions of the different subunits are poorly defined. Here we analyze Chlamydomonas strains expressing different amounts of the dynein 1b light intermediate chain (D1bLIC). Disruption of D1bLIC alters the stability of the dynein 1b complex and reduces both the frequency and velocity of retrograde intraflagellar transport (IFT), but it does not eliminate retrograde IFT. Flagellar assembly, motility, gliding, and mating are altered in a dose-dependent manner. iTRAQ-based proteomics identifies a small subset of proteins that are significantly reduced or elevated in d1blic flagella. Transformation with D1bLIC-GFP rescues the mutant phenotypes, and D1bLIC-GFP assembles into the dynein 1b complex at wild-type levels. D1bLIC-GFP is transported with anterograde IFT particles to the flagellar tip, dissociates into smaller particles, and begins processive retrograde IFT in <2 s. These studies demonstrate the role of D1bLIC in facilitating the recycling of IFT subunits and other proteins, identify new components potentially involved in the regulation of IFT, flagellar assembly, and flagellar signaling, and provide insight into the role of D1bLIC and retrograde IFT in other organisms.

The transmembrane adaptor protein NTAL limits mast cell chemotaxis toward prostaglandin E2

2018 ◽  
Vol 11 (556) ◽  
pp. eaao4354 ◽  
Author(s):  
Ivana Halova ◽  
Monika Bambouskova ◽  
Lubica Draberova ◽  
Viktor Bugajev ◽  
Petr Draber
Keyword(s):  
Mast Cell ◽  
Mast Cells ◽  
T Cell Activation ◽  
Cell Activation ◽  
Adaptor Protein ◽  
Prostaglandin E ◽  
Dependent Manner ◽  
And Function ◽  
Cell Chemotaxis

Chemotaxis of mast cells is one of the crucial steps in their development and function. Non–T cell activation linker (NTAL) is a transmembrane adaptor protein that inhibits the activation of mast cells and B cells in a phosphorylation-dependent manner. Here, we studied the role of NTAL in the migration of mouse mast cells stimulated by prostaglandin E2 (PGE2). Although PGE2 does not induce the tyrosine phosphorylation of NTAL, unlike IgE immune complex antigens, we found that loss of NTAL increased the chemotaxis of mast cells toward PGE2. Stimulation of mast cells that lacked NTAL with PGE2 enhanced the phosphorylation of AKT and the production of phosphatidylinositol 3,4,5-trisphosphate. In resting NTAL-deficient mast cells, phosphorylation of an inhibitory threonine in ERM family proteins accompanied increased activation of β1-containing integrins, which are features often associated with increased invasiveness in tumors. Rescue experiments indicated that only full-length, wild-type NTAL restored the chemotaxis of NTAL-deficient cells toward PGE2. Together, these data suggest that NTAL is a key inhibitor of mast cell chemotaxis toward PGE2, which may act through the RHOA/ERM/β1-integrin and PI3K/AKT axes.

scholarly journals TRPV channel OCR-2 is distributed along C. elegans chemosensory cilia by diffusion in a local interplay with intraflagellar transport

2020 ◽  
Author(s):  
Jaap van Krugten ◽  
Noémie Danné ◽  
Erwin J.G. Peterman
Keyword(s):  
Single Molecule ◽  
Transmembrane Proteins ◽  
Intraflagellar Transport ◽  
Single Molecule Tracking ◽  
C Elegans ◽  
Normal Diffusion ◽  
Cellular Signal ◽  
Underlying Mechanisms ◽  
And Function

AbstractSensing and reacting to the environment is essential for survival and procreation of most organisms. Caenorhabditis elegans senses soluble chemicals with transmembrane proteins (TPs) in the cilia of its chemosensory neurons. Development, maintenance and function of these cilia relies on intraflagellar transport (IFT), in which motor proteins transport cargo, including sensory TPs, back and forth along the ciliary axoneme. Here we use live fluorescence imaging to show that IFT machinery and the sensory TP OCR-2 reversibly redistribute along the cilium after exposure to repellant chemicals. To elucidate the underlying mechanisms, we performed single-molecule tracking experiments and found that OCR-2 distribution depends on an intricate interplay between IFT-driven transport, normal diffusion and subdiffusion that depends on the specific location in the cilium. These insights in the role of IFT on the dynamics of cellular signal transduction contribute to a deeper understanding of the regulation of sensory TPs and chemosensing.

scholarly journals microRNAs Biogenesis, Functions and Role in Tumor Angiogenesis

2020 ◽  
Vol 10 ◽  
Author(s):  
Tiziana Annese ◽  
Roberto Tamma ◽  
Michelina De Giorgis ◽  
Domenico Ribatti
Keyword(s):  
Tumor Angiogenesis ◽  
Vasculogenic Mimicry ◽  
Mirna Biogenesis ◽  
Dependent Manner ◽  
Tumor Initiation ◽  
Rna Molecules ◽  
Angiogenic Switch ◽  
Sprouting Angiogenesis ◽  
And Function

microRNAs (miRNAs) are small non-coding RNA molecules, evolutionary conserved. They target more than one mRNAs, thus influencing multiple molecular pathways, but also mRNAs may bind to a variety of miRNAs, either simultaneously or in a context-dependent manner. miRNAs biogenesis, including miRNA transcription, processing by Drosha and Dicer, transportation, RISC biding, and miRNA decay, are finely controlled in space and time.miRNAs are critical regulators in various biological processes, such as differentiation, proliferation, apoptosis, and development in both health and disease. Their dysregulation is involved in tumor initiation and progression. In tumors, they can act as onco-miRNAs or oncosuppressor-miRNA participating in distinct cellular pathways, and the same miRNA can perform both activities depending on the context.In tumor progression, the angiogenic switch is fundamental. miRNAs derived from tumor cells, endothelial cells, and cells of the surrounding microenvironment regulate tumor angiogenesis, acting as pro-angiomiR or anti-angiomiR.In this review, we described miRNA biogenesis and function, and we update the non-classical aspects of them. The most recent role in the nucleus, as transcriptional gene regulators and the different mechanisms by which they could be dysregulated, in tumor initiation and progression, are treated. In particular, we describe the role of miRNAs in sprouting angiogenesis, vessel co-option, and vasculogenic mimicry. The role of miRNAs in lymphoma angiogenesis is also discussed despite the scarcity of data.The information presented in this review reveals the need to do much more to discover the complete miRNA network regulating angiogenesis, not only using high-throughput computational analysis approaches but also morphological ones.

scholarly journals OsJAZ13 Negatively Regulates Jasmonate Signaling and Activates Hypersensitive Cell Death Response in Rice

2020 ◽  
Vol 21 (12) ◽  
pp. 4379
Author(s):  
Xiujing Feng ◽  
Lei Zhang ◽  
Xiaoli Wei ◽  
Yun Zhou ◽  
Yan Dai ◽  
...  
Keyword(s):  
Cell Death ◽  
Splice Variants ◽  
Adaptor Protein ◽  
Dependent Manner ◽  
Hypersensitive Cell Death ◽  
Cell Death Response ◽  
Jaz Proteins ◽  
Localization Analysis ◽  
And Function

Jasmonate ZIM-domain (JAZ) proteins belong to the subgroup of TIFY family and act as key regulators of jasmonate (JA) responses in plants. To date, only a few JAZ proteins have been characterized in rice. Here, we report the identification and function of rice OsJAZ13 gene. The gene encodes three different splice variants: OsJAZ13a, OsJAZ13b, and OsJAZ13c. The expression of OsJAZ13 was mainly activated in vegetative tissues and transiently responded to JA and ethylene. Subcellular localization analysis indicated OsJAZ13a is a nuclear protein. Yeast two-hybrid assays revealed OsJAZ13a directly interacts with OsMYC2, and also with OsCOI1, in a COR-dependent manner. Furthermore, OsJAZ13a recruited a general co-repressor OsTPL via an adaptor protein OsNINJA. Remarkably, overexpression of OsJAZ13a resulted in the attenuation of root by methyl JA. Furthermore, OsJAZ13a-overexpressing plants developed lesion mimics in the sheath after approximately 30–45 days of growth. Tillers with necrosis died a few days later. Gene-expression analysis suggested the role of OsJAZ13 in modulating the expression of JA/ethylene response-related genes to regulate growth and activate hypersensitive cell death. Taken together, these observations describe a novel regulatory mechanism in rice and provide the basis for elucidating the function of OsJAZ13 in signal transduction and cell death in plants.

PKCϵ has an alcohol-binding site in its second cysteine-rich regulatory domain

2009 ◽  
Vol 421 (3) ◽  
pp. 405-413 ◽  
Author(s):  
Joydip Das ◽  
Satyabrata Pany ◽  
Ghazi M. Rahman ◽  
Simon J. Slater
Keyword(s):  
Protein Kinase ◽  
Binding Site ◽  
Maximum Effect ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Kinase C ◽  
C1 Domain ◽  
Pkc Protein Kinase C ◽  
And Function

Alcohols regulate the expression and function of PKC (protein kinase C), and it has been proposed that an alcohol-binding site is present in PKCα in its C1 domain, which consists of two cysteine-rich subdomains, C1A and C1B. A PKCϵ-knockout mouse showed a significant decrease in alcohol consumption compared with the wild-type. The aim of the present study was to investigate whether an alcohol-binding site could be present in PKCϵ. Here we show that ethanol inhibited PKCϵ activity in a concentration-dependent manner with an EC50 (equilibrium ligand concentration at half-maximum effect) of 43 mM. Ethanol, butanol and octanol increased the binding affinity of a fluorescent phorbol ester SAPD (sapintoxin-D) to PKCϵC1B in a concentration-dependent manner with EC50 values of 78 mM, 8 mM and 340 μM respectively, suggesting the presence of an allosteric alcohol-binding site in this subdomain. To identify this site, PKCϵC1B was photolabelled with 3-azibutanol and 3-azioctanol and analysed by MS. Whereas azibutanol preferentially labelled His236, Tyr238 was the preferred site for azioctanol. Inspection of the model structure of PKCϵC1B reveals that these residues are 3.46 Å (1 Å=0.1 nm) apart from each other and form a groove where His236 is surface-exposed and Tyr238 is buried inside. When these residues were replaced by alanine, it significantly decreased alcohol binding in terms of both photolabelling and alcohol-induced SAPD binding in the mutant H236A/Y238A. Whereas Tyr238 was labelled in mutant H236A, His236 was labelled in mutant Y238A. The present results provide direct evidence for the presence of an allosteric alcohol-binding site on protein kinase Cϵ and underscore the role of His236 and Tyr238 residues in alcohol binding.

scholarly journals Chlamydomonas Basal Bodies as Flagella Organizing Centers

Cells ◽  
2018 ◽  
Vol 7 (7) ◽  
pp. 79 ◽  
Author(s):  
Jenna Wingfield ◽  
Karl-Ferdinand Lechtreck
Keyword(s):  
Structure Function ◽  
Chlamydomonas Reinhardtii ◽  
Developmental Disorders ◽  
Intraflagellar Transport ◽  
Specific Protein ◽  
Basal Bodies ◽  
Unicellular Green Alga ◽  
Flagellar Assembly ◽  
Flagellar Axoneme

During ciliogenesis, centrioles convert to membrane-docked basal bodies, which initiate the formation of cilia/flagella and template the nine doublet microtubules of the flagellar axoneme. The discovery that many human diseases and developmental disorders result from defects in flagella has fueled a strong interest in the analysis of flagellar assembly. Here, we will review the structure, function, and development of basal bodies in the unicellular green alga Chlamydomonas reinhardtii, a widely used model for the analysis of basal bodies and flagella. Intraflagellar transport (IFT), a flagella-specific protein shuttle critical for ciliogenesis, was first described in C. reinhardtii. A focus of this review will be on the role of the basal bodies in organizing the IFT machinery.

HSP27 Protects Malignant Hematopoietic Cells Against VP-16-Induced Apoptosis through Modulation of P38 and C-JUN.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 1276-1276
Author(s):  
Hein Schepers ◽  
Marjan Geugien ◽  
Marco van der Toorn ◽  
Anton L. Bryantsev ◽  
Harm H. Kampinga ◽  
...  
Keyword(s):  
Hematopoietic Cells ◽  
Leukemic Cells ◽  
Dependent Manner ◽  
Apoptosis Pathway ◽  
Hsp27 Expression ◽  
Cd34 Cells ◽  
Induced Apoptosis ◽  
And Function ◽  
Jnk Activation

Abstract In the present study, expression and function of Heat Shock Protein 27 (HSP27) was analyzed in acute myeloid leukemia (AML), since HSP27 expression is linked to unfavourable prognosis. HSP27 protein was predominantly expressed in monocytic blasts (M4-M5, 91%, N = 11) and absent in myeloid leukemic blasts (M1-M2, N = 5). A similar lineage restricted expression was observed in normal hematopoietic cells: high expression in normal CD34+ cells and monocytes, and absent in granulocytes. To study the functional role of HSP27, RNA interference (RNAi) studies were performed in the leukemic TF-1 cell line. These experiments demonstrated a twofold increase in VP-16-induced apoptosis after HSP27 siRNA. In contrast, CD95 Fas-induced apoptosis remained the same, as a result of CD95 Fas-mediated upregulation of HSP27. Additional investigations demonstrated that the increased VP-16-induced apoptosis after HSP27 RNAi, was associated with an enhanced phosphorylation of p38 and c-Jun. This VP-16-induced phosphorylation was subsequently followed by the release of cytochrome c into the cytoplasm, which increased twofold after siRNA treatment. These results indicate that HSP27 plays an important role in the protection against VP-16-induced apoptosis through the modulation of p38 and JNK activation, probably through interference with DAXX-mediated ASK1 activation. This was further underscored by co-immunoprecipitation studies, demonstrating complex formation of DAXX and HSP27 in an ASK1-dependent manner. However, in the investigated AML samples, VP-16-mediated apoptosis correlated moderately with HSP27 expression, although HSP27 was highly expressed and phosphorylated and activated in primitive monocytic AML blasts. This is likely due to the co-expression of p21Waf1/Cip1, which is in the majority of the monocytic AML M4-M5 blasts constitutively localised in the cytoplasm and interferes with apoptosis via the DAXX-ASK1-dependent pathway. Preliminary data indicate that overexpression of a cytoplasmic form of p21 is able to reduce the VP-16-induced apoptosis after RNAi for HSP27 as compared to controls, suggesting a predominant anti-apoptotic role of p21 over HSP27. In summary, we demonstrate a role for HSP27 in the survival of leukemic cells by modulation of the DAXX/p38/JNK apoptosis pathway. This survival advantage can further be promoted by the co-expression of cytoplasmic localised p21Waf1/Cip1 protein, indicating that strategies in AML treatment should be focused on targeting multiple signal transduction pathways.

scholarly journals Caenorhabditis elegans DYF-2, an Orthologue of Human WDR19, Is a Component of the Intraflagellar Transport Machinery in Sensory Cilia

2006 ◽  
Vol 17 (11) ◽  
pp. 4801-4811 ◽  
Author(s):  
Evgeni Efimenko ◽  
Oliver E. Blacque ◽  
Guangshuo Ou ◽  
Courtney J. Haycraft ◽  
Bradley K. Yoder ◽  
...  
Keyword(s):  
Caenorhabditis Elegans ◽  
Aspartic Acid ◽  
Critical Role ◽  
Intraflagellar Transport ◽  
Bardet Biedl Syndrome ◽  
Evolutionarily Conserved ◽  
Sensory Cilia ◽  
Mutant Phenotypes ◽  
And Function

The intraflagellar transport (IFT) machinery required to build functional cilia consists of a multisubunit complex whose molecular composition, organization, and function are poorly understood. Here, we describe a novel tryptophan-aspartic acid (WD) repeat (WDR) containing IFT protein from Caenorhabditis elegans, DYF-2, that plays a critical role in maintaining the structural and functional integrity of the IFT machinery. We determined the identity of the dyf-2 gene by transgenic rescue of mutant phenotypes and by sequencing of mutant alleles. Loss of DYF-2 function selectively affects the assembly and motility of different IFT components and leads to defects in cilia structure and chemosensation in the nematode. Based on these observations, and the analysis of DYF-2 movement in a Bardet–Biedl syndrome mutant with partially disrupted IFT particles, we conclude that DYF-2 can associate with IFT particle complex B. At the same time, mutations in dyf-2 can interfere with the function of complex A components, suggesting an important role of this protein in the assembly of the IFT particle as a whole. Importantly, the mouse orthologue of DYF-2, WDR19, also localizes to cilia, pointing to an important evolutionarily conserved role for this WDR protein in cilia development and function.

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