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Cell Biology
A novel physiological role for ARF1 in the formation of bi-directional tubules from the Golgi
Apr 21, 2017   Molecular Biology Of The Cell
Bottanelli F, Kilian N, Ernst AM, Rivera-Molina F, Schroeder LK, Kromann EB, Lessard MD, Erdmann RS, Schepartz A, Baddeley D, Bewersdorf J, Toomre D, Rothman JE
A novel physiological role for ARF1 in the formation of bi-directional tubules from the Golgi
Apr 21, 2017
Molecular Biology Of The Cell
Capitalizing on CRISPR/Cas9 gene editing techniques and super-resolution nanoscopy, we have explored the role of the small GTPase ARF1 in mediating transport steps at the Golgi. Surprisingly, beside its well-established role in generating COPI vesicles, we find that ARF1 is additionally involved in the formation of long (∼ 3 μm), thin (∼ 110 nm diameter) tubular carriers. The anterograde and retrograde tubular carriers are both largely free of the classical Golgi coat proteins Coatomer (COPI) and Clathrin. Instead, they contain ARF1 along their entire length at a density estimated to be in the range of close packing. Experiments using a mutant form of ARF1 affecting GTP hydrolysis suggest that ARF1[GTP] is functionally required for the tubules to form. Dynamic confocal and stimulated emission depletion (STED) imaging revealed that ARF1-rich tubular compartments fall into two distinct classes containing [either]: i) anterograde cargoes and Clathrin clusters or ii) retrograde cargoes and Coatomer clusters. © 2017 by The American Society for Cell Biology.
Gestational Diabetes alters functions in offspring's umbilical cord cells with implications for cardiovascular health
Apr 21, 2017   Endocrinology
Amrithraj AI, Kodali A, Nguyen L, Teo AKK, Chang CW, Karnani N, Ng KL, Gluckman PD, Chong YS, Stünkel W
Gestational Diabetes alters functions in offspring's umbilical cord cells with implications for cardiovascular health
Apr 21, 2017
Endocrinology
As non-communicable diseases such as type 2 diabetes mellitus have their roots in prenatal development and conditions such as maternal gestational diabetes (GDM), we aimed to test this hypothesis in primary cells derived from the offspring of GDM mothers compared to control subjects. We have assessed primary umbilical cord derived cells such as human umbilical vein endothelial cells (HUVECs) and Wharton's jelly derived mesenchymal stem cells (WJMSCs) from both, the offspring of GDM and non-GDM mothers. We have compared the primary isolates in cell based assays measuring proliferation, mitochondrial oxygen consumption, and the ability to support blood vessel growth. We conducted gene expression microarray studies with subsequent pathway analysis and candidate gene validation. We observed striking differences between the two groups such as lower metabolic rates and impairment of endothelial tube formation in cells with GDM background. HUVECs from subjects with maternal GDM have lower expression of the anti-apoptotic protein BCL-xL suggesting compromised angiogenic capabilities. Comparative gene expression analysis revealed blood vessel formation as a major pathway enriched in the GDM-derived HUVECs with the surface marker CD44 as a significant gene under-expressed in the GDM group. Functional validation of CD44 revealed that it regulates tube formation in HUVECs thereby providing new insights into a novel pathway imprinted in primary umbilical cord-derived cells from GDM offspring. Our data demonstrate that primary cells isolated from the umbilical cord of offspring born to GDM mothers maintain metabolic and molecular imprints of maternal hyperglycemia reflecting an enhanced risk for cardiovascular disease later in life. Copyright © 2017 Endocrine Society.
Gates for soluble and membrane proteins, and two trafficking systems (IFT and LIFT), establish a dynamic ciliary signaling compartment
Apr 22, 2017   Current Opinion In Cell Biology
Jensen VL, Leroux MR
Gates for soluble and membrane proteins, and two trafficking systems (IFT and LIFT), establish a dynamic ciliary signaling compartment
Apr 22, 2017
Current Opinion In Cell Biology
Primary cilia are microtubule-based organelles found on most mammalian cell surfaces. They possess a soluble matrix and membrane contiguous with the cell body cytosol and plasma membrane, and yet, have distinct compositions that can be modulated to enable dynamic signal transduction. Here, we discuss how specialized ciliary compartments are established using a coordinated network of gating, trafficking and targeting activities. Cilium homeostasis is maintained by a size-selective molecular mesh that limits soluble protein entry, and by a membrane diffusion barrier localized at the transition zone. Bidirectional protein shuttling between the cell body and cilium uses IntraFlagellar Transport (IFT), and prenylated ciliary protein delivery is achieved through Lipidated protein IntraFlagellar Targeting (LIFT). Elucidating how these gates and transport systems function will help reveal the roles that cilia play in ciliary signaling and the growing spectrum of disorders termed ciliopathies. Copyright © 2017 Elsevier Ltd. All rights reserved.
Presynaptic morphology and vesicular composition determine vesicle dynamics in mouse central synapses
Apr 22, 2017   ELife
Guillaud L, Dimitrov D, Takahashi T
Presynaptic morphology and vesicular composition determine vesicle dynamics in mouse central synapses
Apr 22, 2017
ELife
Transport of synaptic vesicles (SVs) in nerve terminals is thought to play essential roles in maintenance of neurotransmission. To identify factors modulating SV movements, we performed real-time imaging analysis of fluorescently labeled SVs in giant calyceal and conventional hippocampal terminals. Compared with small hippocampal terminals, SV movements in giant calyceal terminals were faster, longer and kinetically more heterogeneous. Morphological maturation of giant calyceal terminals was associated with an overall reduction in SV mobility and displacement heterogeneity. At the molecular level, SVs over-expressing vesicular glutamate transporter 1 (VGLUT1) showed higher mobility than VGLUT2-expressing SVs. Pharmacological disruption of the presynaptic microtubule network preferentially reduced long directional movements of SVs between release sites. Functionally, synaptic stimulation appeared to recruit SVs to active zones without significantly altering their mobility. Hence, the morphological features of nerve terminals and the molecular signature of vesicles are key elements determining vesicular dynamics and movements in central synapses.
Phosphorylation of iRhom2 at the plasma membrane controls mammalian TACE-dependent inflammatory and growth factor signalling
Apr 22, 2017   ELife
Grieve AG, Xu H, Künzel U, Bambrough P, Sieber B, Freeman M
Phosphorylation of iRhom2 at the plasma membrane controls mammalian TACE-dependent inflammatory and growth factor signalling
Apr 22, 2017
ELife
Proteolytic cleavage and release from the cell surface of membrane-tethered ligands is an important mechanism of regulating intercellular signalling. TACE is a major shedding protease, responsible for the liberation of the inflammatory cytokine TNFα and ligands of the epidermal growth factor receptor. iRhoms, catalytically inactive members of the rhomboid-like superfamily, have been shown to control the ER-to-Golgi transport and maturation of TACE. Here, we reveal that iRhom2 remains associated with TACE throughout the secretory pathway, and is stabilised at the cell surface by this interaction. At the plasma membrane, ERK1/2-mediated phosphorylation and 14-3-3 protein binding of the cytoplasmic amino-terminus of iRhom2 alter its interaction with mature TACE, thereby licensing its proteolytic activity. We show that this molecular mechanism is responsible for triggering inflammatory responses in primary mouse macrophages. Overall, iRhom2 binds to TACE throughout its lifecycle, implying that iRhom2 is a primary regulator of stimulated cytokine and growth factor signalling.
Structure-function analysis of the DNA-binding domain of a transmembrane transcriptional activator
Apr 22, 2017   Scientific Reports
Schlundt A, Buchner S, Janowski R, Heydenreich T, Heermann R, Lassak J, Geerlof A, Stehle R, Niessing D, Jung K, Sattler M
Structure-function analysis of the DNA-binding domain of a transmembrane transcriptional activator
Apr 22, 2017
Scientific Reports
The transmembrane DNA-binding protein CadC of E. coli, a representative of the ToxR-like receptor family, combines input and effector domains for signal sensing and transcriptional activation, respectively, in a single protein, thus representing one of the simplest signalling systems. At acidic pH in a lysine-rich environment, CadC activates the transcription of the cadBA operon through recruitment of the RNA polymerase (RNAP) to the two cadBA promoter sites, Cad1 and Cad2, which are directly bound by CadC. However, the molecular details for its interaction with DNA have remained elusive. Here, we present the crystal structure of the CadC DNA-binding domain (DBD) and show that it adopts a winged helix-turn-helix fold. The interaction with the cadBA promoter site Cad1 is studied by using nuclear magnetic resonance (NMR) spectroscopy, biophysical methods and functional assays and reveals a preference for AT-rich regions. By mutational analysis we identify amino acids within the CadC DBD that are crucial for DNA-binding and functional activity. Experimentally derived structural models of the CadC-DNA complex indicate that the CadC DBD employs mainly non-sequence-specific over a few specific contacts. Our data provide molecular insights into the CadC-DNA interaction and suggest how CadC dimerization may provide high-affinity binding to the Cad1 promoter.
Systematic Synergy of Glucose and GLP-1 to Stimulate Insulin Secretion Revealed by Quantitative Phosphoproteomics
Apr 22, 2017   Scientific Reports
Tang JS, Li QR, Li JM, Wu JR, Zeng R
Systematic Synergy of Glucose and GLP-1 to Stimulate Insulin Secretion Revealed by Quantitative Phosphoproteomics
Apr 22, 2017
Scientific Reports
GLP-1 synergizes with glucose in regulating pancreatic β-cell function, including facilitating β-cell survival and insulin secretion. Though it has been widely accepted that phosphorylation is extremely important in regulating β-cell functions, our knowledge to the global mechanism is still limited. Here we performed a quantitative phosphoproteomics study to systematically present the synergistic regulation of INS-1E cell phosphoproteome mediated by glucose and GLP-1. We generated the largest pancreatic β-cell phosphoproteome by identifying 25,327 accurately localized phosphorylation sites on 5,389 proteins. Our results discovered several novel kinases regulated by glucose, GLP-1 or their synergism, and some of these kinases might act as downstream molecules of GLP-1 mediated PKA signaling cascade. A few phosphosites were regulated by both GLP-1 and glucose alone, and these target proteins were highly related to their biological function on pancreatic β-cells. Finally, we found glucose and GLP-1 executed their synergistic effect at multiple levels, especially at pathway level. Both GLP-1 and glucose participated in regulating every single step of the secretion pathway, and systematically synergized their effects in inducing insulin secretion.
Quiescence of adult oligodendrocyte precursor cells requires thyroid hormone and hypoxia to activate Runx1
Apr 22, 2017   Scientific Reports
Tokumoto Y, Tamaki S, Kabe Y, Takubo K, Suematsu M
Quiescence of adult oligodendrocyte precursor cells requires thyroid hormone and hypoxia to activate Runx1
Apr 22, 2017
Scientific Reports
The adult mammalian central nervous system (CNS) contains a population of slowly dividing oligodendrocyte precursor cells (OPCs), i.e., adult OPCs, which supply new oligodendrocytes throughout the life of animal. While adult OPCs develop from rapidly dividing perinatal OPCs, the mechanisms underlying their quiescence remain unknown. Here, we show that perinatal rodent OPCs cultured with thyroid hormone (TH) under hypoxia become quiescent and acquire adult OPCs-like characteristics. The cyclin-dependent kinase inhibitor p15/INK4b plays crucial roles in the TH-dependent cell cycle deceleration in OPCs under hypoxia. Klf9 is a direct target of TH-dependent signaling. Under hypoxic conditions, hypoxia-inducible factors mediates runt-related transcription factor 1 activity to induce G1 arrest in OPCs through enhancing TH-dependent p15/INK4b expression. As adult OPCs display phenotypes of adult somatic stem cells in the CNS, the current results shed light on environmental requirements for the quiescence of adult somatic stem cells during their development from actively proliferating stem/progenitor cells.
RARβ2 is required for vertebrate somitogenesis
Apr 22, 2017   Development (Cambridge, England)
Janesick A, Tang W, Nguyen TTL, Blumberg B
RARβ2 is required for vertebrate somitogenesis
Apr 22, 2017
Development (Cambridge, England)
During vertebrate somitogenesis, retinoic acid is known to establish the position of the determination wavefront, controlling where new somites are permitted to form along the anteroposterior body axis. Less is understood about how RAR regulates somite patterning, rostral-caudal boundary setting, specialization of myotome subdivisions, or the specific RAR subtype that is required for somite patterning. Characterizing the function of RARβ has been challenging due to the absence of embryonic phenotypes in murine loss-of-function studies. Using the Xenopus system, we show that RARβ2 plays a specific role in somite number and size, restriction of the presomitic mesoderm anterior border, somite chevron morphology and hypaxial myoblast migration. Rarβ2 is the RAR subtype whose expression is most up-regulated in response to ligand and its localization in the trunk somites positions it at the right time and place to respond to embryonic retinoid levels during somitogenesis. RARβ2 positively regulates Tbx3 a marker of hypaxial muscle, and negatively regulates Tbx6 via Ripply2 to restrict the anterior boundaries of the presomitic mesoderm and caudal progenitor pool. These results demonstrate for the first time an early and essential role for RARβ2 in vertebrate somitogenesis. © 2017. Published by The Company of Biologists Ltd.
Ventricular assist device elicits serum natural IgG that correlates with the development of primary graft dysfunction following heart transplantation
Apr 22, 2017   The Journal Of Heart And Lung Transplantation : The Official Publication Of The International Society For Heart Transplantation
See SB, Clerkin KJ, Kennel PJ, Zhang F, Weber MP,   . . . . . .   , Colombo PC, Mancini DM, Schulze PC, Levin B, Zorn E
Ventricular assist device elicits serum natural IgG that correlates with the development of primary graft dysfunction following heart transplantation
Apr 22, 2017
The Journal Of Heart And Lung Transplantation : The Official Publication Of The International Society For Heart Transplantation
Pre-transplant sensitization is a limiting factor in solid-organ transplantation. In heart transplants, ventricular assist device (VAD) implantation has been associated with sensitization to human leukocyte antigens (HLA). The effect of VAD on non-HLA antibodies is unclear. We have previously shown that polyreactive natural antibodies (Nabs) contribute to pre-sensitization in kidney allograft recipients. Here we assessed generation of Nabs after VAD implantation in pre-transplant sera and examined their contribution to cardiac allograft outcome. IgM and IgG Nabs were tested in pre-transplant serum samples collected from 206 orthotopic heart transplant recipients, including 128 patients with VAD (VAD patients) and 78 patients without VAD (no-VAD patients). Nabs were assessed by testing serum reactivity to apoptotic cells by flow cytometry and to the generic oxidized epitope, malondialdehyde, by enzyme-linked immunosorbent assay. No difference was observed in serum levels of IgM Nabs between VAD and no-VAD patients. However, serum IgG Nabs levels were significantly increased in VAD compared with no-VAD patients. This increase was likely due to the presence of the VAD, as revealed by lower serum IgG Nabs levels before implantation. Elevated pre-transplant IgG Nabs level was associated with development of primary graft dysfunction (PGD). Our study demonstrates that VAD support elicits IgG Nabs reactive to apoptotic cells and oxidized epitopes. These findings further support broad and non-specific B-cell activation by VAD, resulting in IgG sensitization. Moreover, the association of serum IgG Nabs levels with development of PGD suggests a possible role for these antibodies in the inflammatory reaction accompanying this complication. Copyright © 2017 International Society for the Heart and Lung Transplantation. Published by Elsevier Inc. All rights reserved.
The Ubiquitin Ligase CHIP Integrates Proteostasis and Aging by Regulation of Insulin Receptor Turnover
Apr 21, 2017   Cell
Tawo R, Pokrzywa W, Kevei É, Akyuz ME, Balaji V, Adrian S, Höhfeld J, Hoppe T
The Ubiquitin Ligase CHIP Integrates Proteostasis and Aging by Regulation of Insulin Receptor Turnover
Apr 21, 2017
Cell
Aging is attended by a progressive decline in protein homeostasis (proteostasis), aggravating the risk for protein aggregation diseases. To understand the coordination between proteome imbalance and longevity, we addressed the mechanistic role of the quality-control ubiquitin ligase CHIP, which is a key regulator of proteostasis. We observed that CHIP deficiency leads to increased levels of the insulin receptor (INSR) and reduced lifespan of worms and flies. The membrane-bound INSR regulates the insulin and IGF1 signaling (IIS) pathway and thereby defines metabolism and aging. INSR is a direct target of CHIP, which triggers receptor monoubiquitylation and endocytic-lysosomal turnover to promote longevity. However, upon proteotoxic stress conditions and during aging, CHIP is recruited toward disposal of misfolded proteins, reducing its capacity to degrade the INSR. Our study indicates a competitive relationship between proteostasis and longevity regulation through CHIP-assisted proteolysis, providing a mechanistic concept for understanding the impact of proteome imbalance on aging. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.
Functional Enhancer Screening in Single Cells
Apr 21, 2017   Molecular Cell
van Arensbergen J, van Steensel B
Functional Enhancer Screening in Single Cells
Apr 21, 2017
Molecular Cell
In this issue of Molecular Cell, Xie et al. (2017) introduce Mosaic-seq, a powerful technology that combines CRISPRi and single-cell RNA-seq. This method enables the high-throughput assessment of contributions of enhancers to gene regulation. Copyright © 2017 Elsevier Inc. All rights reserved.
Patterns of genome-wide diversity and population structure in the Drosophila athabasca species complex
Apr 21, 2017   Molecular Biology And Evolution
Wong Miller KM, Bracewell RR, Eisen MB, Bachtrog D
Patterns of genome-wide diversity and population structure in the Drosophila athabasca species complex
Apr 21, 2017
Molecular Biology And Evolution
The Drosophila athabasca species complex contains three recently diverged, prezygotically isolated semispecies (Western-Northern, Eastern-A, and Eastern-B) that are distributed across North America and share zones of sympatry. Inferences based on a handful of loci suggest that this complex might be an ideal system for studying the genetics of incipient speciation and the evolution of prezygotic isolating mechanisms, but patterns of differentiation have not been characterized systematically. Here, we assembled a draft genome for D. athabasca and analyze whole-genome re-sequencing data for 28 individuals from across the species range to characterize genome-wide patterns of diversity and population differentiation among semispecies. Patterns of differentiation on the X-chromosome vs. autosomes vary, with the X-chromosome showing better phylogenetic resolution and increased levels of between semispecies divergence. Despite low levels of overall differentiation and a lack of phylogenetic resolution of the autosomes for the most closely related semispecies, individuals do exhibit distinct genetic clustering. Demographic analyses provide some support for a model of isolation with migration within D. athabasca, with divergence times
Inhibition of thyroid hormone receptor locally in the retina is a therapeutic strategy for retinal degeneration
Apr 21, 2017   FASEB Journal : Official Publication Of The Federation Of American Societies For Experimental Biology
Ma H, Yang F, Butler MR, Belcher J, Redmond TM, Placzek AT, Scanlan TS, Ding XQ
Inhibition of thyroid hormone receptor locally in the retina is a therapeutic strategy for retinal degeneration
Apr 21, 2017
FASEB Journal : Official Publication Of The Federation Of American Societies For Experimental Biology
Thyroid hormone (TH) signaling regulates cell proliferation, differentiation, and metabolism. Recent studies have implicated TH signaling in cone photoreceptor viability. Using mouse models of retinal degeneration, we demonstrated that antithyroid drug treatment and targeting iodothyronine deiodinases (DIOs) to suppress cellular tri-iodothyronine (T3) production or increase T3 degradation preserves cones. In this work, we investigated the effectiveness of inhibition of the TH receptor (TR). Two genes, THRA and THRB, encode TRs; THRB2 has been associated with cone viability. Using TR antagonists and Thrb2 deletion, we examined the effects of TR inhibition. Systemic and ocular treatment with the TR antagonists NH-3 and 1-850 increased cone density by 30-40% in the Rpe65-/- mouse model of Leber congenital amaurosis and reduced the number of TUNEL+ cells. Cone survival was significantly improved in Rpe65-/- and Cpfl1 (a model of achromatopsia with Pde6c defect) mice with Thrb2 deletion. Ventral cone density in Cpfl1/Thrb2-/- and Rpe65-/- /Thrb2-/- mice was increased by 1- to 4-fold, compared with age-matched controls. Moreover, the expression levels of TR were significantly higher in the cone-degeneration retinas, suggesting locally elevated TR signaling. This work shows that the effects of antithyroid treatment or targeting DIOs were likely mediated by TRs and that suppressing TR protects cones. Our findings support the view that inhibition of TR locally in the retina is a therapeutic strategy for retinal degeneration management.-Ma, H., Yang, F., Butler, M. R., Belcher, J., Redmond, T. M., Placzek, A. T., Scanlan, T. S., Ding, X.-Q. Inhibition of thyroid hormone receptor locally in the retina is a therapeutic strategy for retinal degeneration. © FASEB.
A multi-systemic mitochondrial disorder due to a dominant p.Y955H disease variant in DNA polymerase gamma
Apr 21, 2017   Human Molecular Genetics
Siibak T, Clemente P, Bratic A, Bruhn H, Kauppila TES,   . . . . . .   , Wedell A, Peter B, Freyer C, Falkenberg M, Wredenberg A
A multi-systemic mitochondrial disorder due to a dominant p.Y955H disease variant in DNA polymerase gamma
Apr 21, 2017
Human Molecular Genetics
Mutations in the mitochondrial DNA polymerase, POLG, are associated with a variety of clinical presentations, ranging from early onset fatal brain disease in Alpers syndrome to chronic progressive external ophthalmoplegia. The majority of mutations are linked with disturbances of mitochondrial DNA (mtDNA) integrity and maintenance. On a molecular level, depending on their location within the enzyme, mutations either lead to mtDNA depletion or the accumulation of multiple mtDNA deletions, and in some cases these molecular changes can be correlated to the clinical presentation. We identified a patient with a dominant p.Y955H mutation in POLG, presenting with a severe, early-onset multi-systemic mitochondrial disease with bilateral sensorineural hearing loss, cataract, myopathy, and liver failure. Using a combination of disease models of Drosophila melanogaster (Dm) and in vitro biochemistry analysis, we compare the molecular consequences of the p.Y955H mutation to the well-documented p.Y955C mutation. We demonstrate that both mutations affect mtDNA replication and display a dominant negative effect, with the p.Y955H allele resulting in a more severe polymerase dysfunction. © The Author 2017. Published by Oxford University Press.
Emerging evidence for the role of differential tumor microenvironment in breast cancer racial disparity: a closer look at the surroundings
Apr 21, 2017   Carcinogenesis
Deshmukh SK, Srivastava SK, Tyagi N, Ahmad A, Singh AP, Ghadhban AA, Dyess DL, Carter JE, Dugger K, Singh S
Emerging evidence for the role of differential tumor microenvironment in breast cancer racial disparity: a closer look at the surroundings
Apr 21, 2017
Carcinogenesis
Although increased awareness leading to early detection and prevention, as well as advancements in treatment strategies, have resulted in superior clinical outcomes, African American women with breast cancer continue to have greater mortality rates, compared to Caucasian American counterparts. Moreover, African American women are more likely to have breast cancer at a younger age and be diagnosed with aggressive tumor sub-types. Such racial disparities can be attributed to socioeconomic differences, but it is increasingly being recognized that these disparities may indeed be due to certain genetic and other non-genetic biological differences. Tumor microenvironment, which provides a favorable niche for the growth of tumor cells, is comprised of several types of stromal cells and the various proteins secreted as a consequence of bi-directional tumor-stromal cross-talk. Emerging evidence suggests inherent biological differences in the tumor microenvironment of breast cancer patients from different racial backgrounds. Tumor microenvironment components, affected by the genetic make-up of the tumor cells as well as other non-tumor-associated factors, may also render patients more susceptible to the development of aggressive tumors and faster progression of disease resulting in early onset, thus adversely affecting patients' survival. This review provides an overview of breast cancer racial disparity and discusses the existence of race-associated differential tumor microenvironment and its underlying genetic and non-genetic causal factors. A better understanding of these aspects would help further research on effective cancer management and improved approaches for reducing the racial disparities gaps in breast cancer patients. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.
Ezrin activation by LOK phosphorylation involves a PIP2-dependent wedge mechanism
Apr 21, 2017   ELife
Pelaseyed T, Sauvanet C, Viswanatha R, Filter JJ, Goldberg ML, Bretscher A
Ezrin activation by LOK phosphorylation involves a PIP2-dependent wedge mechanism
Apr 21, 2017
ELife
How cells specify morphologically distinct plasma membrane domains is poorly understood. Prior work has shown that restriction of microvilli to the apical aspect of epithelial cells requires the localized activation of the membrane-F-actin linking protein ezrin. Using an in vitro system, we now define a multi-step process whereby the kinase LOK specifically phosphorylates ezrin to activate it. Binding of PIP2 to ezrin induces a conformational change permitting the insertion of the LOK C-terminal domain to wedge apart the membrane and F-actin-binding domains of ezrin. The N-terminal LOK kinase domain can then access a site 40 residues distal from the consensus sequence that collectively direct phosphorylation of the appropriate threonine residue. We suggest that this elaborate mechanism ensures that ezrin is only phosphorylated at the plasma membrane, and with high specificity by the apically localized kinase LOK.
CRISPR-Cas9-mediated genome editing in one blastomere of two-cell embryos reveals a novel Tet3 function in regulating neocortical development
Apr 21, 2017   Cell Research
Wang L, Li MY, Qu C, Miao WY, Yin Q,   . . . . . .   , Wong CC, Xu G, Jing N, Yu X, Li J
CRISPR-Cas9-mediated genome editing in one blastomere of two-cell embryos reveals a novel Tet3 function in regulating neocortical development
Apr 21, 2017
Cell Research
Studying the early function of essential genes is an important and challenging problem in developmental biology. Here, we established a method for rapidly inducing CRISPR-Cas9-mediated mutations in one blastomere of two-cell stage embryos, termed 2-cell embryo-CRISPR-Cas9 injection (2CC), to study the in vivo function of essential (or unknown) genes in founder chimeric mice. By injecting both Cre mRNA and CRISPR-Cas9 targeting the gene of interest into fluorescent reporter mice, the 2CC method can trace both wild-type and mutant cells at different developmental stages, offering internal control for phenotypic analyses of mutant cells. Using this method, we identified novel functions of the essential gene Tet3 in regulating excitatory and inhibitory synaptic transmission in the developing mouse cerebral cortex. By generating chimeric mutant mice, the 2CC method allows for the rapid screening of gene function in multiple tissues and cell types in founder chimeric mice, significantly expanding the current armamentarium of genetic tools.Cell Research advance online publication 21 April 2017; doi:10.1038/cr.2017.58.
The Mitochondrion of Euglena gracilis
Apr 21, 2017   Advances In Experimental Medicine And Biology
Zimorski V, Rauch C, van Hellemond JJ, Tielens AGM, Martin WF
The Mitochondrion of Euglena gracilis
Apr 21, 2017
Advances In Experimental Medicine And Biology
In the presence of oxygen, Euglena gracilis mitochondria function much like mammalian mitochondria. Under anaerobiosis, E. gracilis mitochondria perform a malonyl-CoA independent synthesis of fatty acids leading to accumulation of wax esters, which serve as the sink for electrons stemming from glycolytic ATP synthesis and pyruvate oxidation. Some components (enzymes and cofactors) of Euglena's anaerobic energy metabolism are found among the anaerobic mitochondria of invertebrates, others are found among hydrogenosomes, the H2-producing anaerobic mitochondria of protists.
Human circulating and tissue gastric cancer stem cells display distinct epithelial-mesenchymal features and behaviors
Apr 21, 2017   Journal Of Cancer Research And Clinical Oncology
Zhang S, Shang Y, Chen T, Zhou X, Meng W, Fan C, Lu R, Huang Q, Li X, Hong X, Zhou Z, Hu J, Mo X
Human circulating and tissue gastric cancer stem cells display distinct epithelial-mesenchymal features and behaviors
Apr 21, 2017
Journal Of Cancer Research And Clinical Oncology
Metastasis is a leading cause of cancer-related-deaths worldwide. Recently, cancer stem cells (CSCs) have been believed to be responsible for tumor initiation and metastasis, but till now, difference of cellular features and behaviors between CSCs from tumor tissues (TCSCs) and circulation (CCSCs) remains largely unknown, which hinders the progression of targeted therapies for metastasis. Here, we provide the features of circulating gastric cancer stem cells (CGCSCs) isolated from human gastric adenocarcinoma. The CGCSCs and TGCSCs were culture in a same serum free stem cell culture medium, however the morphology are different with each other. EMT-associated markers were measured by Immunofluorescence, Western Blotting, and RT-PCR methods, and the results indicated that the CGCSCs and TGCSCs carry different epithelial-mesenchymal features. And then, proliferation and apoptosis assays revealed that the CGCSCs exhibited characteristics of higher proliferation and resistance to apoptosis in vitro. Soft agar assay and nude mice tumorigenicity assay displayed strong tumorigenicity of CGCSCs. Finally, Matrigel invasion assays and in vivo experimental metastasis assay were also performed, which demonstrated that CGCSCs carry high invasive and metastatic capabilities than TGCSCs. As expected, the CGCSCs indeed showed extremely invasive and metastatic properties. They also exhibited distinctive mesenchymal phenotypes, high self-renewal, proliferative capabilities, tumor induction and low apoptosis. Interestingly, CGCSCs show small cell-size than TGCSCs (tissue gastric cancer stem cells). The findings might help us to understand the biological characteristic of CGCSCs deeply, and give light to strategies for cancer therapies.
Uncovering the Roles of Septins in Cilia
Apr 21, 2017   Frontiers In Cell And Developmental Biology
Palander O, El-Zeiry M, Trimble WS
Uncovering the Roles of Septins in Cilia
Apr 21, 2017
Frontiers In Cell And Developmental Biology
Septins are a family of GTP-binding proteins that associate with cellular membranes and the cytoskeleton. Their ability to polymerize into filamentous structures permits them to serve as diffusion barriers for membrane proteins and as multi-molecular scaffolds that recruit components of signaling pathways. At the cellular level, septins contribute to the regulation of numerous processes, including cytokinesis, cell polarity, cell migration, and many others. In this review, we discuss emerging evidence for roles of mammalian septins in the biogenesis and function of flagella and cilia, and how this may impact human diseases such as ciliopathies.
Lipid droplets and liver disease: from basic biology to clinical implications
Apr 21, 2017   Nature Reviews. Gastroenterology & Hepatology
Gluchowski NL, Becuwe M, Walther TC, Farese RV
Lipid droplets and liver disease: from basic biology to clinical implications
Apr 21, 2017
Nature Reviews. Gastroenterology & Hepatology
Lipid droplets are dynamic organelles that store neutral lipids during times of energy excess and serve as an energy reservoir during deprivation. Many prevalent metabolic diseases, such as the metabolic syndrome or obesity, often result in abnormal lipid accumulation in lipid droplets in the liver, also called hepatic steatosis. Obesity-related steatosis, or NAFLD in particular, is a major public health concern worldwide and is frequently associated with insulin resistance and type 2 diabetes mellitus. Here, we review the latest insights into the biology of lipid droplets and their role in maintaining lipid homeostasis in the liver. We also offer a perspective of liver diseases that feature lipid accumulation in these lipid storage organelles, which include NAFLD and viral hepatitis. Although clinical applications of this knowledge are just beginning, we highlight new opportunities for identifying molecular targets for treating hepatic steatosis and steatohepatitis.
Fructose-driven glycolysis supports anoxia resistance in the naked mole-rat
Apr 21, 2017   Science (New York, N.Y.)
Park TJ, Reznick J, Peterson BL, Blass G, Omerbašić D,   . . . . . .   , Smith ESJ, Larson J, Gotthardt M, Kempa S, Lewin GR
Fructose-driven glycolysis supports anoxia resistance in the naked mole-rat
Apr 21, 2017
Science (New York, N.Y.)
The African naked mole-rat's (Heterocephalus glaber) social and subterranean lifestyle generates a hypoxic niche. Under experimental conditions, naked mole-rats tolerate hours of extreme hypoxia and survive 18 minutes of total oxygen deprivation (anoxia) without apparent injury. During anoxia, the naked mole-rat switches to anaerobic metabolism fueled by fructose, which is actively accumulated and metabolized to lactate in the brain. Global expression of the GLUT5 fructose transporter and high levels of ketohexokinase were identified as molecular signatures of fructose metabolism. Fructose-driven glycolytic respiration in naked mole-rat tissues avoids feedback inhibition of glycolysis via phosphofructokinase, supporting viability. The metabolic rewiring of glycolysis can circumvent the normally lethal effects of oxygen deprivation, a mechanism that could be harnessed to minimize hypoxic damage in human disease. Copyright © 2017, American Association for the Advancement of Science.
COPII-coated membranes function as transport carriers of intracellular procollagen I
Apr 21, 2017   The Journal Of Cell Biology
Gorur A, Yuan L, Kenny SJ, Baba S, Xu K, Schekman R
COPII-coated membranes function as transport carriers of intracellular procollagen I
Apr 21, 2017
The Journal Of Cell Biology
The coat protein complex II (COPII) is essential for the transport of large cargo, such as 300-nm procollagen I (PC1) molecules, from the endoplasmic reticulum (ER) to the Golgi. Previous work has shown that the CUL3-KLHL12 complex increases the size of COPII vesicles at ER exit sites to more than 300 nm in diameter and accelerates the secretion of PC1. However, the role of large COPII vesicles as PC1 transport carriers was not unambiguously demonstrated. In this study, using stochastic optical reconstruction microscopy, correlated light electron microscopy, and live-cell imaging, we demonstrate the existence of mobile COPII-coated vesicles that completely encapsulate the cargo PC1 and are physically separated from ER. We also developed a cell-free COPII vesicle budding reaction that reconstitutes the capture of PC1 into large COPII vesicles. This process requires COPII proteins and the GTPase activity of the COPII subunit SAR1. We conclude that large COPII vesicles are bona fide carriers of PC1. © 2017 Gorur et al.
The plastid outer membrane localized LPTD1 is important for glycerolipid remodeling under phosphate starvation
Apr 22, 2017   Plant, Cell & Environment
Hsueh YC, Ehmann C, Flinner N, Ladig R, Schleiff E
The plastid outer membrane localized LPTD1 is important for glycerolipid remodeling under phosphate starvation
Apr 22, 2017
Plant, Cell & Environment
Glycerolipid synthesis in plants is coordinated between plastids and the endoplasmic reticulum (ER). A central step within the glycerolipid synthesis is the transport of phosphatidic acid from ER to chloroplasts. The chloroplast outer envelope protein TGD4 belongs to the LptD family conserved in bacteria and plants and selectively binds and may transports phosphatidic acid. We describe a second LptD-family protein in A. thaliana (atLPTD1; At2g44640) characterized by a barrel domain with an amino acid signature typical for cyanobacterial LptDs. It forms a cation selective channel in vitro with a diameter of about 9 Å. atLPTD1 levels are induced under phosphate starvation. Plants expressing an RNAi construct against atLPTD1 show a growth phenotype under normal conditions. Expressing the RNAi against atLPTD1 in the tgd4-1 background renders the plants more sensitive to light stress or phosphate limitation than the individual mutants. Moreover, lipid analysis revealed that digalactosyldiacylglycerol and sulfoquinovosyldiacylglycerol levels remain constant in the RNAi mutants under phosphate starvation, while these two lipids are enhanced in wild-type. Based on our results we propose a function of atLPTD1 in the transport of lipids from ER to chloroplast under phosphate starvation, which is combinatory with the function of TGD4. This article is protected by copyright. All rights reserved.

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