Role of flotillins in the endocytosis of GPCR in salivary gland epithelial cells

Park, M.Y.,Kim, N.,Wu, L.L.,Yu, G.Y.,Park, K. Academic Press 2016 Biochemical and biophysical research communication Vol.476 No.4

Endocytosis has numerous functions in cellular homeostasis. Defects in the endocytic pathway of receptors may lead to dysfunction of salivary gland secretion. Therefore, elucidating the complex mechanisms of endocytosis may facilitate solutions for disease treatment and prevention. The muscarinic type 3 receptor (M3R), a G-protein-coupled receptor (GPCR) located in the plasma membrane, is involved in numerous physiological activities such as smooth muscle contraction and saliva secretion. M3R enters cells through clathrin-mediated endocytosis (CME), while flotillins (flot-1 and -2), highly conserved proteins residing in lipid-raft microdomains, make use of clathrin-independent endocytosis (CIE) for their internalization. Since these two proteins use two discrete pathways for endocytic entry, the association of flotillins with CME is poorly understood. We examined whether flotillins play a role in CME of M3R using immunoblotting, immunocytochemistry, confocal immunofluorescence microscopy, co-immunoprecipitation, and RNA interference techniques in secretory epithelial cells. Upon stimulation with a cholinergic agonist, M3R, flot-1, and flot-2 each internalized from the plasma membrane into intracellular sites. The addition of chlorpromazine and cytochalasin D, well-known inhibitors of CME, inhibited internalization of M3R via CME. Filipin III and methyl-β-cyclodextrin (mβCD) acting as lipid raft inhibitors disrupted internalization of flot-½ via CIE. Interestingly, filipin III and mβCD slightly reduced expression level of M3R whereas chlorpromazine and cytochalasin D did not affect internalization of the flotillin isoforms. M3R and flot-½ colocalized and interacted with each other as they entered the cytosol during limited periods of incubation. Moreover, knockdown of flot-1 or -2 by flotillin-specific siRNA prevented internalization and reduced the endocytic efficiency of M3R. Our results suggest that flot-1 and -2 are partially involved in CME of M3R by facilitating its internalization.

SEARCH FOR PULSED γ-RAY EMISSION FROM GLOBULAR CLUSTER M28

Wu, J. H. K.,Hui, C. Y.,Wu, E. M. H.,Kong, A. K. H.,Huang, R. H. H.,Tam, P. H. T.,Takata, J.,Cheng, K. S. IOP Publishing 2013 ASTROPHYSICAL JOURNAL LETTERS - Vol.765 No.2

<P>Using the data from the Large Area Telescope on board the Fermi Gamma-ray Space Telescope, we have searched for gamma-ray pulsations from the direction of the globular cluster M28 (NGC 6626). We report the discovery of a signal with a frequency consistent with that of the energetic millisecond pulsar (MSP) PSR B1821-24 in M28. A weighted H-test test statistic of 28.8 is attained, which corresponds to a chance probability of similar to 10(-5) (4.3 sigma detection). With a phase-resolved analysis, the pulsed component is found to contribute similar to 25% of the total observed gamma-ray emission from the cluster. However, the unpulsed level provides a constraint for the underlying MSP population and the fundamental plane relations for the scenario of inverse Compton scattering. Follow-up timing observations in radio/X-ray are encouraged to further investigate this periodic signal candidate.</P>

Functional effects of β₃-adrenoceptor on pacemaker activity in interstitial cells of Cajal from the mouse colon

Wu, M.J.,Shin, D.H.,Kim, M.Y.,Park, C.G.,Kim, Y.D.,Lee, J.,Park, I.K.,Choi, S.,So, I.,Park, J.S.,Jun, J.Y. North-Holland ; Elsevier Science Ltd 2015 european journal of pharmacology Vol.754 No.-

<P>We investigated the presence of beta(3)-adrenoceptor and its functional effects on pacemaker potentials in colonic interstitial cells of Cajal (ICCs) from mice. The whole cell patch clamp technique was used to record pacemaker potentials in cultured ICCs and reverse transcription polymerase chain reaction (RT-PCR) was performed to detect the mRNA transcript levels beta-adrenoceptors. The beta(3)-adrenoceptor agonist, BRL37344, reduced the frequency of pacemaker potentials in a Concentration dependent manner. The inhibitory effects of BRL37344 wore blocked by the pretreatment of propranolol, a nonspecific beta-adrenoceptor antagonist, but not by the selective beta(1)-adrenoceptor antagonist atenolol and the selective beta(2)-adrenoceptor antagonist butoxamine. beta(3)-adrenocepto antagonists SR59230A and L748337 blocked the inhibitory effects of BRL37344. RT-PCR revealed mRNA transcripts of beta(1)- and beta(3)-adrenoceptor, but not beta(2)-adrenoceptor, in c-kit- and Ano-1-positive colonic ICCs. The K+ channel blockers tetraethylammoniu, apamin, and glibenclamide did not block the effects of BRL37344. N-omega-Nitio-L-arginiue methyl ester hydrochloride (L-NAME), an NO synthase inhibitor, and chelerythrine, a protein Kinase C inhibitor, also did not block the effects of BRL37344. Noradrenaline mimicked the effects of BRL37344 in colonic ICCs. However, the inhibitory effects of noradrenaline on pacemaker potentials were blocked only by pretreatment with atenolol but not by butoxamine, SR59230A, or L748337 in small intestinal ICCs, BRL37344 had no effect On pacemaker potentials and mRNA transcripts of beta(1)-and beta(2)-adrenoceptor, but not beta(3)-adrenoceptor were detected. These results suggest that beta(3)-adrenoceptors are present in colonic ICCs and may play a role in regulating gastrointestinal motility by the inhibition or pacemaker potentials. (C) 2015 Elsevier By. All rights reserved.</P>

Long-lived K isomer and enhanced γ vibration in the neutron-rich nucleus ¹⁷²Dy: Collectivity beyond double midshell

Watanabe, H.,Zhang, G.X.,Yoshida, K.,Walker, P.M.,Liu, J.J.,Wu, J.,Regan, P.H.,Soderstrom, P.A.,Kanaoka, H.,Korkulu, Z.,Lee, P.S.,Nishimura, S.,Yagi, A.,Ahn, D.S.,Alharbi, T.,Baba, H.,Browne, F.,Bruce North-Holland Pub. Co 2016 Physics letters. Section B Vol.760 No.-

The level structure of <SUP>172</SUP>Dy has been investigated for the first time by means of decay spectroscopy following in-flight fission of a <SUP>238</SUP>U beam. A long-lived isomeric state with T<SUB>½</SUB>=0.71(5) s and K<SUP>π</SUP>=8<SUP>-</SUP> has been identified at 1278 keV, which decays to the ground-state and γ-vibrational bands through hindered electromagnetic transitions, as well as to the daughter nucleus <SUP>172</SUP>Ho via allowed β decays. The robust nature of the K<SUP>π</SUP>=8<SUP>-</SUP> isomer and the ground-state rotational band reveals an axially-symmetric structure for this nucleus. Meanwhile, the γ-vibrational levels have been identified at unusually low excitation energy compared to the neighboring well-deformed nuclei, indicating the significance of the microscopic effect on the non-axial collectivity in this doubly mid-shell region. The underlying mechanism of enhanced γ vibration is discussed in comparison with the deformed Quasiparticle Random-Phase Approximation based on a Skyrme energy-density functional.

Defining the Stressome of Mycobacterium avium subsp. paratuberculosis In Vitro and in Naturally Infected Cows

Wu, Chia-wei,Schmoller, Shelly K.,Shin, Sung Jae,Talaat, Adel M. American Society for Microbiology 2007 Journal of Bacteriology Vol.189 No.21

<B>ABSTRACT</B><P><I>Mycobacterium avium</I> subsp. <I>paratuberculosis</I> causes an enteric infection in cattle, with a great impact on the dairy industry in the United States and worldwide. Characterizing the gene expression profile of <I>M. avium</I> subsp. <I>paratuberculosis</I> exposed to different stress conditions, or shed in cow feces, could improve our understanding of the pathogenesis of <I>M. avium</I> subsp. <I>paratuberculosis</I>. In this report, the stress response of <I>M. avium</I> subsp. <I>paratuberculosis</I> on a genome-wide level (stressome) was defined for the first time using DNA microarrays. Expression data analysis revealed unique gene groups of <I>M. avium</I> subsp. <I>paratuberculosis</I> that were regulated under in vitro stressors while additional groups were regulated in the cow samples. Interestingly, acidic pH induced the regulation of a large number of genes (<I>n</I> = 597), suggesting the high sensitivity of <I>M. avium</I> subsp. <I>paratuberculosis</I> to acidic environments. Generally, responses to heat shock, acidity, and oxidative stress were similar in <I>M. avium</I> subsp. <I>paratuberculosis</I> and <I>Mycobacterium tuberculosis</I>, suggesting common pathways for mycobacterial defense against stressors. Several sigma factors (e.g., <I>sigH</I> and <I>sigE</I>) were differentially coregulated with a large number of genes depending on the type of each stressor. Subsequently, we analyzed the virulence of six <I>M. avium</I> subsp. <I>paratuberculosis</I> mutants with inactivation of differentially regulated genes using a murine model of paratuberculosis. Both bacterial and histopathological examinations indicated the attenuation of all gene mutants, especially those selected based on their expression in the cow samples (e.g., <I>lipN</I>). Overall, the employed approach profiled mycobacterial genetic networks triggered by variable stressors and identified a novel set of putative virulence genes. A similar approach could be applied to analyze other intracellular pathogens.</P>

CHANDRA DETECTION OF A NEW DIFFUSE X-RAY COMPONENT FROM THE GLOBULAR CLUSTER 47 TUCANAE

Wu, E. M. H.,Hui, C. Y.,Kong, A. K. H.,Tam, P. H. T.,Cheng, K. S.,Dogiel, V. A. University of Chicago Press for the American Astro 2014 ASTROPHYSICAL JOURNAL LETTERS - Vol.788 No.2

In re-analyzing the archival Chandra data of the globular cluster 47 Tucanae, we have detected a new diffuse X-ray emission feature within the half-mass radius of the cluster. The spectrum of the diffuse emission can be described by a power-law model plus a plasma component with photon index Gamma similar to 1.0 and plasma temperature kT similar to 0.2 keV. While the thermal component is apparently uniform, the non-thermal contribution falls off exponentially from the core. The observed properties could possibly be explained in the context of multiple shocks resulting from the collisions among the stellar wind in the cluster and the inverse Compton scattering between the pulsar wind and the relic photons.

Transient Repairing Annealing on Patterned CoFeB-based Magnetic Tunnel Junctions

K. M. Wu,C. H. Huang,M. J. Kao,M. J. Tsai,J. C. Wul,L. Horng 한국자기학회 2007 한국자기학회 학술연구발표회 논문개요집 Vol.- No.-

Electrical Resistivity and Demagnetization Characteristics of Salt-added Nd-Fe-B-type Magnet

M. S. Kang,K. M. Kim,H. W. Kwon,D. Wu,M. Yue,M. C. Kang,C. W. Yang,D. H. Kim,J. G. Lee,J. H. Yu 한국자기학회 2019 Journal of Magnetics Vol.24 No.3

Nd-Fe-B-type die-upset magnet with high electrical resistivity was fabricated by hot-deforming the mixture of melt-spun Nd-Fe-B-type flakes (MQU-F: Nd13.6Fe73.6Co6.6Ga0.6B5.6) and Dy-containing salts: eutectic (DyF₃–LiF) salt mixture and DyF₃ single salt. Profound electrical resistivity enhancement was feasible in the Nd-Fe-B-type die-upset magnet by adding Dy-containing salts. More profound electrical resistivity enhancement was achieved in the magnet added with dielectric eutectic (DyF₃–LiF) salt mixture with respect to the magnet added with single DyF₃ salt. This was attributed to better electrical insulation between the flakes by forming more continuous coverage of the flake interface with the easily melted dielectric salt. Coercivity of the die-upset magnet was also profoundly enhanced by optimal addition of Dy-containing salts, and this was attributed to substitution of some Nd in the Nd₂Fe14B-type grains near flake surface by Dy atoms from the added salt. Kerr microscopy revealed that for both the magnets with or without salt addition, formation of reverse domain initiated mostly inside the flake. Reversed domain started to form at higher reverse field for the magnet added with Dy-containing salt than for the magnet without salt addition. Practical demagnetization occurred largely by formation of new reverse domains at random places rather than enlargement of previously formed reverse domain for both the magnets with or without salt addition.

ORBITAL-PHASE-DEPENDENT γ-RAY EMISSIONS FROM THE BLACK WIDOW PULSAR

Wu, E. M. H.,Takata, J.,Cheng, K. S.,Huang, R. H. H.,Hui, C. Y.,Kong, A. K. H.,Tam, P. H. T.,Wu, J. H. K. IOP Publishing 2012 The Astrophysical journal Vol.761 No.2

<P>We report on evidence for orbital phase dependence of the gamma-ray emission from the PSR B1957+20 black widow system using data from the Fermi Large Area Telescope. We divide an orbital cycle into two regions: one containing the inferior conjunction and the other containing the rest of the orbital cycle. We show that the observed spectra for the different orbital regions are fitted by different functional forms. The spectrum of the orbital region containing the inferior conjunction can be described by a power law with an exponential cutoff (PLE) model, which also gives the best-fit model for the orbital phase without the inferior conjunction, plus an extra component above similar to 2.7 GeV. The emission above 3 GeV in this region is detected with a similar to 7 sigma confidence level. The gamma-ray data above similar to 2.7 GeV are observed to be modulated at the orbital period at the similar to 2.3 sigma level. We anticipate that the PLE component dominant below similar to 2.7 GeV originates from the pulsar magnetosphere. We also show that inverse Compton scattering of the thermal radiation of the companion star off a 'cold' ultrarelativistic pulsar wind can explain the extra component above similar to 2.7 GeV. The black widow pulsar PSR B1957+20 may be a member of a new class of object, in the sense that the system is showing gamma-ray emission with both magnetospheric and pulsar wind origins.</P>

DISCOVERY OF GAMMA-RAY EMISSION FROM THE SUPERNOVA REMNANT Kes 17 WITH <i>FERMI</i> LARGE AREA TELESCOPE

Wu, J. H. K.,Wu, E. M. H.,Hui, C. Y.,Tam, P. H. T.,Huang, R. H. H.,Kong, A. K. H.,Cheng, K. S. IOP Publishing 2011 ASTROPHYSICAL JOURNAL LETTERS - Vol.740 No.1

<P>We report the discovery of GeV emission at the position of supernova remnant Kes 17 by using the data from the Large Area Telescope on board the Fermi Gamma-ray Space Telescope. Kes 17 can be clearly detected with a significance of similar to 12 sigma in the 1-20 GeV range. Moreover, a number of gamma-ray sources were detected in its vicinity. The gamma-ray spectrum of Kes 17 can be described well by a simple power law with a photon index of Gamma similar to 2.4. Together with the multiwavelength evidence for its interactions with the nearby molecular cloud, the gamma-ray detection suggests that Kes 17 is a candidate acceleration site for cosmic rays.</P>