2839 articles

72 publishers

Join mailing list

Jan 2021 • bioRxiv

Polyclonal lymphoid expansion drives paraneoplastic autoimmunity in neuroblastoma

Miriam I Rosenberg, Erez Greenstein, Martin Buchkovich, Martin Mikl, Ayelet Peres, Eric Santoni-Rugiu, Dan Reshef, Amy J Salovin, David L Gibbs, Meredith S Irwin, Arlene Naranjo, Igor Ulitsky, Pedro A de Alarcon, Victor Weigman, Gur Yaari, Jessica A Panzer, Nir Friedman, John M Maris

Neuroblastoma is a lethal childhood solid tumor of developing peripheral nerves. Two percent of children with neuroblastoma develop Opsoclonus Myoclonus Ataxia Syndrome (OMAS), a paraneoplastic disease characterized by cerebellar and brainstem-directed autoimmunity, but typically with outstanding cancer-related outcomes. We compared tumor transcriptomes and tumor infiltrating T- and B-cell repertoires from 38 OMAS subjects with neuroblastoma to 26 non-OMAS associated neuroblastomas. We found greater B- and T-cell infiltration in OMAS-associated tumors compared to controls, but unexpectedly showed that both were polyclonal expansions. Tertiary lymphoid structures (TLS) were enriched in OMAS-associated tumors. We identified significant enrichment of the MHC Class II allele HLA-DOB*01:01 in OMAS patients. OMAS severity scores were associated with the expression of several candidate autoimmune genes. We propose a model in which polyclonal autoreactive B lymphocytes act as antigen presenting cells and drive TLS formation, thereby crucially supporting both sustained polyclonal T-cell-mediated anti-tumor immunity and paraneoplastic OMAS neuropathology.

Show more

Jan 2021 • bioRxiv

Rapid Ensemble Measurement of Protein Diffusion, Probe Blinking and Photobleaching Dynamics in the Complex Cellular Space

Simon Sehayek, Xiyu Yi, Shimon Weiss, Paul W Wiseman

We present a fluorescence fluctuation image correlation analysis method that can rapidly and simultaneously measure the diffusion coefficient, photoblinking rates, and fraction of diffusing particles of fluorescent molecules in cells. Unlike other image correlation techniques, we demonstrated that our method could be applied irrespective of a non-uniformly distributed, immobile blinking fluorophore population. This allows us to measure blinking and transport dynamics in complex cell morphologies, a benefit for a range of super-resolution fluorescence imaging approaches that rely on probe emission blinking. Furthermore, we showed that our technique could be applied without directly accounting for photobleaching. We successfully employed our technique on several simulations with realistic EMCCD noise and photobleaching models, as well as on Dronpa-C12 labeled beta-actin in living NIH/3T3 and HeLa cells. We found that the diffusion coefficients measured using our method were consistent with previous literature values. We further found that photoblinking rates measured in the live HeLa cells varied as expected with changing excitation power.

Show more

Jan 2021 • Polymers

In Situ Grafting of Silica Nanoparticle Precursors with Covalently Attached Bioactive Agents to Form PVA-Based Materials for Sustainable Active Packaging

Miri Klein, Anat Molad Filossof, Idan Ashur, Sefi Vernick, Michal Natan-Warhaftig, Victor Rodov, Ehud Banin, Elena Poverenov

Sustainable antibacterial–antioxidant films were prepared using in situ graftings of silica nanoparticle (SNP) precursors with covalently attached bioactive agents benzoic acid (ba) or curcumin (cur) on polyvinyl alcohol (PVA). The modified PVA-SNP, PVA-SNP-ba and PVA-SNP-cur films were characterized using spectroscopic, physicochemical and microscopic methods. The prepared films showed excellent antibacterial and antioxidant activity, and increased hydrophobicity providing protection from undesired moisture. The PVA-SNP-ba films completely prevented the growth of the foodborne human pathogen Listeria innocua, whereas PVA-SNP-cur resulted in a 2.5 log reduction of this bacteria. The PVA-SNP-cur and PVA-SNP-ba films showed high antioxidant activity of 15.9 and 14.7 Mm/g TEAC, respectively. The described approach can serve as a generic platform for the formation of PVA-based packaging materials with tailor-made activity tuned by active substituents on silica precursors. Application of such biodegradable films bearing safe bioactive agents can be particularly valuable for advanced sustainable packaging materials in food and medicine. View Full-Text

Show more

Jan 2021 • Molecules

Redox Potential and Crystal Chemistry of Hexanuclear Cluster Compounds

Elena Levi, Doron Aurbach, Carlo Gatti

Most of TM 6-cluster compounds (TM= transition metal) are soluble in polar solvents, in which the cluster units commonly remain intact, preserving the same atomic arrangement as in solids. Consequently, the redox potential is often used to characterize structural and electronic features of respective solids. Although a high lability and variety of ligands allow for tuning of redox potential and of the related spectroscopic properties in wide ranges, the mechanism of this tuning is still unclear. Crystal chemistry approach was applied for the first time to clarify this mechanism. It was shown that there are two factors affecting redox potential of a given metal couple: Lever’s electrochemical parameters of the ligands and the effective ionic charge of TM, which in cluster compounds differs effectively from the formal value due to the bond strains around TM atoms. Calculations of the effective ionic charge of TMs were performed in the framework of bond valence model, which relates the valence of a bond to its length by simple Pauling relationship. It was also shown that due to the bond strains the charge depends mainly on the atomic size of the inner ligands. View Full-Text

Show more

Jan 2021 • International journal of molecular sciences

Weak Electromagnetic Fields Accelerate Fusion of Myoblasts

Dana Adler, Zehavit Shapira, Shimon Weiss, Asher Shainberg, Abram Katz

Weak electromagnetic fields (WEF) alter Ca 2+ handling in skeletal muscle myotubes. Owing to the involvement of Ca 2+ in muscle development, we investigated whether WEF affects fusion of myoblasts in culture. Rat primary myoblast cultures were exposed to WEF (1.75 µT, 16 Hz) for up to six days. Under control conditions, cell fusion and creatine kinase (CK) activity increased in parallel and peaked at 4–6 days. WEF enhanced the extent of fusion after one and two days (by~ 40%) vs. control, but not thereafter. Exposure to WEF also enhanced CK activity after two days (almost four-fold), but not afterwards. Incorporation of 3 H-thymidine into DNA was enhanced by one-day exposure to WEF (~ 40%), indicating increased cell replication. Using the potentiometric fluorescent dye di-8-ANEPPS, we found that exposure of cells to 150 mM KCl resulted in depolarization of the cell membrane. However, prior exposure of cells to WEF for one day followed by addition of KCl resulted in hyperpolarization of the cell membrane. Acute exposure of cells to WEF also resulted in hyperpolarization of the cell membrane. Twenty-four hour incubation of myoblasts with gambogic acid, an inhibitor of the inward rectifying K+ channel 2.1 (K ir 2.1), did not affect cell fusion, WEF-mediated acceleration of fusion or hyperpolarization. These data demonstrate that WEF accelerates fusion of myoblasts, resulting in myotube formation. The WEF effect is associated with hyperpolarization but WEF does not appear to mediate its effects on fusion by activating K ir 2.1 channels. View Full-Text

Show more

Jan 2021 • bioRxiv

Multi-parameter photon-by-photon hidden Markov modeling

Paul David Harris, Shimon Weiss, Eitan Lerner

Single molecule FRET (smFRET) is a useful tool for studying biomolecular sub-populations and their dynamics. Advanced smFRET-based techniques often track multiple parameters simultaneously, increasing the information content of the measurement. Photon-by-photon hidden Markov modelling (H2MM) is a smFRET analysis tool that quantifies FRET dynamics of single biomolecules, even if they occur in sub-milliseconds. However, sub-populations can be characterized by additional experimentally-derived parameters other than the FRET efficiency. We introduce multiparameter H2MM (mpH2MM) that identifies sub-populations and their transition dynamics based on multiple experimentally-derived parameters, simultaneously. We show the use of this tool in deciphering the number of underlying sub-populations, their mean characteristics and the rate constants of their transitions for a DNA hairpin exhibiting milliseconds FRET dynamics, and for the RNA polymerase promoter open complex exhibiting sub-millisecond FRET dynamics of the transcription bubble. Overall, we show that using mpH2MM facilitates the identification and quantification of biomolecular sub-populations in smFRET measurements that are otherwise difficult to identify. Finally we provide the means to use mpH2MM in analyzing FRET dynamics in advanced multi-color smFRET-based measurements.

Show more

Jan 2021 • Energies

In Situ Measurement of Localized Current Distribution in H2-Br2 Redox Flow Batteries

Brenda Berenice Martinez Cantu, Peter Fischer, David Zitoun, Jens Tübke, Karsten Pinkwart

Hydrogen bromine redox flow batteries (RFB) are considered to be one of the most promising storage alternatives, as this technology offers both high energy and high-power density. In this work a printed circuit board type of segmented current collector for the measurement of locally resolved current density was developed. This analytical tool was inserted as hydrogen anode current collector in a hydrogen-bromine test cell. Charging and discharging operation was monitored under different stoichiometric flow conditions and the impact on current distribution is presented. This technique offers the possibility to prove cell limiting conditions with spatial resolution, improving our understanding and determining optimal operating conditions for a given design. View Full-Text

Show more

Jan 2021 • bioRxiv

In vitro and in vivo NIR Fluorescence Lifetime Imaging with a time-gated SPAD camera

Jason T Smith, Alena Rudkouskaya, Shan Gao, Juhi M Gupta, Arin Ulku, Claudio Bruschini, Edoardo Charbon, Shimon Weiss, Margarida Barroso, Xavier Intes, Xavier Michalet

Near-infrared (NIR) fluorescence lifetime imaging (FLI) provides a unique contrast mechanism to monitor biological parameters and molecular events in vivo. Single-photon avalanche photodiode (SPAD) cameras have been recently demonstrated in FLI microscopy (FLIM) applications, but their suitability for in vivo macroscopic FLI (MFLI) in deep tissues remains to be demonstrated. Herein, we report in vivo NIR MFLI measurement with SwissSPAD2, a large time-gated SPAD camera. We first benchmark its performance in well-controlled in vitro experiments, ranging from monitoring environmental effects on fluorescence lifetime, to quantifying Förster Resonant Energy Transfer (FRET) between dyes. Next, we use it for in vivo studies of target-drug engagement in live and intact tumor xenografts using FRET. Information obtained with SwissSPAD2 was successfully compared to that obtained with a gated-ICCD camera, using two different approaches. Our results demonstrate that SPAD cameras offer a powerful technology for in vivo preclinical applications in the NIR window.

Show more

Jan 2021 • Autophagy

Guidelines for the use and interpretation of assays for monitoring autophagy 1

Alim Al-Bari, Alves da Costa, Belló Pérez, Bravo-San Pedro, Calvo-Rubio Barrera, Candido de Almeida, Cebollada Rica, Dal Col, Dalla Valle, Dayalan Naidu, De Amicis, De Bosscher, De Felice, De Franceschi, De Leonibus, Barbosa de Mattos, De Meyer, De Milito, De Nunzio, De Palma, De Santi, De Virgilio, De Zio, Del Rio, Di Cristina, Di Domenico, Di Fazio, Di Fonzo, Di Guardo, Di Guglielmo, Di Leo, Di Malta, Di Nardo, Di Rienzo, Di Sano, Dokmeci Emre, El Andaloussi, Fenz Araujo, Gallolu Kankanamalage, García-Del Portillo, Ghasemipour Afshar, Hasima Nagoor, Yi-Ren Hong, Hyung-Ryong Kim, La Spada, Le Stunff, Martinez Velazquez, Misirkic Marjanovic, Monzio Compagnoni, Mulcahy Levy, Ní Cheallaigh, Pereira de Almeida, Porte Alcon, Ramachandra Rao, Ribeiro de Andrade Ramos, Rodrigues Silva, Srinivas Bharath, Clair St, Van den Berghe, Van Kaer, Von Haefen

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question …

Show more

Jan 2021 • Metamaterials, Metadevices, and Metasystems 2021, 2021

Subwavelength Mie-resonant selenium resonators for mid-infrared meta-optics

Danveer Singh, Tomer Lewi

Efficient light manipulation at subwavelength scales in the mid-infrared (MIR) region is essential for various applications and can be harnessed from intrinsic low-loss dielectric resonators. Here, we demonstrate the fabrication of truncated spherical selenium (Se) resonators with tunable high-quality (Q) factor Mie resonances. Large area amorphous Se subwavelength resonators of varying sizes were grown on different substrates, using a novel CVD process. We demonstrate size-tunable Mie resonances spanning the 2-16 µm range, for single isolated resonators and large area ensembles, respectively. We show strong tunable absorption resonances (90%) in ensembles of resonators in a significantly broad MIR range. Moreover, by coupling resonators to epsilon-near-zero (ENZ) substrates, we engineer high-Q resonances as high as Q=40. These findings open up new possibilities in meta-atom paints, anti …

Show more

Jan 2021 • bioRxiv

Protein degradation analysis by affinity microfluidics

Lev Brio, Danit Wasserman, Efrat Michaely-Barbiro, Doron Gerber, Amit Tzur

Jan 2021 • arXiv preprint arXiv:2101.11380

Countering a fundamental law of attraction with quantum wavepacket engineering

Gal Amit, Yonathan Japha, Tomer Shushi, Ron Folman, Eliahu Cohen

Bohmian mechanics was designed to give rise to predictions identical to those derived by standard quantum mechanics, while invoking a specific interpretation of it - one which allows the classical notion of a particle to be maintained alongside a guiding wave. For this, the Bohmian model makes use of a unique quantum potential which governs the trajectory of the particle. In this work we show that this interpretation of quantum theory naturally leads to the derivation of interesting new phenomena. Specifically, we demonstrate how the fundamental Casimir-Polder force, by which atoms are attracted to a surface, may be temporarily suppressed by utilizing a specially designed quantum potential. We show that when harnessing the quantum potential via a suitable atomic wavepacket engineering, the absorption by the surface can be dramatically reduced. This is proven both analytically and numerically. Finally, an experimental scheme is proposed for achieving the required shape for the atomic wavepacket. All these may enable new insights into Bohmian mechanics as well as new applications to metrology and sensing.

Show more

Jan 2021 • bioRxiv

Diet-induced modifications to human microbiome reshape colonic homeostasis in irritable bowel syndrome

Ayelet Pearl, Hadar Bootz, Ehud Melzer, Efrat Sharon, Shlomi Abuchatzera, Sivan Amidror, Elana Aretz, Irit Shoval, Orly Yaron, Stephen Malnick, Nissan Yissachar

Changes in microbiome composition have been associated with a wide array of human diseases, turning the human microbiota into an attractive target for therapeutic intervention. Yet clinical translation of these findings requires the establishment of causative connections between specific microbial taxa and their functional impact on host tissues. Here, we colonized gut organ cultures with longitudinal microbiota samples collected from newly-diagnosed and therapy-naive irritable bowel syndrome (IBS) patients under low-FODMAP (fermentable Oligo-, Di-, Mono-saccharides and Polyols) diet. We show that post-diet microbiota regulates intestinal expression of inflammatory and neuro-muscular gene-sets. Specifically, we identify Bifidobacterium adolescentis as a diet-sensitive pathobiont that alters tight junction integrity and disrupts gut barrier functions. Collectively, we present a unique pathway discovery approach for mechanistic dissection and identification of functional diet-host-microbiota modules. Our data support the hypothesis that the gut microbiota mediates the beneficial effects of low-FODMAP diet, and reinforce the potential feasibility of microbiome based-therapies in IBS.

Show more

Jan 2021 • Zika Virus Impact, Diagnosis, Control, and Models, 231-241, 2021

Magnetic modulation biosensing: How it works and how it can be used to detect the Zika virus

Shira Roth, Amos Danielli

Current serological diagnostic tests for Zika virus (ZIKV) suffer from reduced sensitivity, high cross-reactivity, low specificity, and lengthy protocols. Here, we begin by reviewing serological and antigenemia assays that are either commercially available or under development. We then focus on a new technology, named magnetic modulation biosensing (MMB), that enables rapid detection of ZIKV immunoglobulin M (IgM) and immunoglobulin G (IgG) antibodies. The MMB system utilizes magnetic beads that are conjugated to recombinant ZIKV nonstructural 1 (NS1) protein, which specifically captures Zika IgM/IgG antibodies. A second fluorescently labeled antibody is then added, forming a “sandwich” with the analyte and the capture protein. An oscillating magnetic field gradient concentrates the beads within the sample volume and transports them in a periodic motion in and out of a laser beam, producing an …

Show more

Jan 2021 • Materials

Effect of Diamond Polishing and Thermal Treatment on Carbon Paramagnetic Centers’ Nature and Structure

Ira Litvak, Avner Cahana, Yaakov Anker, Sharon Ruthstein, Haim Cohen

Diamonds contain carbon paramagnetic centers (stable carbon radicals) in small concentrations (at the level of ~1 × 1012 spins/mg) that can help in elucidating the structure of the nitrogen atoms’ contaminants in the diamond crystal. All diamonds that undergo polishing are exposed to high temperatures, owing to the friction force during the polishing process, which may affect the carbon-centered radicals’ concentration and structure. The temperature is increased appreciably; consequently, the black body radiation in the visible range turns orange. During polishing, diamonds emit an orange light (at a wavelength of about 600 nm) that is typical of a black body temperature of 900 °C or higher. Other processes in which color-enhanced diamonds are exposed to high temperatures are thermal treatments or the high-pressure, high-temperature (HPHT) process in which the brown color (resulting from plastic deformation) is bleached. The aim of the study was to examine how thermal treatment and polishing influence the paramagnetic centers in the diamond. For this purpose, four rough diamonds were studied: two underwent a polishing process, and the other two were thermally treated at 650 °C and 1000 °C. The diamonds were analyzed pre- and post-treatment by EPR (Electron Paramagnetic resonance), FTIR (Fourier transform infrared, fluorescence, and their visual appearance. The results indicate that the polishing process results in much more than just thermal heating the paramagnetic centers.

Show more

Jan 2021 • The European Physical Journal D

Gas-phase studies of the retinal protonated Schiff base chromophore

Y Toker, LH Andersen

Gas-phase studies of the retinal protonated Schiff base chromophore are reviewed. The use of action spectroscopy has solidified the understanding of the spectral-tuning mechanisms of this important chromophore. Ion-mobility spectrometry and gas phase femtosecond pump-probe spectroscopy studies indicate that several of the remarkable photo-isomerization properties of the chromophore such as its specificity and ultrafast nature are intrinsic properties of the chromophore. With a firm understanding of the properties of the isolated retinal chromophore in terms of spectroscopy and dynamics, the influence of the protein is becoming better understood.

Show more

Jan 2021 • Applied Sciences

Protective Measurement—A New Quantum Measurement Paradigm: Detailed Description of the First Realization

Enrico Rebufello, Fabrizio Piacentini, Alessio Avella, Rudi Lussana, Federica Villa, Alberto Tosi, Marco Gramegna, Giorgio Brida, Eliahu Cohen, Lev Vaidman, Ivo Pietro Degiovanni, Marco Genovese

We present a detailed description of the experiment realizing for the first time a protective measurement, a novel measurement protocol which combines weak interactions with a “protection mechanism” preserving the measured state coherence during the whole measurement process. Furthermore, protective measurement allows finding the expectation value of an observable, ie, an inherently statistical quantity, by measuring a single particle, without the need for any statistics. This peculiar property, in sharp contrast to the framework of traditional (projective) quantum measurement, might constitute a groundbreaking advance for several quantum technology related fields. View Full-Text

Show more

Jan 2021 • Zenodo, 2021

Data in: Aging power spectrum of membrane protein transport and other subordinated random walks

Eli Barkai, Diego Krapf

{"Abstract":["Datasets generated in the report "Aging power spectrum of membrane protein transport and other subordinated random walks". Included data are:\n\nNumerical simulations \nRWdata1.mat: 10,000 realizations, subordinated random walk with Hurst exponent, H=0.3 and \\(\\alpha\\)=0.4.\nRWdata3.mat: 10,000 realizations, subordinated random walk with Hurst exponent, H=0.7 and \\(\\alpha\\)=0.4.\nRWdata8.mat: 5,000 realizations, subordinated random walk with Hurst exponent, H=0.75 and \\(\\alpha\\)=0.8.\nRWdataCTRW.mat: 10,000 realizations, continuous time random walk (CTRW), \\(\\alpha\\)=0.7.\n\nSpectra of simulations\nPSDdata1.mat: Power spectral density (PSD) of a subordinated random walk with Hurst exponent, H=0.3 and \\(\\alpha\\)=0.4. Five different realization times are used to compute the PDS: 2^8, 2^10, 2^12, 2^14, and 2^16.\nPSDdata3.mat: PSD of a subordinated random walk with Hurst exponent, H=0.7 and \\(\\alpha\\)=0.4. Five different realization times are used to compute the PDS: 2^8, 2^10, 2^12, 2^14, and 2^16.\nPSDdata8.mat: PSD of a subordinated random walk with Hurst exponent, H=0.75 and \\(\\alpha\\)=0.8. Four different realization times are used to compute the PDS: 2^15, 2^16, 2^17, and 2^18.\nPSDs_CTRW.mat: PSD of a continuous-time random walk (CTRW), \\(\\alpha\\)=0.7. Five different realization times are used to compute the PDS: 2^8, 2^10, 2^12, 2^14, and 2^16.\n\nExperimental data of Nav1.6 channels in the soma of hippocampal neurons\nNavMSDtimes.csv: ensemble-averaged (EA) MSD and time-averaged (TA) MSD. The TA-MSD is measured for three observation times, 64, 128 …

Show more

Jan 2021 • Sensors

Rapid and Sensitive Inhibitor Screening Using Magnetically Modulated Biosensors

Shira Roth, Amos Danielli

Inhibitor screening is an important tool for drug development, especially during the COVID-19 pandemic. The most used in vitro inhibitor screening tool is an enzyme-linked immunosorbent assay (ELISA). However, ELISA-based inhibitor screening is time consuming and has a limited dynamic range. Using fluorescently and magnetically modulated biosensors (MMB), we developed a rapid and sensitive inhibitor screening tool. This study demonstrates its performance by screening small molecules and neutralizing antibodies as potential inhibitors of the interaction between the spike protein 1 (S1) of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and the angiotensin-converting enzyme 2 (ACE2) receptor. The MMB-based assay is highly sensitive, has minimal non-specific binding, and is much faster than the commonly used ELISA (2 h vs. 7–24 h). We anticipate that our method will lead to a remarkable advance in screening for new drug candidates. View Full-Text

Show more

Jan 2021 • NeuroMolecular Medicine

Correction to: Specific Susceptibility to COVID-19 in Adults with Down Syndrome (NeuroMolecular Medicine,(2021), 10.1007/s12017-021-08651-5)

Tomer Illouz, Arya Biragyn, Milana Frenkel-Morgenstern, Orly Weissberg, Alessandro Gorohovski, Eugene Merzon, Ilan Green, Florencia Iulita, Lisi Flores-Aguilar, Mara Dierssen, Ilario De Toma, Hefziba Lifshitz, Stylianos E Antonarakis, Eugene Yu, Yann Herault, Marie Claude Potier, Alexandra Botté, Randall Roper, Benjamin Sredni, Ronit Sarid, Jacqueline London, William Mobley, Andre Strydom, Eitan Okun

The original version of this article unfortunately contained mistakes. The author name Maria del Mar Dierssen Sotos should read as Mara Dierssen and the author’s affiliations should appear as follows:

Show more

Jan 2021 • bioRxiv

The Spliced Leader RNA Silencing (SLS) Pathway in Trypanosoma brucei Is Induced by Perturbations of Endoplasmic Reticulum, Golgi Complex, or Mitochondrial Protein Factors …

Uthman Okalang, Bar Mualem Bar-Ner, K Shanmugha Rajan, Nehemya Friedman, Saurav Aryal, Katarina Egarmina, Ronen Hope, Netaly Khazanov, Hanoch Senderowitz, Assaf Alon, Deborah Fass, Shulamit Michaeli

In the parasite Trypanosoma brucei, the causative agent of human African sleeping sickness, all mRNAs are trans-spliced to generate a common 59 exon derived from the spliced leader RNA (SL RNA). Perturbations of protein translocation across the endoplasmic reticulum (ER) induce the spliced leader RNA silencing (SLS) pathway. SLS activation is mediated by a serine-threonine kinase, PK3, which translocates from the cytosolic face of the ER to the nucleus, where it phosphorylates the TATA binding protein TRF4, leading to the shut-off of SL RNA transcription, followed by induction of programmed cell death. Here, we demonstrate that SLS is also induced by depletion of the essential ER resident chaperones BiP and calreticulin, ER oxidoreductin 1 (ERO1), and the Golgi-localized quiescin sulfhydryl oxidase (QSOX1). Most strikingly, silencing of Rhomboid-like 1(TIMRHOM1) involved in mitochondrial protein import, also induces SLS. The PK3 kinase, which integrates SLS signals, is modified by phosphorylation on multiple sites. To determine which of the phosphorylation events activate PK3, several individual mutations or their combination were generated.These mutations failed to completely eliminate the phosphorylation or translocation of the kinase to the nucleus. The structure of PK3 kinase and its ATP binding domain were therefore modeled. A conserved phenylalanine at position 771 was proposed to interact with ATP, and the PK3F771L mutation completely eliminated phosphorylation under SLS, suggesting that the activation involves most if not all the phosphorylation sites. The study suggests that the SLS occurs broadly in …

Show more


Powered by Articali