Publications by authors named "Ron Rotkopf"

34 Publications

TENT4A Non-Canonical Poly(A) Polymerase Regulates DNA-Damage Tolerance via Multiple Pathways That Are Mutated in Endometrial Cancer.

Int J Mol Sci 2021 Jun 28;22(13). Epub 2021 Jun 28.

Department of Biomolecular Sciences, Weizmann Institute of Science, Rehovot 7610001, Israel.

TENT4A (PAPD7) is a non-canonical poly(A) polymerase, of which little is known. Here, we show that TENT4A regulates multiple biological pathways and focuses on its multilayer regulation of translesion DNA synthesis (TLS), in which error-prone DNA polymerases bypass unrepaired DNA lesions. We show that TENT4A regulates mRNA stability and/or translation of DNA polymerase η and RAD18 E3 ligase, which guides the polymerase to replication stalling sites and monoubiquitinates PCNA, thereby enabling recruitment of error-prone DNA polymerases to damaged DNA sites. Remarkably, in addition to the effect on RAD18 mRNA stability via controlling its poly(A) tail, TENT4A indirectly regulates RAD18 via the tumor suppressor CYLD and via the long non-coding antisense RNA , which had no known function. Knocking down the expression of or , or overexpression of led each to reduced amounts of the RAD18 protein and DNA polymerase η, leading to reduced TLS, highlighting as a new TLS regulator. Bioinformatics analysis revealed that TLS error-prone DNA polymerase genes and their -related regulators are frequently mutated in endometrial cancer genomes, suggesting that TLS is dysregulated in this cancer.
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http://dx.doi.org/10.3390/ijms22136957DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8267958PMC
June 2021

missense mutations in PDAC are associated with enhanced fibrosis and an immunosuppressive microenvironment.

Proc Natl Acad Sci U S A 2021 Jun;118(23)

Department of Molecular Cell Biology, Weizmann Institute of Science, 76100 Rehovot, Israel;

Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal cancer, which is refractory to all currently available treatments and bears dismal prognosis. About 70% of all PDAC cases harbor mutations in the tumor suppressor gene. Many of those are missense mutations, resulting in abundant production of mutant p53 (mutp53) protein in the cancer cells. Analysis of human PDAC patient data from The Cancer Genome Atlas (TCGA) revealed a negative association between the presence of missense mutp53 and infiltration of CD8 T cells into the tumor. Moreover, CD8 T cell infiltration was negatively correlated with the expression of fibrosis-associated genes. Importantly, silencing of endogenous mutp53 in KPC cells, derived from mouse PDAC tumors driven by mutant Kras and mutp53, down-regulated fibrosis and elevated CD8 T cell infiltration in the tumors arising upon orthotopic injection of these cells into the pancreas of syngeneic mice. Moreover, the tumors generated by mutp53-silenced KPC cells were markedly smaller than those elicited by mutp53-proficient control KPC cells. Altogether, our findings suggest that missense p53 mutations may contribute to worse PDAC prognosis by promoting a more vigorous fibrotic tumor microenvironment and impeding the ability of the immune system to eliminate the cancer cells.
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http://dx.doi.org/10.1073/pnas.2025631118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8201917PMC
June 2021

The relationship between the plasma proinflammatory cytokine levels of depressed/anxious children and their parents.

Sci Rep 2021 Jun 3;11(1):11798. Epub 2021 Jun 3.

Department of Psychological Medicine, Schneider Children's Medical Center of Israel, 14 Kaplan Street, 4920235, Petach Tikva, Israel.

Recent studies suggest immune function dysregulation in depression and anxiety disorders. Elevated pro-inflammatory cytokines may be a marker for immune system dysregulation. No study assessed the correlation between the levels of cytokines in children and adolescents with depression/anxiety disorders and their parents. In this study, 92 children and adolescents (mean age 13.90 ± 2.41 years) with depression and/or anxiety disorders were treated with fluoxetine. Blood samples were collected before initiation of treatment. One hundred and sixty-four of their parents (mean age 50.6 ± 6.2 years) and 25 parents of healthy children (mean age 38.5 ± 6.2 years) also gave blood samples. Plasma levels of three pro-inflammatory cytokine (TNF-α, IL-6, IL-1β) were measured by enzyme linked immunosorbent assays (ELISA) and compared between depressed/anxious children and their parents. We also compared cytokine levels between parents of children with depression/anxiety and control parents. Mothers of depressed children had higher TNF-α levels than mothers of controls. No significant difference was detected in the fathers. A positive correlation was found between the IL-1β levels of the depressed/anxious boys and their mothers. No such correlation was observed in the fathers. Our conclusions are that higher levels of proinflammatory cytokines may indicate immune system activation in mothers in response to the distress associated with having depressed/anxious offspring. The correlation between IL-1β levels in the mothers and their depressed/anxious children may indicate familial vulnerability to depression and anxiety. Our observation highlights the need for a better understanding of sexual dimorphism in inflammatory responses to stress.
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http://dx.doi.org/10.1038/s41598-021-90971-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8175361PMC
June 2021

A Synthetic SOD/Catalase Mimic Compound for the Treatment of ALS.

Antioxidants (Basel) 2021 May 22;10(6). Epub 2021 May 22.

Schulich Faculty of Chemistry, Technion-Israel Institute of Technology, Haifa 32000, Israel.

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease affecting motor neurons. To date, the etiology of the disease is still unclear, with evidence of reactive oxygen species, mitochondrial dysfunction, iron homeostasis perturbation, protein misfolding and protein aggregation as key players in the pathology of the disease. Twenty percent of familial ALS and two percent of sporadic ALS instances are due to a mutation in Cu/Zn superoxide dismutase (SOD1). Sporadic and familial ALS affects the same neurons with similar pathology; therefore, the underlying hypothesis is that therapies effective in mutant SOD1 models could be translated to sporadic ALS. Corrole metal complexes have lately been identified as strong and potent catalytic antioxidants with beneficial effects in oxidative stress-related diseases such as Parkinson's disease, Alzheimer's disease, atherosclerosis, diabetes and its complications. One of the most promising candidates is the iron complex of an amphiphilic corrole, . In this study we used the SOD1 G93R mutant zebrafish ALS model to assess whether , as a potent catalytic antioxidant, displays any therapeutic merits in vivo. Our results show that caused a substantial increase in mutant zebrafish locomotor activity (up to 30%), bringing the locomotive abilities of the mutant treated group close to that of the wild type untreated group (50% more than the mutated untreated group). Furthermore, did not affect WT larvae locomotor activity, suggesting that enhances locomotor ability by targeting mechanisms underlying SOD1 ALS specifically. These results may pave the way for future development of as a viable treatment for ALS.
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http://dx.doi.org/10.3390/antiox10060827DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8224677PMC
May 2021

Identification of bacteria-derived HLA-bound peptides in melanoma.

Nature 2021 04 17;592(7852):138-143. Epub 2021 Mar 17.

Department of Melanoma Medical Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.

A variety of species of bacteria are known to colonize human tumours, proliferate within them and modulate immune function, which ultimately affects the survival of patients with cancer and their responses to treatment. However, it is not known whether antigens derived from intracellular bacteria are presented by the human leukocyte antigen class I and II (HLA-I and HLA-II, respectively) molecules of tumour cells, or whether such antigens elicit a tumour-infiltrating T cell immune response. Here we used 16S rRNA gene sequencing and HLA peptidomics to identify a peptide repertoire derived from intracellular bacteria that was presented on HLA-I and HLA-II molecules in melanoma tumours. Our analysis of 17 melanoma metastases (derived from 9 patients) revealed 248 and 35 unique HLA-I and HLA-II peptides, respectively, that were derived from 41 species of bacteria. We identified recurrent bacterial peptides in tumours from different patients, as well as in different tumours from the same patient. Our study reveals that peptides derived from intracellular bacteria can be presented by tumour cells and elicit immune reactivity, and thus provides insight into a mechanism by which bacteria influence activation of the immune system and responses to therapy.
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http://dx.doi.org/10.1038/s41586-021-03368-8DOI Listing
April 2021

20S proteasomes secreted by the malaria parasite promote its growth.

Nat Commun 2021 02 19;12(1):1172. Epub 2021 Feb 19.

Department of Biomolecular Sciences, Weizmann Institute of Science, Rehovot, 7610001, Israel.

Mature red blood cells (RBCs) lack internal organelles and canonical defense mechanisms, making them both a fascinating host cell, in general, and an intriguing choice for the deadly malaria parasite Plasmodium falciparum (Pf), in particular. Pf, while growing inside its natural host, the human RBC, secretes multipurpose extracellular vesicles (EVs), yet their influence on this essential host cell remains unknown. Here we demonstrate that Pf parasites, cultured in fresh human donor blood, secrete within such EVs assembled and functional 20S proteasome complexes (EV-20S). The EV-20S proteasomes modulate the mechanical properties of naïve human RBCs by remodeling their cytoskeletal network. Furthermore, we identify four degradation targets of the secreted 20S proteasome, the phosphorylated cytoskeletal proteins β-adducin, ankyrin-1, dematin and Epb4.1. Overall, our findings reveal a previously unknown 20S proteasome secretion mechanism employed by the human malaria parasite, which primes RBCs for parasite invasion by altering membrane stiffness, to facilitate malaria parasite growth.
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http://dx.doi.org/10.1038/s41467-021-21344-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7895969PMC
February 2021

Brain pathology and cerebellar purkinje cell loss in a mouse model of chronic neuronopathic Gaucher disease.

Prog Neurobiol 2021 02 3;197:101939. Epub 2020 Nov 3.

Department of Biomolecular Sciences, Weizmann Institute of Science, Rehovot, Israel.

Gaucher disease (GD) is currently the focus of considerable attention due primarily to the association between the gene that causes GD (GBA) and Parkinson's disease. Mouse models exist for the systemic (type 1) and for the acute neuronopathic forms (type 2) of GD. Here we report the generation of a mouse that phenotypically models chronic neuronopathic type 3 GD. Gba;Gba mice, which contain a Gba transgene regulated by doxycycline, accumulate moderate levels of the offending substrate in GD, glucosylceramide, and live for up to 10 months, i.e. significantly longer than mice which model type 2 GD. Gba;Gba mice display behavioral abnormalities at ∼4 months, which deteriorate with age, along with significant neuropathology including loss of Purkinje neurons. Gene expression is altered in the brain and in isolated microglia, although the changes in gene expression are less extensive than in mice modeling type 2 disease. Finally, bone deformities are consistent with the Gba;Gba mice being a genuine type 3 GD model. Together, the Gba;Gba mice share pathological pathways with acute neuronopathic GD mice but also display differences that might help understand the distinct disease course and progression of type 2 and 3 patients.
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http://dx.doi.org/10.1016/j.pneurobio.2020.101939DOI Listing
February 2021

Efficacy of Ciprofloxacin/Celecoxib combination in zebrafish models of amyotrophic lateral sclerosis.

Ann Clin Transl Neurol 2020 10 11;7(10):1883-1897. Epub 2020 Sep 11.

The Dead Sea Arava Science Center, Auspices of Ben Gurion University, Central Arava, 86815, Israel.

Objective: To evaluate the efficacy of a fixed-dose combination of two approved drugs, Ciprofloxacin and Celecoxib, as a potential therapeutic treatment for amyotrophic lateral sclerosis (ALS).

Methods: Toxicity and efficacy of Ciprofloxacin and Celecoxib were tested, each alone and in distinct ratio combinations in SOD1 G93R transgenic zebrafish model for ALS. Quantification of swimming measures following stimuli, measurements of axonal projections from the spinal cord, neuromuscular junction structure and morphometric analysis of microglia cells were performed in the combination- treated vs nontreated mutant larvae. Additionally, quantifications of touch-evoked locomotor escape response were conducted in treated vs nontreated zebrafish expressing the TARDBP G348C ALS variant.

Results: When administered individually, Ciprofloxacin had a mild effect and Celecoxib had no therapeutic effect. However, combined Ciprofloxacin and Celecoxib (Cipro/Celecox) treatment caused a significant increase of ~ 84% in the distance the SOD1 G93R transgenic larvae swam. Additionally, Cipro/Celecox elicited recovery of impaired motor neurons morphology and abnormal neuromuscular junction structure and preserved the ramified morphology of microglia cells in the SOD1 mutants. Furthermore, larvae expressing the TDP-43 mutation displayed evoked touch responses that were significantly longer in swim distance (110% increase) and significantly higher in maximal swim velocity (~44% increase) when treated with Cipro/Celecox combination.

Interpretation: Cipro/Celecox combination improved locomotor and cellular deficits of ALS zebrafish models. These results identify this novel combination as effective, and may prove promising for the treatment of ALS.
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http://dx.doi.org/10.1002/acn3.51174DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7545590PMC
October 2020

Piezo2 expressed in proprioceptive neurons is essential for skeletal integrity.

Nat Commun 2020 06 23;11(1):3168. Epub 2020 Jun 23.

Department of Molecular Genetics, Weizmann Institute of Science, Rehovot, 76100, Israel.

In humans, mutations in the PIEZO2 gene, which encodes for a mechanosensitive ion channel, were found to result in skeletal abnormalities including scoliosis and hip dysplasia. Here, we show in mice that loss of Piezo2 expression in the proprioceptive system recapitulates several human skeletal abnormalities. While loss of Piezo2 in chondrogenic or osteogenic lineages does not lead to human-like skeletal abnormalities, its loss in proprioceptive neurons leads to spine malalignment and hip dysplasia. To validate the non-autonomous role of proprioception in hip joint morphogenesis, we studied this process in mice mutant for proprioceptive system regulators Runx3 or Egr3. Loss of Runx3 in the peripheral nervous system, but not in skeletal lineages, leads to similar joint abnormalities, as does Egr3 loss of function. These findings expand the range of known regulatory roles of the proprioception system on the skeleton and provide a central component of the underlying molecular mechanism, namely Piezo2.
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http://dx.doi.org/10.1038/s41467-020-16971-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7311488PMC
June 2020

A minimal constraint device for imaging nuclei in live Drosophila contractile larval muscles reveals novel nuclear mechanical dynamics.

Lab Chip 2020 06 20;20(12):2100-2112. Epub 2020 May 20.

Department of Molecular Genetics, Weizmann Institute of Science, Israel.

Muscle contractions produce reiterated cytoplasmic mechanical variations, which potentially influence nuclear mechanotransduction, however information regarding the dynamics of muscle nuclei (myonuclei) in the course of muscle contraction is still missing. Towards that end, a minimal constraint device was designed in which intact live Drosophila larva is imaged, while its muscles still contract. The device is placed under spinning disc confocal microscope enabling imaging of fluorescently labeled sarcomeres and nuclei during muscle contraction, without any external stimulation. As a proof of principle we studied myonuclei dynamics in wild-type, as well as in Nesprin/klar mutant larvae lacking proper nuclear-cytoskeletal connections. Myonuclei in control larvae exhibited comparable dynamics in the course of multiple contractile events, independent of their position along the muscle fiber. In contrast, myonuclei of mutant larvae displayed differential dynamics at distinct positions along individual myofibers. Moreover, we identified a linear link between myonuclear volume and its acceleration values during muscle contraction which, in Nesprin/klar mutants exhibited an opposite tendency relative to control. Estimation of the drag force applied on individual myonuclei revealed that force fluctuations in time, but not the average force, differed significantly between control and Nesprin/klar mutant, and were considerably higher in the mutant myonuclei. Taken together these results imply significant alterations in the mechanical dynamics of individual myonuclei in the Nesprin/klar myonuclei relative to control. Such differences provide novel mechanical insight into Nesprin function in contractile muscles, and might reveal the mechanical basis underlying Nesprin-related human diseases.
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http://dx.doi.org/10.1039/d0lc00214cDOI Listing
June 2020

A minimal constraint device for imaging nuclei in live Drosophila contractile larval muscles reveals novel nuclear mechanical dynamics.

Lab Chip 2020 06 20;20(12):2100-2112. Epub 2020 May 20.

Department of Molecular Genetics, Weizmann Institute of Science, Israel.

Muscle contractions produce reiterated cytoplasmic mechanical variations, which potentially influence nuclear mechanotransduction, however information regarding the dynamics of muscle nuclei (myonuclei) in the course of muscle contraction is still missing. Towards that end, a minimal constraint device was designed in which intact live Drosophila larva is imaged, while its muscles still contract. The device is placed under spinning disc confocal microscope enabling imaging of fluorescently labeled sarcomeres and nuclei during muscle contraction, without any external stimulation. As a proof of principle we studied myonuclei dynamics in wild-type, as well as in Nesprin/klar mutant larvae lacking proper nuclear-cytoskeletal connections. Myonuclei in control larvae exhibited comparable dynamics in the course of multiple contractile events, independent of their position along the muscle fiber. In contrast, myonuclei of mutant larvae displayed differential dynamics at distinct positions along individual myofibers. Moreover, we identified a linear link between myonuclear volume and its acceleration values during muscle contraction which, in Nesprin/klar mutants exhibited an opposite tendency relative to control. Estimation of the drag force applied on individual myonuclei revealed that force fluctuations in time, but not the average force, differed significantly between control and Nesprin/klar mutant, and were considerably higher in the mutant myonuclei. Taken together these results imply significant alterations in the mechanical dynamics of individual myonuclei in the Nesprin/klar myonuclei relative to control. Such differences provide novel mechanical insight into Nesprin function in contractile muscles, and might reveal the mechanical basis underlying Nesprin-related human diseases.
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http://dx.doi.org/10.1039/d0lc00214cDOI Listing
June 2020

ETS Proteins Bind with Glucocorticoid Receptors: Relevance for Treatment of Ewing Sarcoma.

Cell Rep 2019 10;29(1):104-117.e4

Department of Biological Regulation, Weizmann Institute of Science, Rehovot 76100, Israel. Electronic address:

The glucocorticoid receptor (GR) acts as a ubiquitous cortisol-dependent transcription factor (TF). To identify co-factors, we used protein-fragment complementation assays and found that GR recognizes FLI1 and additional ETS family proteins, TFs relaying proliferation and/or migration signals. Following steroid-dependent translocation of FLI1 and GR to the nucleus, the FLI1-specific domain (FLS) binds with GR and strongly enhances GR's transcriptional activity. This interaction has functional consequences in Ewing sarcoma (ES), childhood and adolescence bone malignancies driven by fusions between EWSR1 and FLI1. In vitro, GR knockdown inhibited the migration and proliferation of ES cells, and in animal models, antagonizing GR (or lowering cortisol) retarded both tumor growth and metastasis from bone to lung. Taken together, our findings offer mechanistic rationale for repurposing GR-targeting drugs for the treatment of patients with ES.
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http://dx.doi.org/10.1016/j.celrep.2019.08.088DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6899513PMC
October 2019

Molecular pathways of senescence regulate placental structure and function.

EMBO J 2019 09 19;38(18):e100849. Epub 2019 Aug 19.

Department of Molecular Cell Biology, The Weizmann Institute of Science, Rehovot, Israel.

The placenta is an autonomous organ that maintains fetal growth and development. Its multinucleated syncytiotrophoblast layer, providing fetal nourishment during gestation, exhibits characteristics of cellular senescence. We show that in human placentas from pregnancies with intrauterine growth restriction, these characteristics are decreased. To elucidate the functions of pathways regulating senescence in syncytiotrophoblast, we used dynamic contrast-enhanced MRI in mice with attenuated senescence programs. This approach revealed an altered dynamics in placentas of p53 , Cdkn2a , and Cdkn2a ;p53 mice, accompanied by histopathological changes in placental labyrinths. Human primary syncytiotrophoblast upregulated senescence markers and molecular pathways associated with cell-cycle inhibition and senescence-associated secretory phenotype. The pathways and components of the secretory phenotype were compromised in mouse placentas with attenuated senescence and in human placentas from pregnancies with intrauterine growth restriction. We propose that molecular mediators of senescence regulate placental structure and function, through both cell-autonomous and non-autonomous mechanisms.
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http://dx.doi.org/10.15252/embj.2018100849DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6745498PMC
September 2019

Robo2 regulates synaptic oxytocin content by affecting actin dynamics.

Elife 2019 06 10;8. Epub 2019 Jun 10.

Department of Molecular Cell Biology, Weizmann Institute of Science, Rehovot, Israel.

The regulation of neuropeptide level at the site of release is essential for proper neurophysiological functions. We focused on a prominent neuropeptide, oxytocin (OXT) in the zebrafish as an in vivo model to visualize and quantify OXT content at the resolution of a single synapse. We found that OXT-loaded synapses were enriched with polymerized actin. Perturbation of actin filaments by either cytochalasin-D or conditional Cofilin expression resulted in decreased synaptic OXT levels. Genetic loss of or displayed decreased synaptic OXT content and mutants displayed reduced mobility of the actin probe Lifeact-EGFP in OXT synapses. Using a novel transgenic reporter allowing real-time monitoring of OXT-loaded vesicles, we show that mutants display slower rate of vesicles accumulation. OXT-specific expression of dominant-negative Cdc42, which is a key regulator of actin dynamics and a downstream effector of Robo2, led to a dose-dependent increase in OXT content in WT, and a dampened effect in mutants. Our results link Slit3-Robo2-Cdc42, which controls local actin dynamics, with the maintenance of synaptic neuropeptide levels.
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http://dx.doi.org/10.7554/eLife.45650DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6590984PMC
June 2019

Unmasking cellular response of a bloom-forming alga to viral infection by resolving expression profiles at a single-cell level.

PLoS Pathog 2019 04 24;15(4):e1007708. Epub 2019 Apr 24.

Department of Plant and Environmental Sciences, Weizmann Institute of Science, Rehovot, Israel.

Infection by large dsDNA viruses can lead to a profound alteration of host transcriptome and metabolome in order to provide essential building blocks to support the high metabolic demand for viral assembly and egress. Host response to viral infection can typically lead to diverse phenotypic outcome that include shift in host life cycle and activation of anti-viral defense response. Nevertheless, there is a major bottleneck to discern between viral hijacking strategies and host defense responses when averaging bulk population response. Here we study the interaction between Emiliania huxleyi, a bloom-forming alga, and its specific virus (EhV), an ecologically important host-virus model system in the ocean. We quantified host and virus gene expression on a single-cell resolution during the course of infection, using automatic microfluidic setup that captures individual algal cells and multiplex quantitate PCR. We revealed high heterogeneity in viral gene expression among individual cells. Simultaneous measurements of expression profiles of host and virus genes at a single-cell level allowed mapping of infected cells into newly defined infection states and allowed detection specific host response in a subpopulation of infected cell which otherwise masked by the majority of the infected population. Intriguingly, resistant cells emerged during viral infection, showed unique expression profiles of metabolic genes which can provide the basis for discerning between viral resistant and susceptible cells within heterogeneous populations in the marine environment. We propose that resolving host-virus arms race at a single-cell level will provide important mechanistic insights into viral life cycles and will uncover host defense strategies.
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http://dx.doi.org/10.1371/journal.ppat.1007708DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6502432PMC
April 2019

Asthma phenotypes and associated comorbidities in a large cohort of adolescents in Israel.

J Asthma 2020 07 24;57(7):722-735. Epub 2019 Apr 24.

Medical Corps, IDF, Israel.

Asthma is a multifactorial, heterogeneous, complex and common chronic respiratory disease driven by diverse mechanisms. Although asthma presents various clinical forms with different levels of severity, it is unclear whether asthma severities are a consequence of disease management or varied etiologies. We sought to investigate this question. This article presents a cross-sectional study of 113,671 Israeli adolescents. Univariate and multivariable logistic regression models were performed to analyze the independent associations between mild asthma and moderate-to-severe asthma phenotypes and coexistent medical conditions within each gender separately. Hierarchical clustering of the odds ratios of the diverse statistically significant medical conditions associated with asthma severity-gender groups was also performed. We focused on the allergic and neurological-cognitive-mental disorders. Among males, two associations were common to both asthma groups (atopic dermatitis and allergic rhinitis), five unique to mild asthma (urticaria/angioedema, Hymenoptera/bee allergies, allergic conjunctivitis, epilepsy and migraine) and two unique to moderate-to-severe asthma (learning disabilities and ADD/ADHD (Attention-deficit disorder/Attention-deficit/hyperactivity disorder)). Among females, two associations were common to both clinical asthma groups (allergic rhinitis and urticaria/angioedema), and five unique to moderate-to-severe asthma (atopic dermatitis, learning disabilities, ADD/ADHD, anxiety/mood disorders and migraine). Allergic rhinitis was the only condition to be associated with all four groups. Learning disabilities and ADD/ADHD were only associated with moderate-to-severe asthma (but not with mild asthma), in both males and females. Hierarchical clustering analysis uncovered two prominent clusters, separating mild from moderate-to-severe asthma. The differences between mild and moderate-to-severe asthma enhance asthma phenotype characterization, with respect to comorbidities, and indicate varied etiologies.
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http://dx.doi.org/10.1080/02770903.2019.1604743DOI Listing
July 2020

Single cell dissection of plasma cell heterogeneity in symptomatic and asymptomatic myeloma.

Nat Med 2018 12 6;24(12):1867-1876. Epub 2018 Dec 6.

Department of Hematology, Tel Aviv Sourasky Medical Center, Tel Aviv, Israel.

Multiple myeloma, a plasma cell malignancy, is the second most common blood cancer. Despite extensive research, disease heterogeneity is poorly characterized, hampering efforts for early diagnosis and improved treatments. Here, we apply single cell RNA sequencing to study the heterogeneity of 40 individuals along the multiple myeloma progression spectrum, including 11 healthy controls, demonstrating high interindividual variability that can be explained by expression of known multiple myeloma drivers and additional putative factors. We identify extensive subclonal structures for 10 of 29 individuals with multiple myeloma. In asymptomatic individuals with early disease and in those with minimal residual disease post-treatment, we detect rare tumor plasma cells with molecular characteristics similar to those of active myeloma, with possible implications for personalized therapies. Single cell analysis of rare circulating tumor cells allows for accurate liquid biopsy and detection of malignant plasma cells, which reflect bone marrow disease. Our work establishes single cell RNA sequencing for dissecting blood malignancies and devising detailed molecular characterization of tumor cells in symptomatic and asymptomatic patients.
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http://dx.doi.org/10.1038/s41591-018-0269-2DOI Listing
December 2018

LATS1 and LATS2 suppress breast cancer progression by maintaining cell identity and metabolic state.

Life Sci Alliance 2018 Oct 30;1(5):e201800171. Epub 2018 Oct 30.

Department of Molecular Cell Biology, Weizmann Institute of Science, Rehovot, Israel.

Deregulated activity of LArge Tumor Suppressor (LATS) tumor suppressors has broad implications on cellular and tissue homeostasis. We examined the consequences of down-regulation of either LATS1 or LATS2 in breast cancer. Consistent with their proposed tumor suppressive roles, expression of both paralogs was significantly down-regulated in human breast cancer, and loss of either paralog accelerated mammary tumorigenesis in mice. However, each paralog had a distinct impact on breast cancer. Thus, LATS2 depletion in luminal B tumors resulted in metabolic rewiring, with increased glycolysis and reduced peroxisome proliferator-activated receptor γ (PPARγ) signaling. Furthermore, pharmacological activation of PPARγ elicited LATS2-dependent death in luminal B-derived cells. In contrast, LATS1 depletion augmented cancer cell plasticity, skewing luminal B tumors towards increased expression of basal-like features, in association with increased resistance to hormone therapy. Hence, these two closely related paralogs play distinct roles in protection against breast cancer; tumors with reduced expression of either LATS1 or LATS2 may rewire signaling networks differently and thus respond differently to anticancer treatments.
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http://dx.doi.org/10.26508/lsa.201800171DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6238411PMC
October 2018

Bacterial virulence against an oceanic bloom-forming phytoplankter is mediated by algal DMSP.

Sci Adv 2018 10 24;4(10):eaau5716. Epub 2018 Oct 24.

Department of Plant and Environmental Sciences, Weizmann Institute of Science, Rehovot 7610001, Israel.

is a bloom-forming microalga that affects the global sulfur cycle by producing large amounts of dimethylsulfoniopropionate (DMSP) and its volatile metabolic product dimethyl sulfide. Top-down regulation of blooms has been attributed to viruses and grazers; however, the possible involvement of algicidal bacteria in bloom demise has remained elusive. We demonstrate that a strain, D7, that we isolated from a North Atlantic bloom, exhibited algicidal effects against upon coculturing. Both the alga and the bacterium were found to co-occur during a natural bloom, therefore establishing this host-pathogen system as an attractive, ecologically relevant model for studying algal-bacterial interactions in the oceans. During interaction, D7 consumed and metabolized algal DMSP to produce high amounts of methanethiol, an alternative product of DMSP catabolism. We revealed a unique strain-specific response, in which strains that exuded higher amounts of DMSP were more susceptible to D7 infection. Intriguingly, exogenous application of DMSP enhanced bacterial virulence and induced susceptibility in an algal strain typically resistant to the bacterial pathogen. This enhanced virulence was highly specific to DMSP compared to addition of propionate and glycerol which had no effect on bacterial virulence. We propose a novel function for DMSP, in addition to its central role in mutualistic interactions among marine organisms, as a mediator of bacterial virulence that may regulate blooms.
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http://dx.doi.org/10.1126/sciadv.aau5716DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6200362PMC
October 2018

A Mutant p53-Dependent Embryonic Stem Cell Gene Signature Is Associated with Augmented Tumorigenesis of Stem Cells.

Cancer Res 2018 10 28;78(20):5833-5847. Epub 2018 Aug 28.

Department of Molecular Cell Biology, The Weizmann Institute of Science, Rehovot, Israel.

Mutations in the tumor suppressor p53 are the most frequent alterations in human cancer. These mutations include p53-inactivating mutations as well as oncogenic gain-of-function (GOF) mutations that endow p53 with capabilities to promote tumor progression. A primary challenge in cancer therapy is targeting stemness features and cancer stem cells (CSC) that account for tumor initiation, metastasis, and cancer relapse. Here we show that cultivation of tumors derived from mutant p53 murine bone marrow mesenchymal stem cells (MSC) gives rise to aggressive tumor lines (TL). These MSC-TLs exhibited CSC features as displayed by their augmented oncogenicity and high expression of CSC markers. Comparative analyses between MSC-TL with their parental mutant p53 MSC allowed for identification of the molecular events underlying their tumorigenic properties, including an embryonic stem cell (ESC) gene signature specifically expressed in MSC-TLs. Knockout of mutant p53 led to a reduction in tumor development and tumorigenic cell frequency, which was accompanied by reduced expression of CSC markers and the ESC MSC-TL signature. In human cancer, MSC-TL ESC signature-derived genes correlated with poor patient survival and were highly expressed in human tumors harboring p53 hotspot mutations. These data indicate that the ESC gene signature-derived genes may serve as new stemness-based prognostic biomarkers as well as novel cancer therapeutic targets. Mesenchymal cancer stem cell-like cell lines express a mutant p53-dependent embryonic stem cell gene signature, which can serve as a potential prognostic biomarker and therapeutic target in cancer. .
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http://dx.doi.org/10.1158/0008-5472.CAN-18-0805DOI Listing
October 2018

Microglial MHC class II is dispensable for experimental autoimmune encephalomyelitis and cuprizone-induced demyelination.

Eur J Immunol 2018 08 7;48(8):1308-1318. Epub 2018 Jun 7.

Departments of Immunology, Weizmann Institute of Science, Rehovot, Israel.

Microglia are resident immune cells in the CNS, strategically positioned to clear dead cells and debris, and orchestrate CNS inflammation and immune defense. In steady state, these macrophages lack MHC class II (MHCII) expression, but microglia activation can be associated with MHCII induction. Whether microglial MHCII serves antigen presentation for critical local T-cell restimulation in CNS auto-immune disorders or modulates microglial signaling output remains under debate. To probe for such scenarios, we generated mice harboring an MHCII deficiency in microglia, but not peripheral myeloid cells. Using the CX CR1 -based approach we report that microglial antigen presentation is obsolete for the establishment of EAE, with disease onset, progression, and severity unaltered in mutant mice. Antigen presentation-independent roles of microglial MHCII were explored using a demyelination model induced by the copper chelator cuprizone. Absence of microglial I-A did not affect the extent of these chemically induced white matter alterations, nor did it affect microglial proliferation or gene expression associated with locally restricted de- and remyelination.
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http://dx.doi.org/10.1002/eji.201847540DOI Listing
August 2018

Expression profiling of host and virus during a coccolithophore bloom provides insights into the role of viral infection in promoting carbon export.

ISME J 2018 03 15;12(3):704-713. Epub 2018 Jan 15.

Departments of Plant and Environmental Sciences, Weizmann Institute of Science, Rehovot, 7610001, Israel.

The cosmopolitan coccolithophore Emiliania huxleyi is a unicellular eukaryotic alga that forms vast blooms in the oceans impacting large biogeochemical cycles. These blooms are often terminated due to infection by the large dsDNA virus, E. huxleyi virus (EhV). It was recently established that EhV-induced modulation of E. huxleyi metabolism is a key factor for optimal viral infection cycle. Despite the huge ecological importance of this host-virus interaction, the ability to assess its spatial and temporal dynamics and its possible impact on nutrient fluxes is limited by current approaches that focus on quantification of viral abundance and biodiversity. Here, we applied a host and virus gene expression analysis as a sensitive tool to quantify the dynamics of this interaction during a natural E. huxleyi bloom in the North Atlantic. We used viral gene expression profiling as an index for the level of active infection and showed that the latter correlated with water column depth. Intriguingly, this suggests a possible sinking mechanism for removing infected cells as aggregates from the E. huxleyi population in the surface layer into deeper waters. Viral infection was also highly correlated with induction of host metabolic genes involved in host life cycle, sphingolipid, and antioxidant metabolism, providing evidence for modulation of host metabolism under natural conditions. The ability to track and quantify defined phases of infection by monitoring co-expression of viral and host genes, coupled with advance omics approaches, will enable a deeper understanding of the impact that viruses have on the environment.
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http://dx.doi.org/10.1038/s41396-017-0004-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5864229PMC
March 2018

Fibroblast recruitment as a tool for ovarian cancer detection and targeted therapy.

Int J Cancer 2016 10 20;139(8):1788-98. Epub 2016 Jun 20.

Department of Biological Regulation, Weizmann Institute of Science, Rehovot, Israel.

Metastatic ovarian cancer, the most lethal of gynecologic malignancies, is typically managed by debulking surgery, followed by chemotherapy. However, despite significant efforts, survival rate remains low. We have previously demonstrated, in mouse models, a specific systemic homing of labeled fibroblasts to solid ovarian tumors. Here, we demonstrate the feasibility of utilizing this specific homing of genetically modified fibroblasts for detection and targeted therapy of orthotopic metastatic ovarian carcinoma model in immune-deficient mice. Using an in vivo metastatic mouse model for ovarian cancer, we demonstrated that fibroblasts expressing fluorescent reporters injected intra-peritoneally, were specifically recruited to peritoneal tumor nodules (resulting in 93-100% co-localization). We further used fibroblasts over expressing the soluble receptor variant of VEGFR1 (s-Flt1). Mice bearing tumors were injected weekly with either control or s-Flt1 expressing fibroblasts. Injection of s-Flt1 expressing fibroblasts resulted in a significant reduction in the ascites volume, reduced vascularization of adherent metastases, and improved overall survival. Using fluorescently labeled fibroblasts for tumor detection with readily available intra-operative fluorescence imaging tools may be useful for tumor staging and directing biopsies or surgical efforts during exploratory or debulking surgery. Fibroblasts may serve as a beacon pointing to the otherwise invisible metastases in the peritoneal cavity of ovarian cancer patients. Utilizing the recruited fibroblasts also for targeted delivery of anti angiogenic or antitumor molecules may aid in controlling tumor progression. Thus, these results suggest a novel approach for targeting ovarian tumor metastases for both tumor detection and therapy.
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http://dx.doi.org/10.1002/ijc.30209DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5565769PMC
October 2016

Adolescent BMI at Northern Israel: From Trends, to Associated Variables and Comorbidities, and to Medical Signatures.

Medicine (Baltimore) 2016 Mar;95(12):e3022

From the Medical Corps, Israel Defense Forces (IDF) (YM, DF, AP, ER, DD, YC), Tel HaShomer; The Weizmann Institute of Science (YM, RR), Rehovot; Shaare Zedek Medical Center (DF, RF, GW), Jerusalem; Schneider Children Medical Center (AP), Petach Tikvah; Assaf Harofeh Medical Center (OT), Zerifin; Israel Center for Disease Control (TS), Ministry of Health, Tel Aviv; Sackler School of Medicine (TS), Tel Aviv University, Tel Aviv, Israel.

The increasing prevalence of abnormal body mass index (BMI), mainly obesity, is becoming a significant public health problem. This cross-sectional study aimed to provide a comprehensive view of secular trends of BMI, and the associated socio-demographic variables and comorbidities among adolescents with abnormal BMI. Individuals of the study population were born mainly between 1970 and 1993, and were examined at 16 to 19 years of age during the years 1987 to 2010, at 1 conscription center in the northern district of Israel.The study population included 113,694 adolescents. Univariate and multivariable logistic regression models were used to investigate the associations between BMI categories, socio-demographic variables, and medical conditions.A downward trend in the prevalence of normal BMI among both male and female adolescents was obtained, while trends of overweight and obesity (in both genders) and underweight (only among females) rose. Socio-demographic variables such as religion, education, family-related parameters, residential environment, country of birth, and origin were all associated with different risks for abnormal BMI. Obesity was associated with higher risk for hyperlipidemia, endocrine disorders (only in males), knee disorders, and hypertension type I + II (in both genders). Overweight was associated with knee disorders (only in females). Underweight, exclusively in males, was associated with increased risk for endocrine disorders, proteinuria, and cardiac disorders. Hierarchical clustering analysis revealed the intricate relations between gender, BMI, and medical signatures. It brought to light novel clusters of diseases that were abundant among populations having above-normal BMI or underweight males. Furthermore, above-normal BMI was associated with a lower rate of cardiac anomalies and scoliosis/kyphosis, whereas being underweight was associated with a lower risk for hypertension and flat foot.This study provides a reliable and in-depth view of secular trends in height, weight, and BMI of male and female adolescents. It supports previous associations between abnormal BMI and demographic variables and comorbidities, while uncovering novel associations, mainly regarding medical signatures of each gender-BMI group. This might lead to better monitoring, early detection, prevention, and treatment of various conditions associated to abnormal BMI categories and gender groups.
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http://dx.doi.org/10.1097/MD.0000000000003022DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4998371PMC
March 2016

The LATS2 tumor suppressor inhibits SREBP and suppresses hepatic cholesterol accumulation.

Genes Dev 2016 Apr 24;30(7):786-97. Epub 2016 Mar 24.

Department of Molecular Cell Biology, The Weizmann Institute of Science, Rehovot 76100, Israel;

The Hippo signaling pathway is a major regulator of organ size. In the liver, Hippo pathway deregulation promotes hyperplasia and hepatocellular carcinoma primarily through hyperactivation of its downstream effector, YAP. The LATS2 tumor suppressor is a core member of the Hippo pathway. A screen for LATS2-interacting proteins in liver-derived cells identified the transcription factor SREBP2, master regulator of cholesterol homeostasis. LATS2 down-regulation caused SREBP activation and accumulation of excessive cholesterol. Likewise, mice harboring liver-specific Lats2 conditional knockout (Lats2-CKO) displayed constitutive SREBP activation and overexpressed SREBP target genes and developed spontaneous fatty liver disease. Interestingly, the impact of LATS2 depletion on SREBP-mediated transcription was clearly distinct from that of YAP overexpression. When challenged with excess dietary cholesterol, Lats2-CKO mice manifested more severe liver damage than wild-type mice. Surprisingly, apoptosis, inflammation, and fibrosis were actually attenuated relative to wild-type mice, in association with impaired p53 activation. Subsequently, Lats2-CKO mice failed to recover effectively from cholesterol-induced damage upon return to a normal diet. Additionally, decreased LATS2 mRNA in association with increased SREBP target gene expression was observed in a subset of human nonalcoholic fatty liver disease cases. Together, these findings further highlight the tight links between tumor suppressors and metabolic homeostasis.
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http://dx.doi.org/10.1101/gad.274167.115DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4826395PMC
April 2016

Posttranslational regulation of coordinated enzyme activities in the Pup-proteasome system.

Proc Natl Acad Sci U S A 2016 Mar 7;113(12):E1605-14. Epub 2016 Mar 7.

Department of Life Sciences, Ben-Gurion University of the Negev, Beer-Sheva 84105, Israel; The National Institute for Biotechnology in the Negev, Ben-Gurion University of the Negev, Beer-Sheva 84105, Israel;

The proper functioning of any biological system depends on the coordinated activity of its components. Regulation at the genetic level is, in many cases, effective in determining the cellular levels of system components. However, in cases where regulation at the genetic level is insufficient for attaining harmonic system function, posttranslational regulatory mechanisms are often used. Here, we uncover posttranslational regulatory mechanisms in the prokaryotic ubiquitin-like protein (Pup)-proteasome system (PPS), the bacterial equivalent of the eukaryotic ubiquitin-proteasome system. Pup, a ubiquitin analog, is conjugated to proteins through the activities of two enzymes, Dop (deamidase of Pup) and PafA (proteasome accessory factor A), the Pup ligase. As Dop also catalyzes depupylation, it was unclear how PPS function could be maintained without Dop and PafA canceling the activity of the other, and how the two activities of Dop are balanced. We report that tight Pup binding and the limited degree of Dop interaction with high-molecular-weight pupylated proteins results in preferred Pup deamidation over protein depupylation by this enzyme. Under starvation conditions, when accelerated protein pupylation is required, this bias is intensified by depletion of free Dop molecules, thereby minimizing the chance of depupylation. We also find that, in contrast to Dop, PafA presents a distinct preference for high-molecular-weight protein substrates. As such, PafA and Dop act in concert, rather than canceling each other's activity, to generate a high-molecular-weight pupylome. This bias in pupylome molecular weight distribution is consistent with the proposed nutritional role of the PPS under starvation conditions.
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http://dx.doi.org/10.1073/pnas.1525185113DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4812726PMC
March 2016

RNF20 Links Histone H2B Ubiquitylation with Inflammation and Inflammation-Associated Cancer.

Cell Rep 2016 Feb 4;14(6):1462-1476. Epub 2016 Feb 4.

Department of Molecular Cell Biology, The Weizmann Institute, Rehovot 7610001, Israel. Electronic address:

Factors linking inflammation and cancer are of great interest. We now report that the chromatin-targeting E3 ubiquitin ligase RNF20/RNF40, driving histone H2B monoubiquitylation (H2Bub1), modulates inflammation and inflammation-associated cancer in mice and humans. Downregulation of RNF20 and H2Bub1 favors recruitment of p65-containing nuclear factor κB (NF-κB) dimers over repressive p50 homodimers and decreases the heterochromatin mark H3K9me3 on a subset of NF-κB target genes to augment their transcription. Concordantly, RNF20(+/-) mice are predisposed to acute and chronic colonic inflammation and inflammation-associated colorectal cancer, with excessive myeloid-derived suppressor cells (MDSCs) that may quench antitumoral T cell activity. Notably, colons of human ulcerative colitis patients, as well as colorectal tumors, reveal downregulation of RNF20/RNF40 and H2Bub1 in both epithelium and stroma, supporting the clinical relevance of our tissue culture and mouse model findings.
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http://dx.doi.org/10.1016/j.celrep.2016.01.020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4761112PMC
February 2016

Recurrent inactivating RASA2 mutations in melanoma.

Nat Genet 2015 Dec 26;47(12):1408-10. Epub 2015 Oct 26.

Molecular Cell Biology Department, Weizmann Institute of Science, Rehovot, Israel.

Analysis of 501 melanoma exomes identified RASA2, encoding a RasGAP, as a tumor-suppressor gene mutated in 5% of melanomas. Recurrent loss-of-function mutations in RASA2 were found to increase RAS activation, melanoma cell growth and migration. RASA2 expression was lost in ≥30% of human melanomas and was associated with reduced patient survival. These findings identify RASA2 inactivation as a melanoma driver and highlight the importance of RasGAPs in cancer.
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http://dx.doi.org/10.1038/ng.3427DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4954601PMC
December 2015
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