Dr. Sharon Schlesinger

In the Shlez lab, we study the epigenetic reprogramming following fertilization. In particular, we are interested in the process leading to the formation of heterochromatin de novo and silencing of transposable elements and endogenous retroviruses. We use embryonic stem (ES) cells from mice and from farm animals to get a deeper understanding of the general characteristics of pluripotency in all mammalian cells.

 

 

The Schlesinger lab – research interests:

 

Epigenetic regulation in embryonic stem cells

 

Retroviral silencing in embryonic stem cells

 

Bovine pluripotent stem cells

 

Chronic inflammation alters DNA methylation patterns in the human gut

 

Generation and characterization of Mesenchymal Stem Cells for use in cell therapy

 

 

many

 

 

 

Curriculum Vitae

Short biography:

1997 – 2000 B.Sc. in Life Sciences, "Etgar" project for excellent students. the Hebrew University, Jerusalem, Israel.

2000 – 2001 M.Sc. studies, The Hebrew University- Hadassah Medical School, Jerusalem, Israel.

2001 – 2008 Ph.D. Molecular Biology and Genetics, The Hebrew University- Hadassah Medical School, Jerusalem, Israel. Prof. Howard Cedar and Prof. Yehudit Bergman, Thesis Title: Allelic exclusion of ribosomal RNA genes.

2009 - 2014 Postdoctoral research fellow, Columbia University Medical Center, NY,USA. Prof. Stephen P. Goff, Department of Biochemistry and Molecular Biophysics. Characterization of the Retroviral Silencing Machinery in Embryonic Stem Cells

2014 – 2015 Research associate, The Hebrew University of Jerusalem Edmond J. Safra Campus (Givat Ram). Department of Genetics, The Institute of Life Sciences. Host: Prof. Eran Meshorer

2015 to date Senior lecture (tenure track), The Hebrew University of Jerusalem Department of animal science, Robert H. Smith Faculty of Agriculture, Food, and Environment, Rehovot, Israel.

 

nanograesc

 

List of Publications

Google Scholar List of Publications

Reviews

• Schlesinger S and Goff SP. 2015. Retroviral Transcriptional Regulation and Embryonic Stem Cells: War and Peace. Molecular and Cellular Biology 35(5):770-777. Epub 2014 Dec 29

Articles

• Schlesinger S , Kaffe B, Melcer S, Aguilera JD, Sivaraman DM, Kaplan T, Meshorer E. A hyperdynamic H3.3 nucleosome marks promoter regions in pluripotent embryonic stem cells. Nucleic Acids Res. 2017 Sep 25.

• Yang BX, El Farran CA, Guo HC, Yu T, Fang HT, Wang HF, Schlesinger S, Seah YF, Goh GY, Neo SP, Li Y, Lorincz MC, Tergaonkar V, Lim TM, Chen L, Gunaratne J, Collins JJ, Goff SP, Daley GQ, Li H, Bard FA and Loh YH. Systematic Identification of Factors for Provirus Silencing in Embryonic Stem Cells. Cell, 2015. 163(1): p.230-45.

• Schlesinger S, Meshorer E and Goff SP. 2014. Asynchronous transcriptional silencing of individual retroviral genomes in embryonic cells Retrovirology, 11(1):31

• Schlesinger S, Lee A, Wang GZ, Green L and Goff SP. 2013. Proviral Silencing in Embryonic Cells is Regulated by Yin Yang 1. Cell Reports, 4(1):50-8

• Schlesinger S and Goff SP. 2013. Silencing of Proviral Expression in Embryonic Cells: Efficiency, Stability, and Chromatin Modifications. Embo Reports. 14:73-79.

• Farago M, Rosenbluh C, Tevlin M, Fraenkel S, Schlesinger S, Masika H, Gouzman M, Teng G, Schatz D, Rais Y, Hanna JH, Mildner A, Jung S, Mostoslavsky G, Cedar H, Bergman Y., 2012. Clonal allelic predetermination of immunoglobulin-kappa rearrangement. Nature. 490(7421): 561–565.

• Schlesinger S, Selig S, Bergman Y, Cedar H., 2009. Allelic inactivation of rDNA loci. Genes and Development. 23(20):2437-47. Selected for Faculty of 1000.

• Mostoslavsky, R., Singh, N., Tenzen, T., Goldmit, M., Gabay, C., Elizur, S., Qi, P., Reubinoff, B.E., Chess, A., Cedar, H., and Bergman, Y., 2001. Asynchronous replication and allelic exclusion in the immune system. Nature. 414: p. 221-225

 

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Research

In the Shlez lab, we study the epigenetic reprogramming following fertilization. In particular, we are interested in the process leading to the formation of heterochromatin de novo and silencing of transposable elements and endogenous retroviruses. We use embryonic stem (ES) cells from mice and from farm animals to get a deeper understanding of the general characteristics of pluripotency in all mammalian cells.

 

Publications in Google Scholar

The Schlesinger lab – research interests:

Epigenetic regulation in embryonic stem cells

Epigenetic regulation in embryonic stem cells

Transposable elements regulation in ES cells

Retroviral silencing in embryonic stem cells

Bovine pluripotent stem cells

Bovine pluripotent stem cells

Chronic inflammation alters DNA methylation patterns in the human gut

Chronic inflammation alters DNA methylation patterns in the human gut

Generation and characterization of Mesenchymal Stem Cells for use in cell therapy

Generation and characterization of Mesenchymal Stem Cells for use in cell therapy

 

Epigenetic regulation in embryonic stem cells

Epigenetic regulation in embryonic stem cells

In mammals, fertilization of an oocyte by the sperm is followed by epigenetic reprogramming, which involves de novo acquisition of chromatin signatures in the two parental genomes. The molecular determinants underlying such reprogramming are not fully understood. In particular, the formation of heterochromatin de novo and the silencing of specific genomic loci is thought to be essential to ensure the subsequent organization of the embryonic epigenome and embryonic development.

Embryonic stem cell (ES cell) lines were first generated by culturing mouse inner cell mass (ICM) explants on feeder layers in 1981, and are being used since then as the model system to study pluripotency. They are unique among primary cells in that they can give rise to all cell types of the body and have a very high self-renewing capacity.

In my research, I study the various epigenetic properties of ES cells and their changes during differentiation.

Chromatin has recently emerged as a key-contributing factor to embryonic stem cell identity and plasticity. There is evidence that both unique features of ES cells - self renewal and pluripotency - are regulated to a large extent by epigenetic modifications. We aim to identify, characterize and functionally study epigenetic factors and chromatin dynamics that govern pluripotency in both human and murine ES cells.

I- Elucidating histone turnover rate in the genome of pluripotent and differentiated cells. While histones are generally among the most stably associated DNA-binding proteins known, a subset of histones exhibit dynamic exchange with the soluble pool of nucleoplasmic histones. In all eukaryotes studied, histone H3 exchange is most rapid at promoters, and is generally slowest over heterochromatic regions. Dynamic histone turnover is linked to a variety of key aspects of chromatin biology. In ES cells this dynamic interaction is enhanced, thus maintaining a hyper-dynamic chromatin state in pluripotent ES cells. This hyper-dynamic state has been proposed to maintain the ES cell genome accessible as a relatively permissive ground state that becomes ‘‘locked down’’ during the process of lineage commitment and subsequent differentiation. Surprisingly, FRAP analysis of the H3 variants show that H3.1 is indeed hyper-dynamic in ES cells, while H3.3 has slow turnover rate in both embryonic and differentiated cells. This is somewhat counterintuitive as H3.3 is specifically enriched at transcriptionally active genes and regulatory elements, where a more dynamic state would be anticipated. What is the functional link between H3 turnover rate and expression levels in ES cells? And what is the meaning of the correlation between hyper-dynamic epigenetic state and heterogeneous (or stochastic/noisy) gene expression50 observed often in ES cells?

Retroviral silencing in embryonic stem cells

Nearly half of the mammalian genome is derived from mobile genetic elements, including members of the long terminal repeat (LTR)-containing endogenous retroviruses (ERVs), and the non-LTR-containing retrotransposons (LINE-1 and SINE-1). These genetic elements are considered a driving force in vertebrate evolution and there are many examples of retroviral genes, non-coding RNAs or promoters that are used by the genome of their hosts. Hence, the expression of these elements must be tightly regulated during embryonic development to balance the potentially damaging effects of widespread retrotransposition against the benefits of promoting genetic diversity. The nature of this regulation is not fully understood.

ES cells have the remarkable ability to potently suppress the expression of endogenous and exogenous retroviral sequences. This transcriptional silencing is critical to the maintenance of their genetic stability, since it allows successful differentiation into mature cell types without lethal genotypic damage from retroviral gene integration.

In my research, I study the various mechanisms of epigenetic silencing in ES cells and during their development. Using retroviral vectors, I have characterized two distinct retroviral silencing mechanisms and their kinetics of action. I showed that following retroviral infection, the establishment of silencing is dependent on the protein Yin Yang 1 (YY1), and that the loss of this protein leads to epigenetic changes of the proviral DNAs, including long-lasting loss of DNA methylation. Together, my results suggest that the study of retroviral silencing provides a useful tool for dissecting the molecular details of the ability of ES cells to mark genes for silencing - a crucial aspect of ES cell identity.

 

Examples for domestication events of ERV sequences by mouse human embryonic cells.

MERV-L elements and their remnant ‘solo’ long terminal repeats (LTRs) have coopted to participate in gene-regulatory networks, by serving as primary alternative promoters of nearby genes. A subset of mouse embryonic stem expresses MERV-L LTR-driven chimeric transcripts, which correlates with increased potency. (b) HERV-H interacts with Oct4 to promote enhancer activity of LTR7 nearby regions and to drive the expression of neighboring lncRNAs and protein-genes essential to hESC identity. (c) ERV1 elements in human and mouse genome carry transcription factor-binding sites for OCT4 and Nanog, which can regulate form the pluripotency network near insertion sites, leading to novel regulatory patterns in evolving mammals.

 

 

Bovine pluripotent stem cells

 

Stem cell (SC) research has been around for nearly 40 years, but only human and mouse SCs have been characterized, have consensus protocols and standardized procedures. For other animals, including domestic ones, this is not the case. There is no established methodology and big efforts are being made by the scientific community in this direction, our lab included. Pluripotent stem cells (PSCs) have the capacity to self-renew indefinitely and to develop into the three primary germ cell layers and therefore to form all lineages of the body.

PSCs can be derived using two parallel approaches: (1) the Inner Cell Mass (ICM) of a bovine blastocyst and (2) reprogramming somatic cells to pluripotent state by ectopic expression of various pluripotency factors -e.g. induced pluripotent stem cells (iPS). Currently, our research focuses on producing iPS from bovine fetal fibroblasts (BFF). In this process, the fibroblasts, which are somatic cells, undergo reprogramming to PSCs.

Bovine fetuses of Holstein cows were obtained from a local abattoir. The fibroblastic tissue was processed under sterile conditions and either directly plated with growth media or digested using a mix of 4 enzymes to obtain the fibroblast cells and plate with the media. The cells were grown for 2 passages and then infected with 7 lentivirus vectors for pluripotent genes. The vectors were then induced with Doxycycline.  Four weeks later, the colonies were isolated and grown separately with appropriate growth conditions for stem cells. RNA was extracted from the colonies at passages 1-2 and qRT-PCR was performed to assess pluripotency. Additionally, IF was performed.

We screened for 11 medias that can support infected fibroblasts reprograming to iPS, and identified 4 potential medias for further examination.  Using these four medias, we managed to culture partly pluripotent cells.

Bovine fibroblast reprograming:
A: RT-PCR results for 3 pluripotent markers. Graph columns: C - different medias, ICM (inner cell mass) – positive control, BFF (bovine fetus fibroblasts) - negative controls .B: IF for 4c media using different antibodies. DAPI labels DNA. Images were taken in confocal microscope (x63, Zeiss).

 

 

Next, we will use the knowledge and skills gained toward the generation of true pluripotent embryonic stem cells from bovine and other domesticated animals. We plan to examine the molecular and epigenetic characteristics of these bovine ESC and compare them to human and mouse ESCs.

Embryonic stem cell-based approach is expected to have many applications in domestic animals such as dogs, pigs, goats and cows. Large animals are considered excellent models for long-term experiments in regenerative medicines and biomedical research in general, because of their similarities in physiology with humans as contrasted with the laboratory mouse or rat. Thus, establishing of naïve embryonic stem cell lines from domesticated animals can benefit both clinical applications to improve human health and agricultural applications.

Chronic inflammation alters DNA methylation patterns in the human gut

Chronic inflammation alters DNA methylation patterns in the human gut

Aberrant methylation following inflammation events in the human gut might result in dysregulated cellular functions and an impaired host-microbial homeostasis. During pediatric development, these changes in the methylome will affect the predisposition of the individual to IBD and CRC. We are using a mouse model that allows experimental interrogation of human gut in vivo and provides a unique opportunity to examine the methylation changes and their long-term effect in the developing human gut.

We Aim to (1) Elucidate changes of DNA methylation in the human gut epithelium during organ development. (2) Explore the effect of chronic tissue inflammation on DNA methylation landscape in the human gut. (3) Validate the results using organoid culture. Combine the data acquired with available online data to define ‘developmental window’ in which DNA methylation changes might affect the predisposition to IBD and CRC.

Inflammatory insults are frequently associated with tumorigenesis, and aberrant DNA methylation is a hallmark of tumor development. Here, we study the link between these two and suggest that minor early life events can influence the steady state of the gut and contribute to tumorigenesis. Our long term goal is to elucidate the mechanism through which environmental ques influence the epigenome and thus predisposition to cancer development.

inflammation
ראה גם: Sharon Schlesinger


Fetal human intestine DNA methylation pattern is being formulated along maturation.
(A) Graphic overview of the experiment design; human fetal intestine will be transplanted subcutaneously into SCID mice and mature for 12-16 weeks. Mature xenografts will be challenged with MAP infection for either 10 days or three month. Matching controls will be treated with PBS. Intestine epithelial cells (IEC) from the implants will be analyzed for RNA expression and DNA methylation. green square denotes preliminary data shown. Kmer analysis for tiles hyper (B) and hypo (C) methylated with age. IEC from human fetal intestine and from mature xenografts were subjected to RRBS analysis revealing genomic regions that are hyper (B) or hypo (C) methylated with age. (D) Age related hyper methylated genes were subjected to GO biological process enrichment using PantherDB, all enriched hits are listed and regulatory pathways are emphasized.

 

Generation and characterization of Mesenchymal Stem Cells for use is cell therapy

Cell therapy, therapy in which cells are used for treatment, is the subject of intense research and presents great potential. Specifically, mesenchymal stem cells (MSC) are emerging as an effective and safe cell-based therapy for the treatment and prevention of many infectious and inflammatory conditions affecting humans and animals. MSCs are non-hematopoietic multipotent stem cells, belonging to the mesoderm layer which is one of the three germ layers. MSCs are most frequently derived from adult tissue sources such as bone marrow and adipose tissue or from birth associated tissue such as placental tissue, amnion, umbilical cord and cord blood, and poses the ability of multi-lineage differentiation into cell types such as adipocytes, osteoblasts, chondrocytes, myocytes, β-pancreatic islets cells and neuronal cells.

In our lab we generate MSCs from bovine umbilical cord, placenta and adipose tissue and murine bone marrow, and characterize them by testing their immunomodulation, their ability to differentiate into adipocytes and osteoblasts, we test their gene expression profile by means of RT-PCR and RNA seq and their epigenetic landscape by bisulfite sequencing and Chromatin Immunoprecipitation (ChIP).

Following systemic and local administration, allogeneic MSCs were shown to target inflamed tissues and to improve cure by immunomodulation and better bacterial clearance. We propose here to study the safety, feasibility and efficacy of MSCs for the treatment of inflammatory diseases such as mastitis and metritis (udder and uterine inflammation) in dairy cows. We hypothesize that allogeneic MSCs will populate inflamed organs and will alleviate inflammation and improve bacteriological cure.

Mastitis, an inflammatory response of the mammary tissue to invading pathogenic bacteria, is a major animal health and welfare problem in the dairy industry and is responsible for multibillion dollar of economic losses. Although improved hygiene and management techniques reduced and even eradicated some forms of the disease, mastitis still prevail in all dairy farms. Currently, the administration of antibiotics is the most common method of treatment and prevention of mastitis. However, this strategy has many disadvantages including low cure rate, increasing occurrence of bacterial resistance, and the presence of antibiotics residues in the milk. Therefore, alternative effective approaches for management of bovine mastitis should be looked for and tested.

(A) Changes in bWJ-MSC marker expression during culture time and passages. Note the decrease at p5. CD45 – leucocytes marker. Ctrl – PSMB gene. (B) Co-culture of bovine MSC (BUC – bovine umbilical cord, WJ – Warton jelly, at different passages) with peripheral blood cells inhibit the activation and proliferation of T cells. (C) bUC-MCS (green) in a cross talk with mammary epithelial cells (red – actin filaments, blue – DNA). (D) bUC-MSC (error head) exchange mitochondria (red) and cytoplasmic material (green) with epithelial cells

Lab members

Dr. Sharon Schlesinger – PI

Sharon.shle@mail.huji.ac.il

 

Dr. Carmit Strauss - Lab manager

carmit.feliks@mail.huji.ac.il

 

Dr. Myah Goldstein - post-doc

myah.goldstein@mail.huji.ac.il

 

Ivana Ribarski-Chorev – PhD student

Ivana.Ribarski-Chorev@mail.huji.ac.il

 

Ayellet Tal – PhD student

ayellet.tal@mail.huji.ac.il

 

 

Chen Shimoni - Msc student

chen.shimoni@mail.huji.ac.il

 

Andres Bernys– international MSc student

wladimir.bernys@mail.huji.ac.il

 

Iftach Schouten - MSc student (with Prof. Nahum Shpigel)

iftach.schouten@mail.huji.ac.il

 

Undergrads working in the lab:

Bar Kaufman (with Prof. Nahum Shpigel)

 

Efrat Harel

 

Alumni

Liad Margalit (MSc)

Yaniv Alon (MSc)

Jose David Aguirre Aguilera (MSc)

 

 

 

Funding

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