Marisa Bartolomei, Ph.D.

Mechanism of Genomic Imprinting: As a postdoctoral fellow in the laboratory of Dr. Shirley Tilghman, I identified the one of the first imprinted genes—the H19 gene and also determined that this gene was adjacent to the imprinted Igf2 gene, representing the first evidence of imprinted gene clusters. In my own laboratory, we study the mechanisms governing imprinted gene expression and have determined that imprinted genes and clusters are regulated by differentially methylated imprinting control regions [ICRs]. In the case of H19 and Igf2, the ICR is a paternally methylated CTCF-dependent insulator. Through the use of cis-acting mutations we have characterized imprinting at the H19 locus and have elucidated elements that are important for setting the DNA methylation imprint in the germline and maintaining imprinting in the embryo after fertilization. Our mouse models contributed to the discovery that epigenetic mutations in the H19 ICR caused Silver-Russell Syndrome in a subset of patients. Additionally, microdeletions in the H19 ICR have been identified in Beckwith-Wiedemann Syndrome. We are generating mouse models as well as using iPS cells to study patient-derived mutations. 

• Bartolomei, M.S., Zemel, S. and S.M. Tilghman. (1991). Parental imprinting of the mouse H19Nature 351:153-155.
• Thorvaldsen, J.L., Duran, K.L. and M.S. Bartolomei. (1998). The H19differentially methylated domain provides multiple regulatory functions in H19 and Igf2 reciprocal imprinting. Genes & Development12:3693-3702.
• Engel, N., West, A.G., Felsenfeld, G., and M.S. Bartolomei. (2004). Antagonism between DNA hypermethylation and enhancer-blocking activity at the H19DMD is uncovered by CpG mutations. Nature Genetics, 36:883-888.
• Ideraabdullah, F.Y., Thorvaldsen, J.L., Myers, J.A. and M.S. Bartolomei. (2014). Tissue specific insulator function at H19/Igf2 revealed by deletions at the imprinting control region. Human Molecular Genetics, 23:6246-6259. PMCID: PMC4222363

Discovery that mouse models epigenetic perturbations identified in Assisted Reproductive Technologies (ART): In the imprinting field, experiments performed in the 1990’s were aimed at elucidating when monoallelic expression was initially set for imprinted genes. Two studies in our laboratory and in the Surani laboratory gave contrasting results for the H19 gene, with our lab showing maternal-specific expression when the gene was activated in the blastocyst and the Surani lab describing biallelic expression at this time. We subsequently determined that culturing mouse preimplantation embryos was associated with loss of imprinted gene expression. It was later shown that ART was associated with a higher than expected number of imprinting syndromes, including Beckwith-Wiedemann Syndrome and Angelman Syndrome, with almost all documented cases caused by loss of DNA methylation of the ICR. The mouse, which has the added benefit of normal fertility, has proven a valuable model to address these observations and determine mechanisms with the eventual goal of improving ART outcomes. We have shown that in vitro culture, embryo transfer, in vitro fertilization and hormonal hyperstimulation contribute to errors in epigenetic gene regulation. We have also determined that placental tissues are especially sensitive to the manipulations involved in ART.

• Doherty, A.S., Mann, M.R.W., Tremblay, K.D., Bartolomei, M.S. and R.M. Schultz. (2000). Differential effects of culture on imprinted H19 expression in the preimplantation mouse embryo. Biology of Reproduction, 62:1526-1535.
• Mann, M.R.W., Lee, S.S., Doherty, A.S., Verona, R.I., Nolen, L.D., Schultz, R.M., and M.S. Bartolomei. (2004). Selective loss of imprinting in the placenta following preimplantation development in culture. Development,131:3727-3735.
• Rivera, R.M., Stein, P., Weaver, J.R., Mager, J., Schultz, R.M. and M.S. Bartolomei. (2008). Manipulations of mouse embryos prior to implantation result in aberrant expression of imprinted genes on day 9.5 of development. Human Molecular Genetics, 17:1-14.
• deWaal, E., Mak, W., Calhoun, S., Stein, P., Ord, T., Krapp, C., Coutifaris, C., Schultz, R.M. and M.S. Bartolomei. (2014). In vitro culture increases the frequency of stochastic epigenetic errors at imprinted genes in placental tissues from mouse concepti produced through assisted reproductive technologies, Biology of Reproduction, 90:1-12. PMCID: PMC4076403

Generation of a mouse model to study endocrine disrupting compounds (EDCs): Numerous experiments in mouse model systems as well as human studies have suggested that exposure to EDCs is associated with aberrant DNA methylation. Because imprinted genes are regulated by DNA methylation, we tested whether imprinting could be disrupted when pregnant dams were exposed to levels of BPA that are comparable to human exposure. We demonstrated that genomic imprinting and DNA methylation of ICRs and repetitive DNA were perturbed in offspring exposed in utero, with a pronounced effect in placenta. We have also shown that metabolic phenotypes observed in F1 offspring exposed in utero were transmitted to their F2 offspring, thus exhibiting multigenerational transmission of the phenotype.

• Susiarjo, M., Sasson, I., Mesaros, C. and M.S. Bartolomei. (2013). Bisphenol A exposure disrupts genomic imprinting in the mouse. PLOS Genetics, 9:e1003401. PMCID: PMC3616904
• Susiarjo, M., Xin, F., Bansal, A., Stefaniak, M., Li, C., Simmons, R.A. and M.S. Bartolomei. (2015). Bisphenol A exposure disrupts metabolic health across multiple generations in the mouse. Endocrinology, Mar 25:en20142027. [Epub ahead of print]. PMCID: In progress

CTCF and insulators: CTCF was initially described as a factor that functioned in transcriptional repression and activation. The first demonstration of its insulator activity was by the Felsenfeld laboratory at the globin locus. It soon became apparent that there were CTCF sites in the H19/Igf2 ICR and that this ICR exhibited insulator activity on the maternal allele. To study the role of CTCF at H19 we generated a knockdown line and showed that in the absence of CTCF, H19 became hypermethylated, and also that embryos died early in development. This study was the first that reported the essential role for CTCF in development. Further experiments demonstrated the extensive role of CTCF in gene activity as well as a role for CTCF interacting with cohesins at a number of developmentally important insulators and regulatory regions. We have also studied CTCF binding in an ES cell differentiation model and elucidated sequence preferences both genomically and biochemically, contributing to the knowledge of the role of CTCF in early developmental decisions, as suggested by the knockdown studies.

• Fedoriw, A.M, Stein, P., Svoboda, P., Schultz, R.M. and M.S. Bartolomei. (2004). Transgenic RNAi reveals essential function for CTCF in H19gene imprinting. Science, 303:238-240.
• Wan, L-B., Pan, H., Hannenhalli, S., Cheng, Y., Ma, J., Fedoriw, A., Lobanenkov, V., Latham, K.E., Schultz, R.M. and M.S. Bartolomei. (2008). Maternal depletion of CTCF reveals multiple functions during ooycte and preimplantation embryo development. Development, 135:2729-2738. PMCID: PMC2596970
• Stedman, W., Kang, H., Lin, S., Kissil, J.L., Bartolomei, M.S. and P.M. Lieberman. (2008). Cohesins localize with CTCF at the KSHV latency control region and at cellular c-Myc and H19/IGF2 insulators, EMBO Journal, 27:654-666. PMCID: PMC2262040
• Plasschaert, R.N., Vigneau, S., Tempera, I., Gupta, R., Maksimoska, J., Everett, L., Davuluri, R., Marmorstein, R., Lieberman, P.M., Schultz, D., Hannenhalli, S. and M.S. Bartolomei. (2014). CTCF binding site sequences differences are associated with unique regulatory and functional trends during embryonic stem cell differentiation, Nucleic Acids Research, 42:774-789. PMCID: PMC3902912

Role of DNA methylation in epigenetic gene regulation in the mouse embryo: In early work as a postdoctoral fellow with Dr. Shirley Tilghman, I demonstrated that DNA methylation was critical for conferring parental identity of imprinted genes. Subsequently, my laboratory demonstrated that DNA methylation was erased in primordial germ cells by the time they enter the gonad and established in a sex-specific manner. We and others have also demonstrated that loss of the maintenance methyltransferase DNMT1 disrupts imprinted gene expression in the embryo as well as the placenta. More recently we have collaborated with a number of groups to understand the mechanism by which DNA methylation imprints are established and erased in the germline, including studies on the role of TDG and unpublished studies on the Tet enzyme family.

• Bartolomei, M.S., Webber, A.L., Brunkow, M.E. and S. M. Tilghman. (1993). Epigenetic mechanisms underlying the imprinting of the mouse H19Genes and Development, 7:1663-1673.
• Davis, T.L., Yang, G.J., McCarrey, J.R. and M.S. Bartolomei. (2000). The H19methylation imprint is erased and reestablished differentially on the parental alleles during male germ cell development. Human Molecular Genetics, 9:2885-2894.
• Weaver, J.R., Sarkisian, G., Krapp, C., Mager, J., Mann, M.R. and M.S. Bartolomei. (2010). Domain-specific response of imprinted genes to reduced DNMT1. Molecular and Cellular Biology, 30: 3916-3928. PMCID: PMC2916450
• Cortellino, S., Xu, J., Sannai, M., Moore, R., Caretti, E., Cigliano, A., Le Coz, M., Devarajan, K., Wessels, A., Soprano, D., Abramowitz, L.K., Bartolomei, M.S., Rambow, F., Bassi, M.R., Bruno, T., Fanciulli, M., Renner, C., Klein-Szanto, A.J., Matsumoto, Y., Kobi, D., Davidson, I., Alberti, C., Larue, L. and A. Bellacosa. (2011). Thymine DNA glycosylase is essential for active DNA demethylation by linked deamination-base excision repair. Cell, 146: 67-79. PMCID: PMC3230223