Carl Schildkraut, PhD

Professor, Department of Cell Biology Chanin Bldg., Room 416 718 430-2097 schildkr@aecom.yu.edu   Biosketch Current laboratory members Selected list of publications Collaborators

Research interests Our laboratory is a part of the Einstein Center for Human Embryonic Stem Cell Research and the Cancer Center.   Epigenetic control of DNA replication and nuclear organization in mouse and human embryonic stem cells; reprogramming mammalian nuclei towards pluripotency.   Human ES cells offer promising potential treatments to a variety of diseases such as Parkinson's disease, MS, spinal cord injury, heart disease, and diabetes. To prevent immunorejection, the ideal approach is to generate human ES cells containing genetically identical DNA by reprogramming the matched somatic cell nucleus in a human oocyte. DNA replication reprogramming is an intrinsic part of nuclear reprogramming. Proper DNA replication reprogramming may improve the efficiency of somatic cell nuclear transfer, which is a major hurdle for therapeutic cloning. Our current work focuses on how the DNA replication program in mammalian cells changes as they differentiate along various developmental pathways. The DNA replication program includes when and where DNA replication initiates and in what direction the replication fork proceeds. While it is in theory possible for distinct cell types to replicate their genomes using the same DNA replication program, we and others have found that cells at different stages of development and cells of different tissues actually have very different DNA replication programs. For example, specific genes and entire loci replicate at different times during the S phase in embryonic stem (ES) cells compared to B cell progenitors. We have studied changes in replication timing at the immunoglobulin heavy chain (Igh) locus which codes for antibody genes (Norio et al. 2005) In order to change this replication-timing program, it is essential that there are concurrent changes in replication initiation sites within the Igh locus. For example, initiation sites that were previously silent in ES cells become activated with B cell development Our studies on DNA replication in mammalian cells have been facilitated by a new and unique way that we have devised, which uses fluorescence microscopy to observe purified individual DNA molecules. We call this approach single-molecule analysis of replicated DNA (SMARD) (Norio and Schildkraut, 2001). We use SMARD to observe the dynamics of replication across the entire length of long individual DNA molecules, revealing the positions of origins, termination sites, sites at which replication forks stall, and fork elongation rates (time to replicate a DNA segment). One of our present goals is to determine whether we can reprogram DNA replication by remodeling the chromatin that is assembled at the replication fork. We are making epigenetic changes in ES cells at sites where there are silenced replication origins and determining whether these changes result in activation of initiation sites and changes in replication timing during the S phase.   Long term interests:   • Regulation and reprogramming of DNA replication during human embryogenesis and in human embryonic stem (ES) cells. • How replication initiation sites affect early steps in human embryonic development. • What prevents reinitiation of DNA replication leading to the amplification of cancer causing genes. Current projects include a wide range of interests:   • Human ES cell DNA replication dynamics. • Reprogramming of DNA replication by modification of chromatin structure in the Igh locus in B lineage cells using gene targeting by Recombinase-Mediated Cassette Exchange (RMCE) in ES cells and transgenic mice   • Changes in nuclear organization and DNA replication during B cell development and their role in immunoglobulin heavy chain (Igh) gene rearrangement using Single Molecule Analysis of Replicated DNA (SMARD) • The role of histone acetylation in the regulation of Igh gene expression and DNA replication in the Igh locus. • Nuclear organization and DNA replication in primary cells from knock-out mice deficient in several B cell specific factors.   SILENCED REPLICATION ORIGINS ARE ACTIVATED DURING B CELL DEVELOPMENT   Shown above are images of individual DNA molecules after hybridization and immunostaining of the DNA molecules. The transition from red to green indicates the position of a replication fork (yellow arrow).These forks are shown at different stages of progression in fluorescent Igh DNA molecules from different cells.   A. Replication forks progress in one direction and no replication origins are detected within this DNA segment containing the Igh constant region genes. The yellow arrows show the direction in which replication forks progress. B. Replication origins are activated within this same DNA segment as development proceeds to the pro-B cell stage.

Recent publications Sfeir, A., Kosiyatrakul, S. T., Hockemeyer, D., MacRae, S. L., Karlseder, J., Schildkraut, C.L., and De Lange, T., Mammalian telomeres resemble fragile sites and require TRF1 for efficient replication. Cell, in press. 2009 Desprat, R., Thierry-Mieg, D., Lailler, N., Schildkraut, C. L., Thierry-Mieg, J., and Bouhassira, E. E., Predictable Dynamic Program of Timing of DNA Replication in Human Cells. Under revision for Genome Research. Schultz, S. S., Desbordes, S. C., Du, Z., Kosiyatrakul, S.T., Lipchina, I., Studer, L., and Schildkraut, C. L. Single Molecule Analysis Reveals Developmental Changes in the DNA Replication Program for the POU5F1 Locus. Under review Guan, Z., Hughes, C. M., Kosiyatrakul, S. T., Norio, P., Sen, R., Fiering, S., Allis, C. D Bouhassira, E. E., and Schildkraut, C. L. Replication Temporal Transition Region Suppresses Replication Origin Activity. Under review Norio, P., Kosiyatrakul, S.,Yang, Q., Guan, Z.,Brown, N., Thomas, S., Riblet, R., and Schildkraut, C.L. Progressive activation of DNA replication initiation in large domains of the immunoglobulin heavy chain locus during B cell development. Mol. Cell 20: 575 - 587(2005). Zhou, J., Saleque, S., Ermakova, O., Sepulveda, M.A., Yang, Q., Eckhardt, L.A., Schildkraut, C.L. and Birshtein, B.K. Changes in replication, nuclear location and transcription of the Igh locus after fusion of a pre B cell line with a T cell line. J. Immunology.,175: 2317-2320 (2005). Yang, Q., Riblet, R., Schildkraut, C.L. Sites that Direct Nuclear Compartmentalization are Near the 5' End of the Mouse Immunoglobulin Heavy Chain Locus. Mol. Cell. Biol. 25: 6021-6030 (2005). Norio P, Schildkraut C.L. Plasticity of DNA replication initiation in Epstein-Barr virus episomes. PLoS Biol 2: 816 - 833. (2004). Zhou J., Ermakova, O., Riblet, R., Birshtein, B. and Schildkraut, C.L. Replication and Subnuclear Location Dyamics of the Immunoglobulin Heavy-Chain Locus in B-Lineage Cells. Mol. Cell. Biol., 22:4876-4889 (2002) Norio, P., Schildkraut, C.L. Visualization of DNA Replication on Individual Epstein-Barr Virus Episomes. Science 294: 2361-2364 (2001).

Carl Schildkraut: Research interests | Biosketch Faculty research at a glance Birshtein | Bouhassira | Edelmann | Fyodorov | Keogh | Kielian | Kitsis | Nathenson | Query Scharff | Schildkraut | Shafritz | Singer | Skoultchi | Stanley | Steidl |Warner | Ye   Home