The entire genome is divided into individual units (chromosomal bands) whose replication timing in the cell cycle is under developmental control. This process plays a wide spread role in the regional regulation of expression, both for tissue specific genes and as a differential marker of monoallelic transcription as seen in genomic imprinting and allelic exclusion. Although little is known about this process, a new approach which takes advantage of Fluorescence In Situ Hybridization (FISH) to interphase nuclei should now allow us to decipher the basic elements which direct replication timing in vivo. Preliminary data indicates that sequences located within the Locus Control Region (LCR) of the human globin domain are involved in regulating replication timing during development. The investigator has developed a new tissue culture assay which can be employed for deciphering the exact cis acting elements which mediate this effect, and this will pave the way towards cloning the trans acting factors that direct this process. This represents an entire new class of molecular effectors which are intimately involved in differentiation. Asynchronous replication is characteristic of monoallelically expressed genes. In order to understand how each allele controls replication timing separately, they have developed a transgenic mouse model system and have already identified the DNA regions which regulate imprinted replication timing patterns. This will be used to map the molecular interactions which bring about imprinting. We have also shown that the lgk domain is subject to primary allelic exclusion in a process which employs replication timing differences to mark each allele. This may represent a new biological strategy for controlling monoallelic gene expression in a clonal manner. Ultimately they would like to understand the relationship between late replication and gene repression. New studies using injection of genes into individual nuclei of cells in culture clearly show that the ability to set up basal transcription complexes is cell cycle dependent and is most active in early S as opposed to late S. These experiments will allow them to decipher the mechanism of S phase transcriptional control and explain the relationship between replication timing and expression.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
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Cell Development and Function Integrated Review Group (CDF)
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Wolfe, Paul B
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Hebrew University of Jerusalem
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Sabag, Ofra; Zamir, Ayelet; Keshet, Ilana et al. (2014) Establishment of methylation patterns in ES cells. Nat Struct Mol Biol 21:110-2
Lande-Diner, Laura; Zhang, Jianmin; Cedar, Howard (2009) Shifts in replication timing actively affect histone acetylation during nucleosome reassembly. Mol Cell 34:767-74
Epsztejn-Litman, Silvina; Feldman, Nirit; Abu-Remaileh, Monther et al. (2008) De novo DNA methylation promoted by G9a prevents reprogramming of embryonically silenced genes. Nat Struct Mol Biol 15:1176-83