MacAlpine Research
Research Focus
One of the most important unanswered questions in the field of replication is where and how metazoan origins are specified. Replication initiates from multiple sites (origins) throughout the chromosome. These origins are defined by both cis- and trans-acting factors. In S. cerevisiae, the characterization of the cis-acting elements that define origins of replication led to the identification of the origin replication complex (ORC), an essential six-subunit complex that marks potential sites of replication initiation. Additional proteins are recruited to origins of replication, in an ORC dependent manner, forming the pre-replicative complex (pre-RC). The pre-RC regulates origin function and prevents re-replication. Despite the conservation of pre-RC components in all eukaryotic species, the sequences that define metazoan origins remain poorly characterized, with only a handful of origins being identified and characterized.
I have developed a systematic approach to identify a large number of replication origins in D. melanogaster by coupling chromatin immunoprecipitation with genomic microarrays. I designed a high density microarray which contains approximately 13 thousand unique PCR products and includes both intergenic and intragenic sequences along the entire left arm of chromosome 2 and all unique sequences on chromosome 4. The high resolution of this microarray, combined with immunoprecipitation allows for the identification of specific protein-DNA interactions at a resolution of 1.5 kb.
Using this technology, I have found evidence that there is a connection between transcription and replication at two distinct levels: a global effect on origin activation and a local effect on origin selection by ORC.
Publications
MacAlpine DM, Bell SP. A genomic view of eukaryotic DNA replication. Chromosome Res. 2005; 13(3):309-26.
Jeonn Y, Bekiranov S, Karnani N, Kapranov P, Ghosh S, MacAlpine DM, Lee C, Hwang DS, Gingeras T, Dutta A. Temporal profile of replication of human chromosomes. Proc. Natl. Acad. Sci. 2005. In Press.
MacAlpine DM, Rodriguez HK, Bell SP, Coordination of replication and transcription along a Drosophila chromosome. Genes Dev. 2004 Dec 15; 18(24). [PDF]
Schubeler D, MacAlpine DM, Scalzo D, Wirbelauer C, Kooperberg C, van Leeuwen F, Gottschling DE, O'Neil LP, Turner BM, Delrow J, Bell SP, Groudine M. The histone modification pattern of active genes revealed through genome-wide chromatin analysis of a higher eukaryote. Genes Dev. 2004 Jun 1; 18(11). [PDF]
Claycomb JM, MacAlpine DM, Evans JG, Bell SP, Orr-Weaver TL. Visualization of replication initiation and elongation in Drosophila. J Cell Biol. 2002 Oct 28;159(2):225-36. [PDF]
MacAlpine, DM. Studies of mitochondrial DNA inheritance in S. cerevisiae: the influence of recombination, DNA packaging, and replication. Thesis 2001. [PDF]
MacAlpine DM, Kolesar J, Okamoto K, Butow RA, Perlman PS. Replication and preferential inheritance of hypersuppressive petite mitochondrial DNA. EMBO J. 2001 Apr 2;20(7):1807-17. [PDF]
MacAlpine DM, Perlman PS, Butow RA. The numbers of individual mitochondrial DNA molecules and mitochondrial DNA nucleoids in yeast are co-regulated by the general amino acid control pathway. EMBO J. 2000 Feb 15;19(4):767-75. [PDF]
MacAlpine DM, Perlman PS, Butow RA. The high mobility group protein Abf2p influences the level of yeast mitochondrial DNA recombination intermediates in vivo. Proc Natl Acad Sci U S A. 1998 Jun 9;95(12):6739-43. [PDF]
Zhang Z, MacAlpine DM, Kapler GM. Developmental regulation of DNA replication: replication fork barriers and programmed gene amplification in Tetrahymena thermophila. Mol Cell Biol. 1997 Oct;17(10):6147-56. [PDF]
MacAlpine DM, Zhang Z, Kapler GM. Type I elements mediate replication fork pausing at conserved upstream sites in the Tetrahymena thermophila ribosomal DNA minichromosome. Mol Cell Biol. 1997 Aug;17(8):4517-25. [PDF]
Andrews DL, MacAlpine DM, Johnson JR, Kelley PM, Cobb BG, Drew MC. Differential induction of mRNAs for the glycolytic and ethanolic fermentative pathways by hypoxia and anoxia in maize seedlings. Plant Physiol. 1994 Dec;106(4):1575-82. [PDF]
Awards and Fellowships
Damon Runyon Cancer Research Foundation, post-doctoral fellowship, Aug. 2001 -- 2004.