Dorothy Pazin


Courtney Sill

As winner, Dorothy Pazin was awarded a prize of $250, and runner-up Courtney Sill was awarded $100.

Sullivan lab student Dorothy Pazin wins in poster competition

Spreading of centromeric chromatin in cis on Drosophila minichromosomes

Dorothy E. Pazin and Beth A. Sullivan

The centromere ensures proper segregation of chromosomes during mitosis and meiosis. Eukaryotic centromeres are typically located within regions of repetitive DNA. However, variant centromeres, called neocentromeres, are able to form at non-centromeric sites (i.e. “euchromatin”). Thus, a centromere may be any sequence that is marked by a change in chromatin structure or is epigenetically regulated. The mechanism of neocentromere formation is unclear, especially since most human neocentromeres are ascertained long after their formation and stabilization. To study neocentromere formation and determinants of centromere identity, we use a Drosophila minichromosome (Dp 8-23) that contains a fully functional centromere. The centromere (CEN) of Dp 8-23 has been defined to a 420 kb region flanked by heterochromatin (Murphy & Karpen 1995). Subsequent irradiation of flies carrying Dp 8-23 resulted in a series of derivative minichromosomes (Dp238 and DpJ21A) that contain variable amounts of CEN DNA and heterochromatin. Some derivatives completely lack the 420kb CEN but are stable, suggesting that they are neocentromeres and have ‘acquired’ centromere function. In this study we investigated if removal of heterochromatin allows CEN to spread on the euchromatic DNA of Dp238 and whether CEN must spread further on the smaller derivative DpJ21A. Our results show that positioning a centromere adjacent to euchromatin (Dp238) allows CID to spread up to 66kb into non-centromeric DNA. On smaller minichromosomes (DpJ21A), CID is spreads even farther, over 150kb. We can conclude that derivative minichromosomes (Dps) are stable because CID has spread over euchromatin, maintaining a functional kinetochore and that euchromatic sequences provide a chromatin environment that can support kinetochore assembly.











Christman lab student Courtney Sill is runner-up in poster competition

Esa1 is the only known yeast histone acetyltransferase (HAT) required for cell viability. It is a member of the MYST family of HAT proteins which includes the homologous human protein Tip60 as well as the Drosophila MOF protein. All three homologs contain a conserved acetyltransferase domain in addition to a chromodomain located near the N-terminus.

While ESA1’s HAT activity is important in processes such as DNA repair, it is likely not its essential function. Mutants with a single point mutation in the active site cysteine are still viable even though their acetyltransferase abilities are abolished [1].

And although the acetyltransferase activities of HAT proteins can positively regulate transcription via modifications to histone tails, chromatin immunoprecipitation assays have shown ESA1 is distributed evenly along the length of chromatin, not localized to specific promoters as would be expected from a HAT protein involved in transcriptional regulation [2].

But, as is the case for other HAT proteins, ESA1’s acetyltransferase activity is significant, however, in processes such as DNA replication, DNA repair and cell cycle progression [1,3]. The aim of my research is to determine the essential function of ESA1 - the catalytic subunit of the yeast HAT complex, NuA4 - which I hypothesize may be DNA replication and/or DNA repair.









References

1. Bird AW, Yu DY, Pray-Grant MG, Qiu Q, Harmon KE, Megee PC, Grant PA, Smith MM, Christman MF. Acetylation of histone H4 by Esa1 is required for DNA double-strand break repair. Nature. 2002 Sep 26;419(6905):411-5.

2. Vogelauer M, Wu J, Suka N, Grunstein M. Global histone acetylation and deacetylation in yeast. Nature. 2000 Nov 23;408(6811):495-8.

3. Clarke AS, Lowell JE, Jacobson SJ, Pillus L. Esa1p is an essential histone acetyltransferase required for cell cycle progression. Mol Cell Biol. 1999 Apr;19(4):2515-26.