Anthony Schwacha

  • Associate Professor
  • DNA replication

Contact

Office: (412) 624-4307
Lab: (412) 624-4307
560B Crawford Hall
4249 Fifth Avenue
Pittsburgh, PA 15260

Cancer is caused by defects in the normal genetic code.  Emerging evidence suggests that regulatory mishaps during DNA replication are often the causative event for this dreadful disease.  To address the fundamental relationship between DNA replication and cancer, my laboratory studies the major regulatory hub of DNA replication – the Mcm2-7 replicative helicase.  As a helicase, Mcm2-7 uses ATP binding and hydrolysis to unwind double-stranded DNA to generate templates for DNA polymerase.  From a regulatory standpoint, the loading, activation, and subsequent DNA unwinding by this complex are all heavily regulated to ensure the precise production of a single exact copy of the genome during each S-phase.  Alterations in this regulation cause damage that has been shown experimentally to lead to cancer.  In combination with various collaborators, my lab uses a multitude of biochemical, genetic, structural, cytological and genomic approaches to address fundamental biological issues using the budding yeast S. cerevisiae, an easy-to-work-with model eukaryote whose DNA replication process is conserved with those of humans. The fusion of complementary in vitro and in vivo methodologies in my lab provides excellent and unique research training opportunities.

 Figure 1. The Mcm gate model specifics an inverse functional relationship between activity and closure of the discontinuity.

The key to Mcm2-7 regulation is its unusual subunit composition. Unlike related helicases formed from six identical subunits, Mcm2-7 is formed from six unique and functionally distinct subunits.  Three Mcm subunits are specialized to unwind DNA (Mcm4, 6, and 7) while the other three (Mcm2, 3, and 5) regulate the activity of the complex.  How the regulatory subunits control Mcm2-7 activity is a critical unsolved question in the replication field.  To address this problem, we have found that the regulatory subunits form a reversible discontinuity within the Mcm2-7 ring structure (Figure 1).  Biochemical activity is inversely related to the discontinuity – closing the ring activates DNA unwinding;  while opening the ring inactivates this activity.  These observations suggest that Mcm2-7 regulation in living cells might occur through cellular factors that regulate the state of the discontinuity.  Recent structural data from our colleagues provide strong support for our hypothesis (Costa et. al. 2011 Nat. Struct. Biol. 18:471).

Figure 2.  Comparison of ciprofloxacin inhibition on yeast growth and helicase activity (Mcm2-7, Mcm467 (a Mcm subcomplex that has helicase activity and lacks the regulatory subunits), and a related helicase (SV-40 large T antigen).

Although this model provides a rational mechanistic explanation for Mcm2-7 regulation, its relevance in living cells remains to be tested.  Current studies in the Schwacha Lab largely focus on this issue by comparing various aspects of Mcm2-7 regulation within living cells with their biochemical effect on the Mcm2-7  discontinuity.  In parallel, we have started to identify small molecule inhibitors of Mcm2-7 DNA unwinding activity (Figure 2).  These compounds are ultimately hoped to provide both useful research tools to study Mcm function as well as therapeutic agents to fight cancer.

For a more complete discussion of current research in the Schwacha Lab, please visit our lab webpage

Tsai FL, Vijayraghavan S., Prinz J., MacAlpine

Tsai FL, Vijayraghavan S., Prinz J., MacAlpine HK., MacAlpine DM., and A. Schwacha. (2015) Mcm2-7 is an Active Player in the DNA Replication Checkpoint Signaling Cascade via Proposed Modulation of its DNA Gate. Mol Cell Biol. 2015 Apr 13. pii: MCB.01357-14. [Epub ahead of print]

Simon, N. and A. Schwacha (2014) The Mcm2-7 Rep

Simon, N. and A. Schwacha (2014) The Mcm2-7 Replicative Helicase: A Promising Chemotherapuetic Target.  Biome Res Int 2014:549719

Simon, N., Bochman, M., Seguin, S., Brodsky, J.

Simon, N., Bochman, M., Seguin, S., Brodsky, J.L., Seibel, W.L., and Schwacha. A. (2013)  Ciprofloxacin is a selective inhibitor of the Mcm2-7 Replicative Helicase. Biosci Rep. 33:783-795

Vijayraghavan, S., and A. Schwacha

Vijayraghavan, S., and A. Schwacha (2012) The Eukaryotic Mcm2-7 Replicative Helicase in The Eukaryotic Replisome: A guide to protein structure and Function. Ed. S. MacNeill, Springer, New York, NY.

Bochman, M.L., and A. Schwacha (2010) The Saccharomyces cerevisiae Mcm6/2 and Mcm5/3 ATPase activ

Bochman, M.L., and A. Schwacha (2010) The Saccharomyces cerevisiae Mcm6/2 and Mcm5/3 ATPase active sites contribute to the function of the putative Mcm2-7 'gate'. Nucleic Acids Res. 38:6078-6088

Bochman, M.L., and A. Schwacha (2009) The Mcm complex: unwinding the mechanism of a replicative h

Bochman, M.L., and A. Schwacha (2009) The Mcm complex: unwinding the mechanism of a replicative helicase. Microbiol Mol Biol Rev 73:652-683

Bochman, M.L., and A. Schwacha (2008) The Mcm2-7 complex has in vitro helicase activity.

Bochman, M.L., and A. Schwacha (2008) The Mcm2-7 complex has in vitro helicase activity. Mol. Cell 31:287-293

Bochman, M.L., S.P. Bell, and A. Schwacha (2008) Subunit organization of Mcm2-7 and the unequal r

Bochman, M.L., S.P. Bell, and A. Schwacha (2008) Subunit organization of Mcm2-7 and the unequal role of active sites in ATP hydrolysis and viability. Mol. Cell Biol. 28:5865-5873

Bochman, M.L., and A. Schwacha (2007) Differences in the single-stranded DNA binding activities o

Bochman, M.L., and A. Schwacha (2007) Differences in the single-stranded DNA binding activities of MCM2-7 and MCM467: MCM2 and 5 define a slow ATP-dependent step. J. Biol. Chem. 282:33795-33804

Schwacha, A., and S.P. Bell (2001) Interactions between two catalytically distinct MCM subgroups

Schwacha, A., and S.P. Bell (2001) Interactions between two catalytically distinct MCM subgroups are essential for coordinated ATP hydrolysis and DNA replication. Mol. Cell 8:1093-1104
Dr. Schwacha received his Ph.D. in 1996 with Nancy Kleckner at Harvard University, performed his postdoctoral studies with Stephen Bell at the Massachusetts Institute of Technology, and joined the Department in 2003.