Ann Skalka, PhD
Senior Advisor to the President
Office Phone: 215-728-2490
The work in our laboratory is focused on obtaining a detailed understanding of the mechanism by which retroviral DNA is integrated into its host cell chromatin, and discovering the epigenetic factors and processes that affect its subsequent expression. Retroviruses are of special interest, not only because they are agents of disease, including cancer, but also because they are important as vehicles for the insertion of desired genes into target cells for scientific investigation and gene therapy. Recently we have also become interested in whether sequences from viruses other than retroviruses have become inserted into vertebrate genomes over an evolutionary time frame, how this may have occurred and, what may be the consequences. We exploit a broad range of investigational methods in our studies from analyses of protein function, to in vivo studies of viral growth and cell biology. This comprehensive approach provides unique insights, and excellent opportunities for cross-discipline training and collaboration. Our continuing overarching goals are to uncover new information of fundamental importance to both virus and cell biology, and to identify new targets for therapies to treat disease.
We have made exciting progress in our investigation of the molecular structure of retroviral integrases, the enzymes that mediate insertion of viral DNA into the DNA of its host cell. The techniques of small angle X-ray scattering (SAXS) and molecular modeling have been used to obtain the first available information concerning the shapes and structures of integrase monomers, dimers, and tetramers in solution. The resulting models are tested using biochemical crosslinking and mass spectrometry. These structures can now be used to identify points of contact between the protein subunits, as well as between the protein and its DNA substrates. Such information will not only add to our understanding of this complex reaction, but may also suggest new ways to interfere with this activity, which is critical for viral replication.
Together with collaborators in the Systems Biology Group at the Institute for Advanced Study in Princeton, we conducted a systematic analysis of the 48 published vertebrate genomes for sequences related to all known viruses with single-stranded RNA or DNA genomes. We discovered numerous examples of sequences related to two families of RNA viruses, the Bornaviruses and Filoviruses, which entered 19 vertebrate genomes about 40 million years ago, most likely facilitated by LINE retrotransposons. We also discovered that the history of integration of sequences related to the small DNA virus families, the Parvoviruses and Circoviruses, spans an even longer evolutionary period from 60 million years ago to recent times. Some of the sequences from both types of ancient viruses have retained open reading frames, indicating that they have been subject to positive selection and must have provided some advantage to the host or the virus over time.
To understand how integrated viral gene expression is controlled, we have developed a system in which we can monitor high-frequency epigenetic gene silencing of freshly integrated retroviral DNA. We believe that such silencing may reflect a cellular anti-viral response. We have published the results from our proof-of-principle studies using silent “reporter cells” to establish the validity of this system for identification of cellular factors that govern this epigenetic silencing – a phenomenon that is important to our understanding of the antiviral response as well as, the roles of epigenetic factors in normal cell development and cancer. With the help of the Fox Chase Cancer Center High Throughput Screening Facility, we have used this reporter cell population to develop a robust and sensitive siRNA-based high throughput screen to identify specific host factors that participate in the maintenance of epigenetic silencing. Our results to date provide compelling evidence for roles of specific host factors and indicate that they are involved in cooperative and reinforcing interactions that mediate epigenetic silencing.Description of research projects
Fox Chase Programs
- Skalka, AM. HIV: Integration triggers death. Nature 2013, 498:305-06. PubMed
- Shalginskikh, N, Poleshko, A, Skalka, AM, Katz, R. Retroviral DNA methylation and epigenetic repression are mediated by the antiviral host protein Daxx. J Virol 2013;87:2137-50. Selected by the Editors as an Article of “Significant Interest.” PMCID: PMC3571491 PubMed
- Bojja, RS, Andrake, MD, Merkel, G, Weigand, S, Dunbrack, RL, Jr, Skalka, AM. Architecture and assembly of HIV integrase multimers in the absence of DNA substrates. J Biol Chem 2013;288:7373-86. PMCID: PMC3591645 PubMed
- Peletskaya, E, Andrake, M, Gustchina, A, Merkel, G, Alexandratos, J, Zhou, D, Bojja, RS, Satoh, T, Potapov, M, Kogon, A, Potapov, V, Wlodawer, A, Skalka, AM. Localization of ASV integrase-DNA contacts by site-directed crosslinking and their structural analysis. PLoS One 2011;6:e27751. PubMed
- Katz RA, Merkel G, Andrake MD, Roder H, Skalka AM. Retroviral integrases promote fraying of viral DNA ends. J Biol Chem. 2011;286:25710-8. PubMed
- Bojja RS, Andrake MD, Weigand S, Merkel G, Yarychkivska O, Henderson A, Kummerling M, Skalka AM. Architecture of a full-length retroviral integrase monomer and dimer, revealed by small angle X-ray scattering and chemical cross-linking. J Biol Chem. 2011 May 13;286(19):17047-59. Epub 2011 Mar 15. PubMed
- Belyi VA, Levine AJ, Skalka AM. Sequences from Ancestral Single Stranded DNA Viruses In Vertebrate Genomes: The Parvoviridae And Circoviridae Are More Than 40-50 Million Years Old. J Virol. 2010;84:12458-12462. PubMed
- Belyi VA, Levine AJ, Skalka AM, 2010 Unexpected Inheritance: Multiple Integrations of Ancient Bornavirus and Ebolavirus/Marburgvirus Sequences in Vertebrate Genomes. PLoS Pathog 6(7): e1001030. doi:10.1371/journal.ppat.1001030. PLoS Pathog
- Poleshko A, Einarson MB, Adams PD, Zhang R, Skalka AM, Katz RA. Identification of a functional network of human epigenetic silencing factors. J Biol Chem. 2010 Jan 1;285(1):422-33. Epub 2009 Oct 30. PubMed
- Merkel G, Andrake MD, Ramcharan J, Skalka AM. Oligonucleotide-based assays for integrase activity. Mechanistic and Pharmacological Analyses of HIV-1 Integration. Alan Engelman, ed. Elsevier Editorial Systems for Methods. 2009 Apr;47(4):243-8. Epub 2008 Nov 14. PubMed
- Katz RA, Daniel R, Skalka AM. Host factors that affect provirus stability and silencing. HIV1 Integrase: Mechanisms Of Action And Inhibitor Design. (Neamati, N, Wand, B, eds.), Chapter 11, pp 141-50. John Wiley and Sons, Inc., UK.
- Flint SJ, Enquist LW, Racaniello VR, Skalka AM. Principles of Virology, 3rd edition. Vol. II. Pathogenesis and Control of Animal Viruses, 419 pages. ASM Press, Washington DC, 2009
- Flint SJ, Enquist LW, Racaniello VR, Skalka AM. Principles of Virology, 3rd edition. Vol I. Molecular Biology, 569 pages. ASM Press, Washington DC, 2009.
- Andrake MD, Sauter MM, Boland K, Goldstein AD, Hussein M, and Skalka AM. Nuclear import of avian sarcoma virus integrase is facilitated by host cell factors. Retrovirology. 2008 August;5:73. PubMed
- Poleshko A, Palagin I, Zhang R, Boimel P, Castagna C, Adams PD, Skalka AM, and Katz RA. Identification of cellular proteins that maintain retroviral epigenetic silencing: evidence for an antiviral response. J Virol. 2008 December;82(5):2313-23. PubMed
- Katz RA, Jack-Scott E, Narezkina A, Palagin I, Boimel P, Kulkosky J, Nicolas E, Greger J.G, and Skalka AM. High frequency epigenetic repression and silencing of retroviruses can be antagonized by HDAC inhibitors and transcriptional activators, but uniform reactivation in cell clones is restricted by additional mechanisms. J Virol. 2007 January;81(6):2592-04. PubMed
- Ramcharan J, Colleluori DM, Merkel G, Andrake MD, and Skalka AM. Mode of Inhibition of HIV-1 integrase by a C-terminal domain-specific monoclonal antibody. Retrovirology. 2006 June; 3:34. PubMed
- Ramcharan J, Skalka AM. Strategies for identification of HIV-1 integrase inhibitors. Future Virol. 2006 November;1:717-31.
- Ason B, Knauss DJ, Balke AM, Merkel G, Skalka AM, Reznikoff WS. Targeting Tn5 transposase identifies HIV-1 inhibitors. Antimicrob. Agents Chemother. 2005 April;49:2035-43. PubMed
- Daniel R, Marusich E, Argyris E, Zhao R Y, Skalka AM, and Pomerantz RJ. Caffeine inhibits HIV-1 transduction of non-dividing cells. J Virol. 2005 February;79(4):2058-65. PubMed
- Greger JG, Katz RA, Ishov AM, Maul G, and Skalka AM. The cellular protein Daxx interacts with Avian Sarcoma Virus integrase and viral DNA to repress viral transcription. J Virol. 2005 March;79(8):4610-4618, 2005. PubMed
- Katz RA, Greger JG, and Skalka AM. Effects of cell cycle status on early events in retroviral replication. J Cell Biochem. 2005 April;94:880-89.
- Skalka AM, and Katz RA. Retroviral DNA integration and the DNA damage response. Cell Death Diff. 2005 August;2:971-78.
- Daniel R, Ramcharan J, Rogakou E, Taganov KD, Greger JG, Bonner W, Nussenzweig A, Katz RA, and Skalka AM Histone H2AX is phosphorylated at sites of retroviral DNA integration, but is dispensable for post-integration repair. J Biol Chem. 2004 August;279:45810-14.PubMed
- Narezkina, A., Taganov, K.D., Litwin, S., Stoyanova, R., Hayashi, J., Seeger, C., Skalka, A.M. and Katz, R.A. Genome-wide analyses of avian sarcoma virus integration sites. J Virol. 2004 October;78(21):11656-63.PubMed
- Daniel R, Katz RA, Greger J, Taganov K, Wu X, Kappes JC, and Skalka, A.M. Evidence that stable retroviral transduction and cell survival following DNA integration depends on components of the NHEJ repair pathway. J Virol. 2004 July;78(16):8573-81.PubMed
- Greger J, Katz RA, Taganov K, Rall GF, and Skalka AM. Transduction of terminally differentiated neurons by Avian Sarcoma Virus. J Virol. 2004 April;78(9):4902-06.PubMed