Virginia A. Zakian
Professor of Molecular Biology
Faculty AssistantMary Gidaro
- B.S., Biology, Cornell University
- Ph.D., Biology, Yale University
Research AreaGenetics & Genomics
Research FocusDNA replication and chromosome structure in yeast; telomeres; replication fork progression
DNA replication and chromosome structure in yeast; telomeres; replication fork progression
Research in the Zakian lab focuses on structures and processes that ensure the faithful maintenance of eukaryotic chromosomes, mainly using the yeast Saccharomyces cerevisiae as a model system. A major goal is to understand how telomeres, the physical ends of chromosomes, contribute to chromosome stability. Yeast chromosomes end in ~350 bps of C1-3A/TG1-3 DNA and the many proteins that associate with this DNA. Telomeres are essential for the stable maintenance of yeast chromosomes (Sandell and Zakian, 1993): they protect the end of the chromosome against degradation, help the cell distinguish intact from broken DNA, and promote replication of the very end of the chromosome. In addition, yeast telomeres are specialized sites for gene expression: genes placed near yeast telomeres are transcriptionally repressed, a phenomenon called telomere position effect (TPE) (Gottschling et al., 1990). In most eukaryotes, telomeric DNA is synthesized by a telomere specific reverse transcriptase called telomerase, but recombination provides an alternative mechanism for lengthening telomeric tracts in both yeast (Teng et al., 2000) and mammals.
Our lab uses a combination of genetic, biochemical, and cell biological approaches to identify proteins that affect telomeres and to determine their mechanism of action. For example, using a genetic strategy, we identified the Pif1p DNA helicase as an inhibitor of telomere lengthening and especially of telomere formation (Schulz and Zakian, 1994). Mutations that eliminate the helicase activity of Pif1p in vitro fail to inhibit telomerase mediated telomere lengthening in vivo (Zhou et al., 2000). Thus, Pif1p is a catalytic inhibitor of telomerase. Because Pif1p is telomere associated in vivo, it likely affects telomere replication directly. We also study other proteins that inhibit (for example, the Rif proteins; Teng et al., 2000) or promote (eg., Cdc13p; Qi and Zakian, 2000) the telomerase pathway.
Using a combination of database searches and gene isolation strategies, we found that Pif1p is the prototype member of a helicase subfamily, conserved from yeast to humans (Zhou et al., 2000). Saccharomyces has a second Pif1-like protein, called Rrm3p. Although Rrm3p also affects replication of telomeric DNA, it has an additional and very important role in replication of ribosomal DNA (rDNA). By virtue of its helicase activity, Rrm3p promotes fork progression throughout the rDNA and is especially important to resolve the converging replication forks that accumulate at the end of rDNA replication (Ivessa et al., 2000). Again, because Rrm3p is rDNA associated in vivo, it probably affects replication directly. Rrm3p has the appropriate properties to be the replicative helicase for rDNA.
The lab also uses yeast as a model system to study trinucleotide repeats. Expansion of trinucleotide repeats is the causative mutation in ~20 human genetic diseases. In many of these diseases, the expanded repeat array is a preferred site of chromosome breakage, a so-called "fragile site." Remarkably, both CTG (Freudenreich et al., 1998) and CGG (Balakumaran et al., 2000) tracts are length-dependent fragile sites in yeast, a behavior that provides the basis for genetic assays to identify genes affecting repeat expansion.
PIF1 family DNA helicases suppress R-loop mediated genome instability at tRNA genes. Nat Commun. 2017 ;8:15025. .
Pfh1 Is an Accessory Replicative Helicase that Interacts with the Replisome to Facilitate Fork Progression and Preserve Genome Integrity. PLoS Genet. 2016 ;12(9):e1006238. .
Telomerase RNA stem terminus element affects template boundary element function, telomere sequence, and shelterin binding. Proc Natl Acad Sci U S A. 2015 ;112(36):11312-7. .
Proteomics of yeast telomerase identified Cdc48-Npl4-Ufd1 and Ufd4 as regulators of Est1 and telomere length. Nat Commun. 2015 ;6:8290. .
Telomere les(i/s)ons from a telomerase RNA mutant. Cell Cycle. 2015 ;14(24):3769-70. .
21st Century Genetics: Mass Spectrometry of Yeast Telomerase. Cold Spring Harb Symp Quant Biol. 2015 ;80:111-6. .
Virginia A. Zakian is the Harry C. Weiss Professor in the Life Sciences in the Department of Molecular Biology at Princeton University. She received her B.S. in Biology from Cornell University and her Ph.D. in Biology in 1975 for research carried out in the lab of Dr. Joseph G. Gall at Yale University. In 1979, after three years of post doctoral work (first at Princeton University with Dr. Arnold J. Levine in Biochemistry and then the University of Washington with Dr.Walton F. Fangman in Genetics), she started her own lab at the Fred Hutchinson Cancer Research Center in Seattle WA. She became a tenured full professor in 1987. In 1995, she joined the faculty of the Department of Molecular Biology at Princeton University where she is currently the Harry C. Wiess Professor in the Life Sciences where her research concerns chromosome structure and replication, with a focus on telomeres, the ends of chromosomes and replication fork progression through natural replication barriers. Throughout her career, she has been an advocate for women in science.
Research in the Zakian lab uses diverse genetic, biochemical, and cell biological approaches to understand chromosome behaviour. Most of their experiments are done with either budding or fission yeasts, although other model organisms, e.g., ciliated protozoa and mammalian cultured cells, are also used.
Virginia A. Zakian has made critical contributions in two areas of chromosome biology: telomeres and replication fork progression. Her work is characterized by a willingness to tackle important questions, use of a wide variety of experimental approaches, and novel findings. Her lab used ciliates to isolate the first telomere single-‐strand DNA binding proteins, the prototype of Pot1, and demonstrated that they protect DNA ends from degradation. Her lab discovered telomeric silencing and cell cycle dependent degradation of C-‐strand telomeric DNA in budding yeast, features now known to occur in diverse eukaryotes. They also identified proteins and events required for cell cycle and length-‐dependent regulation of telomerase.
Her work on fork progression focuses on the Pif1 family of DNA helicases, which her lab showed are conserved from bacteria to humans. These studies began with the discovery that the budding yeast Pif1 acts catalytically to eject telomerase from telomeres and double strand breaks, thereby inhibiting telomerase action. Two other members of the Pif1 helicase family, budding yeast Rrm3 and fission yeast Pfh1, promote semi-‐conservative replication through telomeric DNA. Moreover, budding and fission yeast Pif1 family helicases have more general roles in combatting the replication stress that arises at naturally occurring replication barriers, such as stable protein complexes, converged replication forks, and DNA secondary structures; e.g., the Zakian lab used a combination of biochemistry and genetics to demonstrate an evolutionarily conserved role for Pif1 family helicases in promoting replication and suppressing DNA damage at G-‐quadruplex DNA. Her lab also discovered that highly transcribed RNA polymerase II genes are the most potent obstacles for DNA replication in wild type yeast cells, a conclusion later found to apply to diverse organisms.
- Featured speaker, EMBO conference Telomeres, Telomerase and Disease, Brussels, Belgium
- Keynote speaker, GRS for Gordon Res Conf., Chromosome Dynamics
- Keynote speaker, EMBO workshop, Telomere chromatin and telomere fragility,, Singapore
- Keynote speaker, Annual meeting, Swedish Society Biochemistry, Biophysics and Molecular Biology, Marstrand, Sweden
- Speaker Annual NIA IRP Postbac Day, National Institutes of Aging
- Diamonds are Forever: Celebrating First 75 Years, Princeton Adult School
- Leading Edge Lecture, City of Hope
- Keynote speaker, Anat Krauskopf Symposium, Tel Aviv IS,
- Barnum Museum Lecture, Tufts University
- Magni Lecture, Milan, Italy
- President's Lecture Series, Princeton University
- Barnum Museum Lecture, Tufts University
- President’s Lecture series, Princeton University
- Keynote Speaker, College of NJ, Advancement program Symposium
- Danny Kaye Lecture, St. Jude’s, Memphis
- Wall of Fame, Upper Darby Sr HS, Upper Darby PA
- Second Annual Athena Lecture, Royal Society, London
- Honors Program Lecture, NYU School of Medicine
- Distinguished Lecture, Lawrence Berkeley National Laboratory, Life Sciences Division
- Merit Award, NIGMS of the National Institutes of Health
- Elkin Distinguished Lectureship, Winship Cancer Center, Emory University
- Harold Varmus’ NIH Wednesday Afternoon Lectureship, National Institutes of Health
- Blaffer Seminar, University of Texas M.D. Anderson Cancer Center
- Distinguished Lecture Series, NIEHS
- June Wood Lecture, Indiana University
- Women in Cell Biology, Senior Woman Award, American Society of Cell Biology
- Fellow, American Academy of Microbiology
- Travel Fellowship, Ministry of Education, Japan
- Fellow, American Academy for the Advancement of Science