ZFIN ID: ZDB-LAB-080806-1
Wolfe Laboratory
PI/Director: Wolfe, Scot A.
Contact Person: Wolfe, Scot A.
Email: scot.wolfe@umassmed.edu
Address: University of Massachusetts Medical School 6th Floor, Lazare Research Building 364 Plantation St. Worcester, MA 01605-2324
Country: United States
Phone: (508) 856 3953
Fax: (508) 856 5460
Line Designation: None assigned


My research program is focused on three inter-related areas:

1) Understanding fundamental aspects of protein-DNA recognition
2) Engineering artificial transcription factors for targeted gene regulation and modification
3) Developing selection technologies to characterize and engineer protein-DNA interactions

Protein-DNA recognition - Our research on protein-DNA recognition is focused primarily on two of the most abundant families of DNA-binding domains in metazoans:

Cys2His2 Zinc fingers & Homeodomains

We have recently performed the first comprehensive analysis of homeodomain specificities in a metazoan (D. melanogaster – fruit fly) in collaboration with Michael Brodsky (UMMS-PGFE) & Gary Stormo (Wash. U) (Noyes et al., Cell 2008). Using this information we can build simple qualitative models of recognition that allow the design of homeodomains with novel DNA-binding specificity. This dataset can also be used to broadly predict the specificity of family members from other species (see ural.wustl.edu/flyhd). We continue to build upon this work to understand fundamental aspects of DNA-recognition for the homeodomain and zinc finger families with the goal of broadly and accurately predicting the specificity of naturally-occurring family members in all species. These studies will also provide a valuable resource for understanding specificity determinants within each family for rationally engineering the specificity of these DNA-binding domains.

B1H selection systems - We continue to develop a bacterial one-hybrid system for rapidly characterizing the DNA-binding specificities of sequence-specific transcription factors, both naturally-occurring and engineered. Using this technology we intend to characterize all of the sequence-specific transcription factors in the D. melanogaster genome in collaboration with the laboratory of Michael Brodsky (UMMS – PGFE). This dataset will be used to unravel transcription factor regulatory networks within the fly in collaboration with Saurabh Sinha (UI-Urbana Champaign). We have already begun building computational tools to allow the scientific community to identify cis-regulatory modules using clusters of phylogenetically conserved binding sites for the ~15% of the TFs in the fly genome that we have characterized to date (GenomeSurveyor - biotools.umassmed.edu/genomesurveyor).

ZFNs in Zebrafish - We have utilized our selection technology to create zinc finger nucleases that recognize specific genes in the Zebrafish genome in collaboration with Nathan Lawson (UMMS – PGFE). Zinc finger nucleases (ZFNs) are tailor-made restriction endonucleases that can generate a double-stranded break at a specific DNA sequence defined by the specificity of the attached zinc fingers. Using this technology we have made the first targeted gene knockouts in the zebrafish (Meng et al. Nat. Biotech 2008). We continue to develop these DNA-targeting and cleavage tools with the goal of creating an accessible resource for model organism communities that will allow them to disrupt, or modify, a desired gene in any model organism. This technology should revolutionize reverse genetic approaches in most model organisms and may allow the straightforward creation of tailor-made human disease models with profound implications for the development of treatments for a variety of diseases.

McNulty, Joseph Post-Doc Chu, Stephanie Graduate Student Gupta, Ankit Graduate Student
Noyes, Marcus Graduate Student

Shin, M., Nozaki, T., Idrizi, F., Isogai, S., Ogasawara, K., Ishida, K., Yuge, S., Roscoe, B., Wolfe, S.A., Fukuhara, S., Mochizuki, N., Deguchi, T., Lawson, N.D. (2019) Valves Are a Conserved Feature of the Zebrafish Lymphatic System. Developmental Cell. 51(3):374-386.e5
Liu, P., Luk, K., Shin, M., Idrizi, F., Kwok, S., Roscoe, B., Mintzer, E., Suresh, S., Morrison, K., Frazão, J.B., Bolukbasi, M.F., Ponnienselvan, K., Luban, J., Zhu, L.J., Lawson, N.D., Wolfe, S.A. (2019) Enhanced Cas12a editing in mammalian cells and zebrafish. Nucleic acids research. 47(8):4169-4180
Kok, F.O., Shin, M., Ni, C., Gupta, A., Grosse, A.S., van Impel, A., Kirchmaier, B.C., Peterson-Maduro, J., Kourkoulis, G., Male, I., DeSantis, D.F., Sheppard-Tindell, S., Ebarasi, L., Betsholtz, C., Schulte-Merker, S., Wolfe, S.A., Lawson, N.D. (2015) Reverse Genetic Screening Reveals Poor Correlation between Morpholino-Induced and Mutant Phenotypes in Zebrafish. Developmental Cell. 32(1):97-108
Weicksel, S.E., Gupta, A., Zannino, D.A., Wolfe, S.A., Sagerström, C.G. (2014) Targeted germ line disruptions reveal general and species-specific roles for paralog group 1 hox genes in zebrafish. BMC Developmental Biology. 14:25
Kok, F.O., Gupta, A., Lawson, N.D., and Wolfe, S.A. (2014) Construction and application of site-specific artificial nucleases for targeted gene editing. Methods in molecular biology (Clifton, N.J.). 1101:267-303
Gupta, A., Hall, V.L., Kok, F.O., Shin, M., McNulty, J.C., Lawson, N.D., and Wolfe, S.A. (2013) Targeted Chromosomal Deletions and Inversions in Zebrafish. Genome research. 23(6):1008-17
Shin, J., Padmanabhan, A., de Groh, E.D., Lee, J.S., Haidar, S., Dahlberg, S., Guo, F., He, S., Wolman, M.A., Granato, M., Lawson, N.D., Wolfe, S.A., Kim, S.H., Solnica-Krezel, L., Kanki, J.P., Ligon, K.L., Epstein, J.A., and Look, A.T. (2012) Zebrafish neurofibromatosis type 1 genes have redundant functions in tumorigenesis and embryonic development. Disease models & mechanisms. 5(6):881-894
Gupta, A., Christensen, R.G., Rayla, A.L., Lakshmanan, A., Stormo, G.D., and Wolfe, S.A. (2012) An optimized two-finger archive for ZFN-mediated gene targeting. Nature Methods. 9(6):588-590
Zhu, C., Smith, T., McNulty, J., Rayla, A.L., Lakshmanan, A., Siekmann, A.F., Buffardi, M., Meng, X., Shin, J., Padmanabhan, A., Cifuentes, D., Giraldez, A.J., Look, A.T., Epstein, J.A., Lawson, N.D., and Wolfe, S.A. (2011) Evaluation and application of modularly assembled zinc-finger nucleases in zebrafish. Development (Cambridge, England). 138(20):4555-4564
Lawson, N.D., and Wolfe, S.A. (2011) Forward and reverse genetic approaches for the analysis of vertebrate development in the zebrafish. Developmental Cell. 21(1):48-64
Bussmann, J., Wolfe, S.A., and Siekmann, A.F. (2011) Arterial-venous network formation during brain vascularization involves hemodynamic regulation of chemokine signaling. Development (Cambridge, England). 138(9):1717-1726
Gupta, A., Meng, X., Zhu, L.J., Lawson, N.D., and Wolfe, S.A. (2011) Zinc finger protein-dependent and -independent contributions to the in vivo off-target activity of zinc finger nucleases. Nucleic acids research. 39(1):381-392
Cifuentes, D., Xue, H., Taylor, D.W., Patnode, H., Mishima, Y., Cheloufim, S., Ma, E., Mane, S., Hannon, G.J., Lawson, N.D., Wolfe, S.A., and Giraldez, A.J. (2010) A novel miRNA processing pathway independent of Dicer requires Argonaute2 catalytic activity. Science (New York, N.Y.). 328(5986):1694-1698
Siekmann, A.F., Standley, C., Fogarty, K.E., Wolfe, S.A., and Lawson, N.D. (2009) Chemokine signaling guides regional patterning of the first embryonic artery. Genes & Development. 23(19):2272-2277
Meng, X., Noyes, M.B., Zhu, L.J., Lawson, N.D., and Wolfe, S.A. (2008) Targeted gene inactivation in zebrafish using engineered zinc-finger nucleases. Nat. Biotechnol.. 26(6):695-701