Fire Lab Research Program

The genetic landscape faced by a living cell is constantly changing. Developmental transitions, environmental shifts, and pathogenic invasions lend a dynamic character to both the genome and its activity pattern. We study a variety of natural mechanisms that are utilized by cells adapting to genetic change. These include mechanisms activated during normal development and systems for detecting and responding to foreign or unwanted genetic activity. At the root of these studies are questions of how a cell can distinguish "self" versus "nonself" and "wanted" versus "unwanted" gene expression.

Silencing Responses to Foreign DNA and RNA:

We can mimic certain aspects of genetic response to viral pathogens by injecting foreign DNA or RNA into cells. Using the nematode C. elegans as a model system, we find that injected DNA or RNA is often poorly expressed, even when carried to the proper cellular compartment. More surprisingly, experiments in which the introduced DNA or RNA is homologous to a cellular gene often result in the silencing of the endogenous gene, even in cases where that gene provides an essential physiological function for the organism.

Double stranded RNA as a major mediator in gene silencing:

In investigating the mechanisms by which injected genetic information is silenced in C. elegans, we have found a number of structural features that provide an indication of unwanted nucleic acid. The most prominent of these features has been double stranded RNA (dsRNA). Absent during "normal" gene expression, dsRNA is an essential component in the life cycle of most viruses. By flagging dsRNA as an indicator of unwanted RNA replication, and by scrupulously avoiding the production of dsRNA during most normal gene expression, the cell acquires a modicum of protection from viral infection. The mechanism by which dsRNA segments are utilized to trigger silencing of homologous genes, termed RNAi, has been a major focus in the lab for the last six years, with our efforts primarily directed toward understanding the efficacy of a system that can use just a few molecules of dsRNA to silence a large population of target molecules.

Other mediators of gene silencing:

Double stranded RNA is not the only trigger of genetic silencing for C. elegans. In addition, several aspects of DNA context (copy number, position, and tertiary structure) and RNA structure (as yet unknown) allow the system to recognize foreign invaders in the genetic makeup of the cell. The precise character of these determinants and the mechanisms underlying their ability to evoke a response are currently under investigation.

Gene silencing processes in development and pathogenesis:

We continue to investigate the roles for gene silencing components in normal development. C. elegans is an opportune system for these analyses, as development is readily followed and analyzed on a number of levels. Many of the components that have key roles in gene silencing also have critical developmental functions in C. elegans. Particularly interesting are a number of connections between silencing processes and germline development. Of all tissues, the germline goes to greatest lengths to prevent expression of unwanted genes. In particular we observe strikingly efficient silencing for factors whose normal role is to restrict cell fates in somatic tissue. Mechanisms for recognizing and silencing of such genes are essential to maintain a totipotent germline and are being analyzed by a combination of genetic and biochemical approaches.

Silencing responses to unusual DNA and RNA structures are by no means specific to C. elegans. Although mechanistic details may be different in different organisms, the underlying responses to several "aberrant" DNA and RNA triggers appear to be present in plants and fungi in addition to a broad spectrum of animal species. Silencing responses in plants have clearly been shown to reflect and respond to pathogenic challenges. In animal systems, the connections have yet to be fully worked out. In particular, we are interested in the roles of gene silencing processes in viral pathogenesis and tumor progression in mammalian systems.

Some Recent Publications

Fire, A., Xu, S., Montgomery, M.K., Kostas, S.A., Driver, S.E., and Mello, C.C. (1998) Potent and specific genetic interference by double stranded RNA in C. elegans, Nature 391, 806-811

Kelly, W., and Fire, A. (1998) Chromatin silencing and the maintenance of a functional germline in Caenorhabditis elegans, Development 125:2451-2456.

Hsieh, J., Liu, J., Kostas, S., Chang, C., Sternberg, P., and Fire, A. (1999) The RING finger/B-Box factor TAM-1 and a retinoblastoma-like protein LIN-35 modulate context-dependent gene silencing in Caenorhabditis elegans. Genes and Development, 13:2958-2970

Parrish, S., Fleenor, J., Xu, S., Mello, C., and Fire, A.(2000). Functional anatomy of a dsRNA trigger: differential requirement for the two trigger strands in RNA interference. Molecular Cell 6, 1077-1087.

Grishok, A., Pasquinelli, A., Conte, D., Li, N., Parrish, S., Ha, I., Baillie, D., Fire, A., Ruvkun, G, and Mello, C (2001) Genes and mechanisms related to RNA interference regulate expression of the small temporal RNAs that control C. elegans developmental timing. Cell. 106:23-34

Caplen, N., Parrish, S., Imani, F., Fire, A., and Morgan, R. (2001) Specific inhibition of gene expression by small double-stranded RNAs in invertebrate and vertebrate systems. Proc Natl Acad Sci U S A. 98:9742-9747.

Sijen, T., Fleenor, J., Simmer, F., Thijssen, K, Parrish, S., Timmons, L., Plasterk, R., and Fire, A. (2001) On the role of RNA amplification in dsRNA-triggered gene silencing. Cell 107:465-476.

Kostas, S., and Fire, A. (2002) The TBox factor MLS-1 acts as a molecular switch distinguishing two non-striated muscle cell types in Caenorhabditis elegans. Genes and Development 16:257-259.

Timmons, L., Tabara, H., Mello, C., and Fire, A. (2003) Inducibility of systemic RNA silencing in Caenorhabditis elegans Mol. Biol. Cell, in press

Or Try: Pubmed Searches: Our Publications, E. elegans, RNAi, Carnegie, Stanford

Lab Members (as of July 2003)

P.I.:

Andrew Fire

Technician:

Jamie Fleenor

Postdoctoral Fellow:

Judith Yanowitz

Graduate Students:

Rosa Alcazar

Ky Sha

Daniel Blanchard

Fred Tan (Joint Student with Blake Hill's Lab, JHU)

Undergraduate Student:

Amy Goh

Lab Alumni

PREVIOUS PH.D. STUDENTS (and current positions)

Verena Plunger (University of Vienna) Ph.D. 1993. Current Position: Research Associate, Department of Botany, University of Vienna. Advisor: Dr. Josef Loidl

Lihsia Chen (Johns Hopkins University) Ph.D. 1995. Current Position: Assistant Professor, Department of Cell Biology, University of Minnesota

Brian Harfe (Johns Hopkins University) Ph.D. 1997. Current Position: Assistant Professor of Molecular Genetics and Microbiology, University of Florida)

Mitch Kostich (Joint advisee with D. Fambrough, JHU) Ph.D. 1998. Current Position: Department of Bioinformatics, Schering-Plough, Madison, NJ

Jenny Hsieh (Johns Hopkins University) Ph.D. 2000. Current Position: Postdoctoral Fellow, Salk Institute, La Jolla, Ca. Advisor: Dr. Fred Gage.

Stephen Kostas (Johns Hopkins University) Ph.D. 2001. Current: University of Chicago School of Law.

Susan Parrish (Johns Hopkins University) Ph.D. 2002. Current: Postdoctoral Fellow, NIH, Bethesda, MD. Advisor: Bernard Moss.

PREVIOUS POSTDOCTORAL FELLOWS (and current positions)

Dennis Dixon (Department of Education, Province of British Columbia)

Peter Okkema (Associate Professor, Biological Sciences, University of Illinois)

Geraldine Seydoux (Associate Professor, Johns Hopkins University School of Medicine)

Joohong Ahnn (Associate Professor, Kwangju Institute of Science and Technology)

Mary Montgomery (Assistant Professor, Department of Biology, Macalester College)

William Kelly (Assistant Professor, Department of Biology, Emory Univerisity)

Jun Liu (Assistant Professor, Department of Biology, Cornell Univerisity)

Lisa Timmons (Assistant Professor, Department of Biology, University of Kansas)

PREVIOUS TECHNICIANS (and current positions)

Susan White Harrison (Postdoctoral Fellow, University of Kentucky)

SiQun Xu (Shanghai, China / St. Louis, Mo.)

Mei Hsu (Graduate Student, Dartmouth University)

PREVIOUS UNDERGRADUATE STUDENTS (and most recent info)

Beth Johnston (<need update>)

Arash Aryana (Medical Resident, Creighton University)

Ilil Carmi (Editor, Cell Press)

Andrew Godbey (MPH Student, Tulane University School of Public Health)

Staci Getz (<need update>)

Meagan Jacoby (MD/PhD Graduate, Washington University)

Caroline Macarah (Graduate Student, Loyola Marymount University)

Luiz Pantalena-Filho (Graduate Student, Stanford University)

Javier Lopez-Molina (Technician, Johns Hopkins University School of Medicine)

PREVIOUS SABBATICAL VISITORS

Kirsten Crossgrove (Loyola College of Maryland, Baltimore, MD)

Casonya Johnson (Morgan State University, Baltimore, MD)

Departmental Affiliations

Department of Embryology, Carnegie Institution of Washington

(our location until 11/03)

http://www.ciwemb.edu/

... Late Breaking News ...

We will be moving as of November 2003

from Carnegie Institution in Baltimore

to Stanford University in California

New Address after 11/03

Departments of Pathology and Genetics, Stanford University

http://pathology.stanford.edu/

http://genetics.stanford.edu/

Institutional Affiliations and Support

Carnegie Institution of Washington

http://www.ciw.edu/

Johns Hopkins University

http://www.bio.jhu.edu/

U.S. National Institutes of Health

http://www.nih.gov/

Stanford University (after 11/03)

http://www.stanford.edu/

Web Links

Our Resources

Fire Lab Vector Kits

http://www.ciwemb.edu/pages/firelab.html

Fire Lab RNAi protocols

http://www.ciwemb.edu/pages/firelab.html

Some customized Pubmed Searches

Pubmed search for recent papers from the Fire lab

http://www.ncbi.nlm.nih.gov:80/entrez/query.fcgi?cmd=search&term=fire+a[au]

Pubmed search for recent papers utilizing C. elegans

http://www.ncbi.nlm.nih.gov:80/entrez/query.fcgi?cmd=search&term=(C.+elegans)+OR+(Caenorhabditis+elegans)

Pubmed search for recent papers involving dsRNA or RNAi

http://www.ncbi.nlm.nih.gov:80/entrez/query.fcgi?cmd=search&term=(RNAi)+OR+(dsRNA)+OR+(double+stranded+RNA)+OR+(RNA+interference)+OR+(RNA-mediated+interference)+OR+(co-suppression)+OR+(cosuppression)+OR+(post-transcri

Pubmed search for recent papers from Stanford University

http://www.ncbi.nlm.nih.gov:80/entrez/query.fcgi?cmd=search&term=94305+[AD]

Pubmed search for recent papers from Carnegie Embryology

http://www.ncbi.nlm.nih.gov:80/entrez/query.fcgi?cmd=search&term=21210+[AD]

Other Resources

Natioal Center for Biotechnology Information

http://ncbi.nih.gov/

C. elegans information

http://elegans.swmed.edu/

http://www.wormbase.org/

C. elegans stock center

http://biosci.umn.edu/CGC/CGChomepage.htm

Applying to join the lab

As we make the big move from Maryland to California, we welcome new applicants to the lab. Prospective postdoctoral applicants should send a resume and summary of research to Dr. Fire at the above address, and arrange to have 3-4 letters of reference likewise sent to this address in electronic or paper format. Prospective graduate students are encouraged to apply to the Stanford Genetics Ph.D. program http://genome-www.stanford.edu/genetics/

or to any of the biosciences Ph.D. programs which can be browsed at the bioscience website http://www.med.stanford.edu/school/biosciences/ .