Progress 02/01/96 to 09/30/11
Outputs
OUTPUTS: We have identified the basis of specificity in certain mRNA-protein interactions and are now developing methods to manipulate proteins to obtain new, designer proteins with novel specificities. We have determined how PUF proteins promote deadenylation: they interact directly with a multifunctional deadenylase complex. We have identified new classes of poly(A) and poly(U) adding enzymes. The GlD2 and PUF proteins control stem cells in diverse systems, including, as we have shown, in the germ lines of C. Elegans and Drosophila. We have begun studying the poly(A) adding enzyme of these proteins in the nervous system, and already show that Gld2 is required for long term memory. PARTICIPANTS: Marv Wickens oversaw all aspects of the work. A. Cooke worked on regulation of GLD2 by Bic-C protein, and isolation of GLD-2 complexes. J. Chritton worked on development of in vitro assays for mRNA control. C. Stumpf worked on RNA binding specificity of PUF proteins in yeast and Xenopus and their interactions with protein partners. Yvonne Koh worked on the nature of PUF protein specificity. Dan Wilinski studies the evolution of RNA regulatory circuitry. Zak Campbell studies the dyanmics of mRNA control. N. Buter was the research specialist who performed minipreps, transformations of E. coli and yeast, and she helped with 3 hybrid screens and analysis. TARGET AUDIENCES: Nothing significant to report during this reporting period. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
The work has led to two patents. The objects are to manipulate the fate and function of mRNAs. In addition, both the RNA-binding proteins and the poly(A) adding enzymes may have critical roles in stem cells.
Publications
- Chritton, J.J. and Wickens, M. (2010) Translational repression by PUF proteins in vitro. RNA. 16, 1217-1225.
- Cooke, A, Prigg, A., and Wickens, M. (2010) Translational repression by deadenylases. J. Biol. Chem. 16, 28506-28513.
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Progress 01/01/09 to 12/31/09
Outputs
OUTPUTS: We have identified the basis of specificity in certain mRNA-protein interactions and are now developing methods to manipulate proteins to obtain new, designer proteins with novel specificities. We have determined how PUF proteins promote deadenylation: they interact directly with a multifunctional deadenylase complex. We have identified new classes of poly(A) and poly(U) adding enzymes. The GlD2 and PUF proteins control stem cells in diverse systems, including, as we have shown, in the germ lines of C. Elegans and Drosophila. We have begun studying the poly(A) adding enzyme of these proteins in the nervous system, and already show that Gld2 is required for long term memory. PARTICIPANTS: Marv Wickens oversaw all aspects of the work. A. Cooke worked on regulation of GLD2 by Bic-C protein, and isolation of GLD-2 complexes. J. Chritton worked on development of in vitro assays for mRNA control. C. Stumpf worked on RNA binding specificity of PUF proteins in yeast and Xenopus and their interactions with protein partners. Yvonne Koh works on the nature of PUF protein specificity. Dan Wilinski studies the evolution of RNA regulatory circuitry. Zak Campbell studies the dyanmics of mRNA control. N. Buter was the research specialist who performed minipreps, transformations of E. coli and yesat, and she helped with 3 hybrid screens and analysis. TARGET AUDIENCES: Nothing significant to report during this reporting period. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
The work has led to two patents. The objects are to manipulate the fate and function of mRNAs. In addition, both the RNA-binding proteins and the poly(A) adding enzymes may have critical roles in stem cells.
Publications
- Suh N, Crittenden SL, Goldstrohm A, Hook B, Thompson B, Wickens M, Kimble J. (2009) FBF and Its Dual Control of gld-1 Expression in the Caenorhabditis elegans Germline. Genetics. 181, 1249-1260.
- Koh, Y, Opperman, L. Stumpf, C., Mandan, A., Keles, S., and Wickens, M. (2009) A singe C. elegans PUF protein binds RNA in multiple modes. RNA. 15, 1090-1099.
- Zhu D, Stumpf CR, Krahn JM, Wickens M, Hall TM. (2009) A 5 prime cytosine binding pocket in Puf3p specifies regulation of mitochondrial mRNAs. Proc Natl Acad Sci U S A. In Press.
- Wang Y, Opperman L, Wickens M, Hall TM. (2009) Structural basis for specific recognition of multiple mRNA targets by a PUF regulatory protein. Proc Natl Acad Sci U S A.
- Goldstrohm, A.C., B.A. Hook, and M. Wickens. (2008). Regulated deadenylation in vitro. In RNA Turnover in Eukaryotes: Nucleases, Pathways and Analysis of mRNA Decay of the Methods in Enzymology Series (L.E. Maquat and M. Kiledjian eds.), Elsevier Inc. Vol. 448, Chapter 5, 77-106
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Progress 01/01/08 to 12/31/08
Outputs
OUTPUTS: We have identified the basis of specificity in certain mRNA-protein interactions and are now developing methods to manipulate proteins to obtain new, designer proteins with novel specificities. We have determined how PUF proteins promote deadenylation: they interact directly with a multifunctional deadenylase complex. We have identified new classes of poly(A) and poly(U) adding enzymes. The GlD2 and PUF proteins control stem cells in diverse systems, including, as we have shown, in the germ lines of C. Elegans and Drosophila. We have begun studying the poly(A) adding enzyme of these proteins in the nervous system, and already show that Gld2 is required for long term memory. TARGET AUDIENCES: Research Community PARTICIPANTS: Marv Wickens oversaw all aspects of the work. A. Cooke worked on regulation of GLD2 by Bic-C protein, and isolation of GLD-2 complexes. Jae Eun Kwak worked on mechanisms of GLD2 action and its role in memory. J. Chritton worked on development of in vitro assays for mRNA conrtrol. L. Opperman worked on GLd2 as an activator of translation and stability and also worked on PUF specificity, regulation in frog oocytes, and the targeting of designer PUF proteins to new mRNAs. A. Goldstrohm was a key researcher on the project. C. Stumpf worked on RNA binding specificity of PUF proteins in yeast and Xenopus and their interactions with protein partners. N. Buter was the research specialist who performed minipreps, transformations of E. coli and yesat, and she helped with 3 hybrid screens and analysis. TARGET AUDIENCES: Research Community PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
The work has led to two patents. The objects are to manipulate the fate and function of mRNAs. In addition, both the RNA-binding proteins and the poly(A) adding enzymes may have critical roles in stem cells.
Publications
- 1. Benoit, P., Papin, C., Kwak, J.E., Wickens, M., and Simonelig, M. (2008). PAP- and GLD-2-type poly(A) polymerases are required sequentially in cytoplasmic polyadenylation and oogenesis in Drosophila. Development. 135, 1969-1979.
- 2. Kwak, J.E., Drier, E., Barbee, S.A., Ramaswami, M., Yin, J.C.P., and M. Wickens. (2008) GLD2 poly(A) polymerase is specifically required for long-term memory. Proc Natl Acad Sci U.S.A. 105, 14644-16649.
- 3. Stumpf, C.R., J. Kimble, and M. Wickens. (2008) A C. elegans PUF protein family with distinct RNA binding specificity. RNA. 14, 1550-1557.
- 4. Goldstrohm, A.C. and M. Wickens (2008). Multifunctional deadenylase complexes diversify mRNA control. Nature Reviews Molecular Cell Biology. 9, 337-344.
- 5. C. Stumpf. L. Opperman and M. Wickens, (2008). The three-hybrid system to analyze RNA protein interactions. Methods in Enzymology, In Press.
- 6. Wickens, M. and J.E. Kwak. (2008). Molecular Biology. A Tail Tale for U. Science. 19, 1344-1345.
- 7. Goldstrohm, A. and M. Wickens. (2008). Cell-free systems to study RNA decay. Methods in Enzymlogy. In Press.
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Progress 01/01/07 to 12/31/07
Outputs
OUTPUTS: We have identified the basis of specificity in certain mRNA-protein interactions and are now developing methods to manipulate proteins to obtain new, designer proteins with novel specificities. We have determined how PUF proteins promote deadenylation: they interact directly with a multifunctional deadenylase complex. We have identified new classes of poly(A) and poly(U) adding enzymes. The GlD2 and PUF proteins control stem cells in diverse systems, including, as we have shown, in the germ lines of C. Elegans and Drosophila. We have begun studying the poly(A) adding enzyme of these proteins in the nervous system, and already show that Gld2 is required for long term memory
TARGET AUDIENCES: Research Community
Impacts
The work has led to two patents. The objects are to manipulate the fate and function of mRNAs. In addition, both the RNA-binding proteins and the poly(A) adding enzymes may have critical roles in stem cells.
Publications
- Rouhana, L. and M. Wickens (2007), Autoregulation of GLD-2 cytoplasmic poly(A) polymerase. RNA, 13, 188-199.
- Goldstrohm, A.C., D..J. Seay, B.A. Hook, and M. Wickens (2007). PUF protein-mediated deadenylation is catalyzed by Ccr4p. J. Biol. Chem. 282, 109-114.
- Kwak, J.E. and M. Wickens (2007) A family of poly(U) polymerases. RNA, 13, 860-867.
- Hook, B., Goldstrohm, A.C., Seay, D.J. and M. Wickens (2007) Two yeast PUF proteins negatively regulate a single mRNA. J. Biol. Chem, 282, 15430-15438.
- B. Thompson, M. Wickens and J. Kimble. 2007. Translational Control in Development. In Translational Control in Biology and Medicine (Michael Mathews, Nahum Sonenberg, and John W.B. Hershey eds.) Cold Spring Harbor Press, New York, Monograph Series 48, pp. 507-544.
- J. Coller and M. Wickens. 2007. Tethered function assays: An adaptable approach to study RNA regulatory proteins. In Translation Initiation: Extract Systems and Molecular Genetics (Jon Lorsch, ed.) Methods in Enzymology Series. Elsevier, Inc. Vol. 429, pp. 299-321.
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Progress 01/01/06 to 12/31/06
Outputs
We have identified the basis of specificity in certain mRNA-protein interactions and are now developing methods to manipulate proteins to obtain new, designer proteins with novel specificities. We have identified new classes of poly(A) and poly(V) adding enzymes. C. elegans participates in the control of stem cell proliferation and differentiation. We have begun the poly(A) adding enzyme studies of these proteins in the nervous system.
Impacts
The work has led to two patents. The objects are to manipulate the fate and function of mRNAs. In addition, both the RNA-binding proteins and the polY(A) adding enzymes may have critical roles in stem cells.
Publications
- Lee, M.H., B. Hook, L.B. Lamont, M. Wickens, and J. Kimble (2006). LIP-1 phosphatase controls the extent of germline proliferation in Caenorhabditis elegans. EMBO J. 25, 88-96.
- Goldstrohm, A.C., B.A. Hook, D.J. Seay, and M. Wickens (2006) PUF proteins bind Pop2p to regulate messenger RNAs. Nature Structural & Molecular Biology. 13, 533-539.
- Seay, D., B. Hook, K. Evans, and M. Wickens (2006) A three-hybrid screen identifies mRNAs controlled by a regulatory protein. RNA. 12, 1594-1600.
- Suh N, Jedamzik B, Eckmann CR, Wickens M and Kimble J. (2006). The GLD-2 poly(A) polymerase activates gld-1 mRNA in the Caenorhabditis elegans germ line. Proc Natl Acad Sci U.S.A. 103, 15108-15112.
- Rouhana, L. and M. Wickens. (2007), Autoregulation of GlD-2 cytoplasmic poly(A) polymerase. RNA, 13, 1-12.
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Progress 01/01/05 to 12/31/05
Outputs
We have identified the basis of specificity in certain mRNA-protein interactions and are now developing methods to manipulate proteins to obtain new, designer proteins with novel specificities. We have identified a new class of poly(A) adding enzymes and have shown that in C. elegans, this enzyme participates in the control of stem cell proliferation and differentiation.
Impacts
The work has led to two patents. The objects are to manipulate the fate and function of mRNAs. In addition, both the RNA-binding proteins and the polY(A) adding enzymes may have critical roles in stem cells.
Publications
- Hook, B., D. Bernstein, B. Zhang, and M. Wickens. (2005). RNA-protein interactions in the yeast three-hybrid system: affinity, sensitivity and enhanced library screening. RNA, 11, 227-233.
- Bernstein, D., B. Hook, A. Hajarnavis, L. Opperman, and M. Wickens, (2005). Binding specificity and mRNA targets of a C. elegans PUF protein, FBF-1, RNA. 11, 447-458.
- Rouhana, L., L. Wang, N. Buter, J.E. Kwak, C. Schiltz, T. Gonzalez, A. Kelley, C.F. Landry, and M. Wickens. (2005). Vertebrate GLD2 poly(A) polymerases in the germline and the brain. RNA, 11, 1117-1130.
- Thompson, B.E., D.S. Bernstein, J.L. Bachorick, A.G. Petcherski, M. Wickens, and J. Kimble. (2005). Dose-dependent control of proliferation and sperm specification by FOG-1/CPEB. Development. 132, 3471-3481.
- Opperman, L., B. Hook, M. DeFino, D.S. Bernstein, and M. Wickens. (2005). A single spacer nucleotide determines the specificities of two mRNA regulatory proteins. Nature Structural & Molecular Biology. 12, 945-951.
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Progress 01/01/04 to 12/31/04
Outputs
We have identified the basis of specificity in certain mRNA-protein interactions and are now developing methods to manipulate proteins to obtain new, designer proteins with novel specificities. We have identified a new class of poly(A) adding enzymes and have shown that in C. elegans, this enzyme participates in the control of stem cell proliferation and differentiation.
Impacts
The work has led to two patents. The objects are to manipulate the fate and function of mRNAs. In addition, both the RNA-binding proteins and the polY(A) adding enzymes may have critical roles in stem cells.
Publications
- S.E. Butcher and M. Wickens. 2004. STAR-studded circuitry. Nature Structural & Molecular Biology, 11, 2-3.
- Wickens, Marvin and Tania N. Gonzalez (2004). Knives, accomplices and RNA. Science, 306, 1299-1300.
- Kwak, J.E., Wang, L., Ballantyne, S., Kimble, J. and Wickens, M. (2004). Mammalian GLD-2 homologs are poly(A) polymerases. Proc. Natl. Acad. Sci., 101, 4407-4412.
- Wang, L., Kimble, J. and Wickens, M. (2004). Tissue-specific C-to-U editing of gld-2 mRNA in C. elegans: likely C-to-U editing. RNA, 10, 1444-1448.
- Lamont, L.B., S.L. Crittenden, D. Bernstein, M. Wickens, J. Kimble. (2004). FBF-1 and FBF-2 regulate the size of the mitotic region in the C. elegans germline. Dev. Cell. 7, 697-707.
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Progress 01/01/03 to 12/31/03
Outputs
We have identified the basis of specificity in certain mRNA-protein interactions and are now developing methods to manipulate proteins to obtain new, designer proteins with novel specificities. We have identified a new class of poly(A) adding enzymes and have shown that in C. elegans, this enzyme participates in the control of stem cell proliferation and differentiation.
Impacts
The work has led to two patents. The objects are to manipulate the fate and function of mRNAs. In addition, both the RNA-binding proteins and the polY(A) adding enzymes may have critical roles in stem cells.
Publications
- Crittenden, S.L, Eckmann, C.R., Wang, L., Bernstein, D.S., Wickens, M., and Kimble, J. 2003. Regulation of the mitosis/meiosis decision in the Caenorhabditis elegans germline. Phil. Trans. R. Soc. Lond. B, 358:1359-1362.
- Goldstrom, A. and M. Wickens. 2003. A place to die, a place to sleep. Science, 300:753-755.
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Progress 01/01/02 to 12/31/02
Outputs
We have analyzed and identified multiple regulators of mRNA expression in the C. elegans germline, and now are studying their regulation. In addition, we are studying the functions of their relatives in other species, including yeast and vertebrates.The proteins we have discovered likely play roles in many species, including humans. We have patented multiple aspects of our work. Understanding how the regulatory proteins work, and how they are controlled, may pave the way to new treatments and diagnostics.
Impacts
The proteins we have discovered likely play roles in many species, including humans. We have patented multiple aspects of our work. Understanding how the regulatory proteins work, and how they are controlled, may pave the way to new treatments and diagnostics.
Publications
- Eckmann, C.R., Kraemer, B., Wickens, M. and Kimble, J. (2002) GLD-3, a Bicaudal-C homolog that inhibits FBF to control germline sex determination in C. elegans. Developmental Cell, in press.
- M. Wickens, D. Bernstein, J. Kimble and R. Parker. 2002. A PUF family portrait: 3'UTR regulation as a way of life. Trends in Genetics. 18, 150-157.
- D. Bernstein, N. Buter, C. Stumpf and M. Wickens. 2002. Analyzing mRNA-protein complexes using a yeast three-hybrid. In RNA-Protein Interactions (L.E. Maquat, ed.) of the Methods in Enzymology Series. Academic Press, San Diego, CA. Vol. 26, No. 2, pp. 123-141.
- J. Coller and M. Wickens. 2002. Tethered function assays using 3'UTRs. In RNA-Protein Interactions (L.E. Maquat, ed.) of the Methods in Enzymology Series. Academic Press, San Diego, CA. Vol. 26, No. 2, pp. 142-150.
- Crittenden, S.L., Bernstein, D.S, Bachorik, J.L., Thompson, B.E., Gallegos, M., Petcherski, A.G., Moulder, G., Barstead, R., Wickens, M. and J. Kimble (2002) A conserved RNA-binding protein controls germline stem cells in Caenorhabditis elegans. Nature 417, 660-663.
- Wang, L., Eckmann, C.R., Kadyk, L.C., Wickens, M. and J. Kimble (2002) A novel cytoplasmic poly(A) polymerase in Caenorhabditis elegans. Nature 419, 312-316.
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Progress 01/01/01 to 12/31/01
Outputs
We have developed an efficient means to assess the interactions between RNAs and proteins in yeast. We have developed a variety of vectors, plasmids and strains that simplify detecting and analyzing RNA-protein interactions. These vectors enable one to screen a library of cDNAs for proteins that bind to a specific DNA sequence, to screen a library of RNAs for ones that bind specifically to a known protein, and to dissect a known RNA protein interaction in detail.
Impacts
The system has been patented and has been provided to some 400 academic laboratories worldwide. A company now sells a "kit" for research purposes, and several companies have licensed the use of the approach for their needs. Since RNA-protein interactions are vital in disease, viral infection, and fundamental cellular processes required for cell growth and regulation, this system promises to have impact in these areas. To date, it has been used to identify proteins involved in regulating key components of the chromosome, factors involved in chromosome replication, and proteins required for proper sex determination and development.
Publications
- B. Kraemer, B. Zhang, D. SenGupta, S. Fields and M. Wickens, 2000. Using the yeast three-hybrid system to detect and analyze interactions between RNA and protein. In RNA-Ligand Interactions. (Daniel W. Celander and John N. Abelson, eds.) of the Methods in Enzymology Series. Academic Press, New York. Vol. 328: Part III, pp. 297-321.
- K. S. Dickson, S.R. Thompson, N.K. Gray and M. Wickens, 2001. Poly(A) polymerase and the regulation of cytoplasmic polyadenylation. J. Biol. Chem. 276, 41810-41816.
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Progress 01/01/00 to 12/31/00
Outputs
We have developed an efficient means to assess the interactions between RNAs and proteins in yeast. We have developed a variety of vectors, plasmids and strains that simplify detecting and analyzing RNA-protein interactions. These vectors enable one to screen a library of cDNAs for proteins that bind to a specific DNA sequence, to screen a library of RNAs for ones that bind specifically to a known protein, and to dissect a known RNA protein interaction in detail.
Impacts
The system has been patented and has been provided to some 400 academic laboratories worldwide. A company now sells a "kit" for research purposes, and several companies have licensed the use of the approach for their needs. Since RNA-protein interactions are vital in disease, viral infection, and fundamental cellular processes required for cell growth and regulation, this system promises to have impact in these areas. To date, it has been used to identify proteins involved in regulating key components of the chromosome, factors involved in chromosome replication, and proteins required for proper sex determination and development.
Publications
- S. R. Thompson, E.B. Goodwin and M. Wickens. 2000. Rapid deadenylation and poly(A)-dependent translational repression mediated by the C. elegans tra-2 3'UTR in Xenopus embryos. Mol & Cell Biol. 20, 2129-2137.
- A. F. Barkoff, K.S. Dickson, N.K. Gray and Marvin Wickens. 2000. Translational control of cyclin B1 mRNA during meiotic maturation: coordinated repression and cytoplasmic polyadenylation. Developmental Biology 220, 97-109.
- N. K. Gray, J.M. Coller and Marvin Wickens. 2000. Multiple portions of poly(A)-binding protein stimulate translation in vivo through a poly(A)-independent mechanism. EMBO J. 19, 4723-4733.
- C. Luitjens, M. Gallegos, B. Kraemer, J. Kimble and M. Wickens. 2000. CPEB proteins control two key steps in spermatogenesis in C. elegens. Genes Dev. 14, 2596-2609.
- Marvin Wickens, Elizabeth B. Goodwin, Judith Kimble, Sidney Strickland, and Matthias Hentze. 2000. Translational Control in Developmental Decisions. In Translational Control of Gene Expression. (Nahum Sonenberg, John W. B. Hershey, and Michael Mathews, eds.) Cold Spring Harbor Press, Cold Spring Harbor, New York. Monograph 39, pp. 295-370.
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Progress 01/01/99 to 12/31/99
Outputs
Our work has identified a network of regulators that control mRNAs during germline development, and in other biological contexts. The work is sigifnicant for two reasons. It has led to the development of new methods for analyzing and manipulating RNA-protein interactions, and for identifying new regulatory molecules. Both are the subject of patents. Second, it has revealed circuitry in genetic regulation that likely is conserved throughout the animal and plant kingdomws. Manipulation of the circuitry may be useful in controlling fertility.
Impacts
The practical impact of the work is long-term. Both the methods we have developed and the universal biology are of interest in trying to approach clinical problems that involve mRNA-protein interactions. The work does not have obvious environmental or social impact.
Publications
- Eun-gyung Lee, Ashley Yeo, Brian Kraemer, Marvin Wickens and Maxine L. Linial. 1999. The Gag domains required for avian retroviral RNA encapsidation determined by using two independent assays. Journal of Virology 73, 6282-6292.
- Dhruba J. SenGupta, Marvin Wickens and Stanley Fields, 1999. Identification of RNAs that bind to a specific protein using the yeast three-hybrid system. RNA 5, 596-601.
- Dickson, Kirsten S., Bilger, A., Ballantyne, Scott, and Wickens, Marvin. 1999. CPSF in X. laevis oocytes: A cytoplasmic factor involved in regulated polyadenylation. Mol. & Cell. Biol. 19, 5707-5717.
- Kraemer, B., Crittenden, S., Gallegos, M., Moulder, G. Barstead, R. Kimble, J. and M. Wickens. 1999. NANOS-3 and FBF proteins physically interact to control the sperm/oocyte switch in C. elegans. Current Biology 9, 1009-1018.
- Thompson, S.R., E.B. Goodwin and M. Wickens. 2000. Rapid deadenylation and poly(A)-dependent translational repression mediated by the C. Elegans tra-2 3'UTR in Xenopus embryos. Mol & Cell Biol. 20
- Barkoff, A.F., K.S. Dickson, N.K. Gray and Marvin Wickens. 2000. Translational control of cyclin B1 mRNA during meiotic maturation: coordinated repression and cytoplasmic polyadenylation. In Press.
- Gray, N.K., J.M. Coller and Marvin Wickens. 2000. Multiple portions of poly(A)-binding protein stimulate translation in vivo through a poly(A)-independent mechanism. Submitted.
- Astrom, Jonas, S.R. Thompson, and Marvin Wickens. 2000. Activation of an RNA-turnover pathway sensitive to the cap and poly(A) tail during the first cleavage divisions of Xenopus embryos. Submitted.
- Beilin Zhang, Brian Kraemer, Dhruba SenGupta, Stanley Fields and Marvin Wickens. 1999. A three-hybrid system to detect and analyze RNA-protein interactions in vivo. In Expression of Recombinant Genes in Eukaryotic Systems. (J. Glorioso and M. Schmidt, eds.) of the Methods in Enzymology Series. Academic Press, New York. Vol. 306, pp. 93-113.
- Beilin Zhang, Brian Kraemer, Dhruba SenGupta, Stanley Fields and Marvin Wickens. 2000. A yeast three-hybrid system to detect and analyze interactions between RNA and protein. In RNA-Ligand Interactions. (Daniel W. Celander and John N. Abelson, eds.) of the Methods in Enzymology Series. Academic Press, New York. Submitted.
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