Mailing Address:
University of California at Berkeley
Dept. of Integrative Biology
3060 VLSB #3140
Berkeley, CA 94720-3140
Lab TEL (510) 643-4201
FAX (510) 643-5022
Email: ejayrehm[at]berkeley.edu
Research Summary
My research involves three aspects of our larger effort to establish the amphipod crustacean Parhyale hawaiensis as a new model organism for developmental studies within the arthropods: (1) developing tools for in vivo gene expression, (2) Parhyale genomics, (3) the structural and functional characterization of Parhyale orthologs of Drosophila pair-rule genes, which are dynamically expressed in the developing germ band.
I identified and characterized an endogenous constitutive promoter/enhancer and developed a simple procedure for expressing genes of interest in Parhyale embryos via the microinjection of supercoiled DNA. A genomic region spanning the beginning of a Parhyale Ef1-alpha gene drives strong and constitutive expression of a DsRed marker protein when microinjected as supercoiled DNA into early blastomeres. DsRed protein fluorescence commences during gastrulation, is seen in all tissues, and persists until hatching. This cell autonomous and ubiquitous marker is useful for tracing cell fate patterns: clones produced from the progeny of individually injected blastomeres reveal distinct cell lineages characteristic of Parhyale. This technique will potentially allow us to efficiently assay control regions of developmentally regulated genes for their ability to drive fluorescent protein expression in a predicted manner.
In an effort to characterize the Parhayle genome, and in collaboration with the Joint Genome Institute (JGI), we have undertaken (1) EST sequencing using a normalized embryonic cDNA library and (2) directed BAC sequencing of 70 BAC clones containing developmentally interesting genes. In addition, (3) I will shortly provide JGI with another supply of Parhyale embryonic cDNA to be used for 454 sequencing. We anticipate that a few hundred thousand of the very short reads provided by 454 sequencing will significantly complement the roughly 13,000 clusters generated by paired-end sequencing 30,000 cDNA clones. This additional sequence information should facilitate annotation of the existing clusters as well as reveal additional genes. Finally, while sequencing the Parhyale genome is cost prohibitive due to its large size (3,600 Mb), (4) we are preparing to sequence the genome of a closely related amphipod crustacean with an unusually small genome, Jassa slatteyri.
http://www.jgi.doe.gov/sequencing/why/CSP2006/crustaceans.html
http://www.genome.gov/10002154
http://www.genome.gov/Pages/Research/Sequencing/SeqProposals/EcdysozoaProposalFinalPDF.pdf.
Following a candidate gene approach, graduate student Ron Parchem cloned a number of Parhyale orthologs of Drosophila pair-rule genes by means of degenerate PCR. Five odd-skipped (odd) family members and a two odd-paired (opa) family members have been identified. A number of these odd and opa orthologs are dynamically expressed in the segmenting germ band: odd-1, odd-2, odd-5, and opa-1. I have been working to characterize these genes and investigate their role in segmentation. I generated full-length cDNA clones for each ortholog. It has become clear that at least one of these genes, opa-1, encodes alternatively spliced mRNAs that result in two embryonic transcripts with distinct 3’ termini. I have designed in situ RNA probes specific to each of these variant transcripts to determine their respective patterns of expression. I have also identified BAC clones containing each of the 7 Parhyale odd and opa orthologs, which will shortly be sequenced. Genomic sequence flanking these genes should provide important information concerning their cis-regulation. To investigate gene function I have built transgenic over-expression constructs for those odd and opa orthologs expressed in the germband. I currently have transgenic over-expression lines established for one odd-skipped (odd-2) and one odd-paired gene (opa-1). To compliment these experiments I am also attempting to knockdown gene function by injecting small targeted siRNAs into Parhyale embryos.
Publications
Rehm, E. J, R. Hannibal, R.C. Chaw, M. Vargas-Vila and N. H. Patel. 2009. The Crustacean Parhyale hawaiensis: A New Model for Arthropod Development. In Emerging Model Organisms: A Laboratory Manual, Volume 1 (Cold Spring Harbor Press, NY), pp 373-404.
Rehm, E. J. and N. H. Patel. 2005. Gene Expression from DNA Injected into Embryos of the Amphipod Crustacean Parhyale hawaiensis. 64th Annual Society for Developmental Biology Meeting, San Francisco, CA.
Sink, H., E. J. Rehm, L. Richstone, Y. M. Bulls, and C. S. Goodman. 2001. sidestep encodes a target-derived attractant essential for motor axon guidance in Drosophila. Cell 105(1): 57-67.
Sink, H., E. J. Rehm, L. Richstone, Y. M. Bulls, and C. S. Goodman. 2001. Motor axon guidance in the embryonic periphery is mediated by the Sidestep protein. 42nd Annual Drosophilia Research Conference, Washington, D. C.
Huang, A., E. J. Rehm, G. M. Rubin. 2000. Recovery of DNA Sequences Flanking P-element Insertions: Inverse PCR and Plasmid Rescue. In Drosophila Protocols. Sullivan, W., M. Ashburner and R. S. Hawley, eds. Cold Spring Harbor Laboratory Press, 429-437.
Liao, G. C., E. J. Rehm, G.M. Rubin. 2000. Insertion site preferences of the P transposable element in Drosophila melanogaster. Proceedings of the National Academy of Sciences U. S. A. 97:7 3347-51.
Spradling, A., D. Stern, A. Beaton, E. J. Rehm, T. Laverty, N. Mozden, S. Misra, G. M. Rubin. 1999. The Berkeley Drosophila Genome Project Gene Disruption Project: Single P-Element Insertions Mutating 25% of Vital Drosophila Genes. Genetics 153: 135-177.
Rorth, P., K. Szabo, A. Bailey, T. Laverty, J. Rehm, G. M. Rubin, K. Weigmann, M. Milan, V. Benes, W. Ansorge and S. M. Cohen. 1998. Systematic gain-of-function genetics in Drosophila. Development 125(6): 1049-57.
Hayward, D. C., N. Patel, E. J. Rehm, C. S. Goodman, and E. E. Ball. 1995. Sequence and Expression of the Antennapedia Gene of the Grasshopper, Shistocerca americana: comparison to Drosophila. Developmental Biology 172(2): 452-465.
McAllister, L., E. J. Rehm, C. S. Goodman and Kai Zinn. 1992. Alternative splicing of micro-exons creates multiple forms of the insect cell adhesion molecule fasciclin I. Journal of Neuroscience 12(3): 895-905.
Grenningloh, G., E. J. Rehm and C. S. Goodman. 1991. Genetic analysis of growth cone guidance in Drosophila: fasciclin II functions as a neuronal recognition molecule. Cell 67(1): 45-57.
Ball, E. E., E. J. Rehm, and C. S. Goodman. 1991. Cloning of a grasshopper c DNA coding for a protein homologous to the A1, A2/B1 proteins of mammalian hnRNP. Nucleic Acids Research 19(2): 397.
Grenningloh, G., A. J. Bieber, E. J. Rehm, P. M. Snow, Z. R. Traquina, M. Hortsch, N. Patel, and C. S. Goodman. 1990. Molecular Genetics of neuronal recognition in Drosophila: evolution and function of immunoglobulin superfamily cell adhesion molecules. Cold Spring Harbor Symp. Quant. Biol., Molecular Neurobiology 55: 327-340.
Education
WHITNEY MUSEUM INDEPENDENT STUDY PROGRAM
New York, NY
2002-2003
SCHOOL OF THE ART INSTITUTE OF CHICAGO
Chicago, Illinois
1998-2001
BFA in Fine Art, 2001
UNIVERSITY OF CALIFORNIA AT BERKELEY
Berkeley, California
1997-1998
Art History
STANFORD UNIVERSITY
Stanford, California
1982-1986
AB in History, 1987
BS in Biology, 1987
Additional Research Experience
SENIOR RESEARCH TECHNOLOGIST II (1999 – 2002)
Laboratory of Professor Susan Lindquist, Department of Molecular Genetics and Cellular Biology, University of Chicago, Howard Hughes Medical Institute
STAFF RESEARCH ASSOCIATE III (1995 – 1998)
Berkeley Drosophila Genome Project (BDGB), Professor Gerald Rubin, Director, Department of Molecular and Cellular Biology, Division of Genetics, University of California-Berkeley, Howard Hughes Medical Institute
STAFF RESEARCH ASSOCIATE II (1987 – 1995)
Laboratory of Professor Corey S. Goodman, Department of Molecular and Cellular Biology, Division of Neurobiology, University of California-Berkeley, Howard Hughes Medical Institute
My research involves three aspects of our larger effort to establish the amphipod crustacean Parhyale hawaiensis as a new model organism for developmental studies within the arthropods: (1) developing tools for in vivo gene expression, (2) Parhyale genomics, (3) the structural and functional characterization of Parhyale orthologs of Drosophila pair-rule genes, which are dynamically expressed in the developing germ band.
I identified and characterized an endogenous constitutive promoter/enhancer and developed a simple procedure for expressing genes of interest in Parhyale embryos via the microinjection of supercoiled DNA. A genomic region spanning the beginning of a Parhyale Ef1-alpha gene drives strong and constitutive expression of a DsRed marker protein when microinjected as supercoiled DNA into early blastomeres. DsRed protein fluorescence commences during gastrulation, is seen in all tissues, and persists until hatching. This cell autonomous and ubiquitous marker is useful for tracing cell fate patterns: clones produced from the progeny of individually injected blastomeres reveal distinct cell lineages characteristic of Parhyale. This technique will potentially allow us to efficiently assay control regions of developmentally regulated genes for their ability to drive fluorescent protein expression in a predicted manner.
In an effort to characterize the Parhayle genome, and in collaboration with the Joint Genome Institute (JGI), we have undertaken (1) EST sequencing using a normalized embryonic cDNA library and (2) directed BAC sequencing of 70 BAC clones containing developmentally interesting genes. In addition, (3) I will shortly provide JGI with another supply of Parhyale embryonic cDNA to be used for 454 sequencing. We anticipate that a few hundred thousand of the very short reads provided by 454 sequencing will significantly complement the roughly 13,000 clusters generated by paired-end sequencing 30,000 cDNA clones. This additional sequence information should facilitate annotation of the existing clusters as well as reveal additional genes. Finally, while sequencing the Parhyale genome is cost prohibitive due to its large size (3,600 Mb), (4) we are preparing to sequence the genome of a closely related amphipod crustacean with an unusually small genome, Jassa slatteyri.
http://www.jgi.doe.gov/sequencing/why/CSP2006/crustaceans.html
http://www.genome.gov/10002154
http://www.genome.gov/Pages/Research/Sequencing/SeqProposals/EcdysozoaProposalFinalPDF.pdf.
Following a candidate gene approach, graduate student Ron Parchem cloned a number of Parhyale orthologs of Drosophila pair-rule genes by means of degenerate PCR. Five odd-skipped (odd) family members and a two odd-paired (opa) family members have been identified. A number of these odd and opa orthologs are dynamically expressed in the segmenting germ band: odd-1, odd-2, odd-5, and opa-1. I have been working to characterize these genes and investigate their role in segmentation. I generated full-length cDNA clones for each ortholog. It has become clear that at least one of these genes, opa-1, encodes alternatively spliced mRNAs that result in two embryonic transcripts with distinct 3’ termini. I have designed in situ RNA probes specific to each of these variant transcripts to determine their respective patterns of expression. I have also identified BAC clones containing each of the 7 Parhyale odd and opa orthologs, which will shortly be sequenced. Genomic sequence flanking these genes should provide important information concerning their cis-regulation. To investigate gene function I have built transgenic over-expression constructs for those odd and opa orthologs expressed in the germband. I currently have transgenic over-expression lines established for one odd-skipped (odd-2) and one odd-paired gene (opa-1). To compliment these experiments I am also attempting to knockdown gene function by injecting small targeted siRNAs into Parhyale embryos.
Publications
Rehm, E. J, R. Hannibal, R.C. Chaw, M. Vargas-Vila and N. H. Patel. 2009. The Crustacean Parhyale hawaiensis: A New Model for Arthropod Development. In Emerging Model Organisms: A Laboratory Manual, Volume 1 (Cold Spring Harbor Press, NY), pp 373-404.
Rehm, E. J. and N. H. Patel. 2005. Gene Expression from DNA Injected into Embryos of the Amphipod Crustacean Parhyale hawaiensis. 64th Annual Society for Developmental Biology Meeting, San Francisco, CA.
Sink, H., E. J. Rehm, L. Richstone, Y. M. Bulls, and C. S. Goodman. 2001. sidestep encodes a target-derived attractant essential for motor axon guidance in Drosophila. Cell 105(1): 57-67.
Sink, H., E. J. Rehm, L. Richstone, Y. M. Bulls, and C. S. Goodman. 2001. Motor axon guidance in the embryonic periphery is mediated by the Sidestep protein. 42nd Annual Drosophilia Research Conference, Washington, D. C.
Huang, A., E. J. Rehm, G. M. Rubin. 2000. Recovery of DNA Sequences Flanking P-element Insertions: Inverse PCR and Plasmid Rescue. In Drosophila Protocols. Sullivan, W., M. Ashburner and R. S. Hawley, eds. Cold Spring Harbor Laboratory Press, 429-437.
Liao, G. C., E. J. Rehm, G.M. Rubin. 2000. Insertion site preferences of the P transposable element in Drosophila melanogaster. Proceedings of the National Academy of Sciences U. S. A. 97:7 3347-51.
Spradling, A., D. Stern, A. Beaton, E. J. Rehm, T. Laverty, N. Mozden, S. Misra, G. M. Rubin. 1999. The Berkeley Drosophila Genome Project Gene Disruption Project: Single P-Element Insertions Mutating 25% of Vital Drosophila Genes. Genetics 153: 135-177.
Rorth, P., K. Szabo, A. Bailey, T. Laverty, J. Rehm, G. M. Rubin, K. Weigmann, M. Milan, V. Benes, W. Ansorge and S. M. Cohen. 1998. Systematic gain-of-function genetics in Drosophila. Development 125(6): 1049-57.
Hayward, D. C., N. Patel, E. J. Rehm, C. S. Goodman, and E. E. Ball. 1995. Sequence and Expression of the Antennapedia Gene of the Grasshopper, Shistocerca americana: comparison to Drosophila. Developmental Biology 172(2): 452-465.
McAllister, L., E. J. Rehm, C. S. Goodman and Kai Zinn. 1992. Alternative splicing of micro-exons creates multiple forms of the insect cell adhesion molecule fasciclin I. Journal of Neuroscience 12(3): 895-905.
Grenningloh, G., E. J. Rehm and C. S. Goodman. 1991. Genetic analysis of growth cone guidance in Drosophila: fasciclin II functions as a neuronal recognition molecule. Cell 67(1): 45-57.
Ball, E. E., E. J. Rehm, and C. S. Goodman. 1991. Cloning of a grasshopper c DNA coding for a protein homologous to the A1, A2/B1 proteins of mammalian hnRNP. Nucleic Acids Research 19(2): 397.
Grenningloh, G., A. J. Bieber, E. J. Rehm, P. M. Snow, Z. R. Traquina, M. Hortsch, N. Patel, and C. S. Goodman. 1990. Molecular Genetics of neuronal recognition in Drosophila: evolution and function of immunoglobulin superfamily cell adhesion molecules. Cold Spring Harbor Symp. Quant. Biol., Molecular Neurobiology 55: 327-340.
Education
WHITNEY MUSEUM INDEPENDENT STUDY PROGRAM
New York, NY
2002-2003
SCHOOL OF THE ART INSTITUTE OF CHICAGO
Chicago, Illinois
1998-2001
BFA in Fine Art, 2001
UNIVERSITY OF CALIFORNIA AT BERKELEY
Berkeley, California
1997-1998
Art History
STANFORD UNIVERSITY
Stanford, California
1982-1986
AB in History, 1987
BS in Biology, 1987
Additional Research Experience
SENIOR RESEARCH TECHNOLOGIST II (1999 – 2002)
Laboratory of Professor Susan Lindquist, Department of Molecular Genetics and Cellular Biology, University of Chicago, Howard Hughes Medical Institute
STAFF RESEARCH ASSOCIATE III (1995 – 1998)
Berkeley Drosophila Genome Project (BDGB), Professor Gerald Rubin, Director, Department of Molecular and Cellular Biology, Division of Genetics, University of California-Berkeley, Howard Hughes Medical Institute
STAFF RESEARCH ASSOCIATE II (1987 – 1995)
Laboratory of Professor Corey S. Goodman, Department of Molecular and Cellular Biology, Division of Neurobiology, University of California-Berkeley, Howard Hughes Medical Institute
NIPAM H. PATEL
PHOTO ABOVE LEFT: A day old Parhyale embryo that was injected at the 1-cell stage with supercoiled plasmid construct pECE-DsRed-NLS. This construct consists of a ~9kb promoter/enhancer fragment from an endogenous Ef1-alpha gene that constitutively drives expression of a nuclear localized variant of DsRed fluorescent protein. In fact, expression appears to be sub-nuclear, for unknown reasons. The nuclei of all cells are fluorescing. Fluorescence first appears at around 18 hours, about the time of gastrulation. We believe that this coincides with the onset of zygotic transcription. In the pictured embryo a germdisc has formed at the anterior end of the egg (top). A fluorescent image has been overlaid on a brightfield image.
PHOTO ABOVE RIGHT: Fluorescent image of a day old Parhyale embryo that was injected at the 1-cell stage with supercoiled plasmid construct pECE-Timer. In this case, the ef1alpha promoter/enhancer drives cytoplasmic expression of Timer, a DsRed variant that fluoresces green for a few hours before the protein folds into its mature conformation, at which point it fluoresces red. In the pictured embryo a germdisc has formed at the anterior end of the egg (top).
PHOTO ABOVE RIGHT: Fluorescent image of a day old Parhyale embryo that was injected at the 1-cell stage with supercoiled plasmid construct pECE-Timer. In this case, the ef1alpha promoter/enhancer drives cytoplasmic expression of Timer, a DsRed variant that fluoresces green for a few hours before the protein folds into its mature conformation, at which point it fluoresces red. In the pictured embryo a germdisc has formed at the anterior end of the egg (top).
PHOTO ON LEFT: Parhyale odd-paired (opa-1) expression in the germband (anterior is up). The darker yellow line traces the first of two highly organized rounds of mitotic division, which begin medially before spreading laterally as a wavefront. Below this lie transverse rows of parasegment precursor cells, each of which represents a future parasegment. The lighter yellow line indicates the second mitotic wavefront, which results in a parasegment containing 4 cell rows. As is clear from the false-colored in situ (red), opa-1 expression begins shortly after the first mitotic division. It is expressed only in the more posterior row of daughter cells following the first mitotic event. Anterior to the germband, opa-1 is expressed in appendage fields of the head parasegments, which are further along in development. (In situ image courtesy of Ron Parchem.)