F. Bryan Pickett
Assistant Department Chair
Zebrafish Transgenics as a System to Analyze the Transcriptional Regulation of the Retinoid Synthesis Network and Heart Contractile Protein Functions in Vertebrates
The role of patterned expression of the Retinoic Acid biosynthetic enzymes in zebrafish embryonic pattern formation is controversial. Experimental data in support of alternative hypotheses stating that patterned RA synthesis, or degradation, or synthesis combined with degradation is important for producing instructive bursts and gradients of RA accumulation in the embryo has been presented in the literature. To better understand the regulation of the biosynthetic enzyme pathway producing RA, we are defining minimal promoter/enhancer regions for biosynthetic enzymes, and are doing traditional promoter analysis and evolutionary conserved region analysis to define the cis regulatory modules that drive expression of these genes. Using the aldh1a2/RaldH2 promoter, we have defined a minimal region that recapitulates most normal expression as assessed by YFP/fluorescent reporter expression, and are using a combination of deletion analysis, enhancer testing, and epigenetic regulation analysis to better understand the control modularity of this region. Regions of interest have recently been shown to be evolutionarily conserved in humans, and to be the targets of hypermethylation in human cancer cell lines, thus our fish study may have translational value if human aldh1a is valuable as a potential tumor suppressor gene in human cancers. Identification of specific, evolutionarily conserved, regulators of the expression of this gene in fish may lead to identification of protein targets for small molecules that might be useful in activating the expression of this gene in a tumor context in man.
In collaboration with the laboratory of Dr. Pieter de Tombe, Stritch School of Medicine, we are also developing the zebrafish as a system to study the role of the cardiac contractile apparatus in vertebrate heart physiology and disease. We are partially and completely humanizing fish contractile proteins and are also testing specific mutations to determine if target genes may play a role in the Frank-Startling mechanism of the regulation of cardiac output.
The Evolutionary Fate of Genes and Networks After Gene and Genome Duplication
In collaboration with Drs. Allan Force and William Cresko we are developing simulations to test the impact of the Duplication, Degeneration and Complementation (DDC) model of gene functional evolution, which we co-developed with Dr. Force (Force, Lynch, Pickett et al 1999), on the evolution of whole genomic genetic network architectures and the total information of evolved interaction networks. This work begins to extend our gene level model to the level of the whole genome. Initial results of our modeling suggest that network informational complexity may act as an internal constraint on the number and density of redundant network interactions that are maintained after whole or partial genome duplication, depending in part on the mutational patterns that resolve redundancies and the level at which redundancy endures in the network over time. This work has surprising connections to the missing heritability problem in quantitative genetics.
Meristem Function and Embryonic Patterning in the Plant Arabidopsis thaliana
The shoot apical meristem of higher plants acts similarly to animal embryos, providing stem cells for organization into tissues and organs. My laboratory uses a genetic approach to identify mutations in the plant Arabidopsis thaliana which perturb the function of the shoot apical meristem. Since organ production is the main job of the meristem, we have screened for temperature sensitive mutations that fail to produce new leaves and other organs following germination.
Three mutations with strict developmental arrests at high temperature have been identified to date. I have designated these mutations arrested development (add) 1, 2, and 3. Current and future projects include the molecular cloning of the ADD1 gene because this mutation was induced by the disruption of the gene by insertion of foreign DNA. This "DNA tag" has been used to isolate genomic DNA near the point of insertion and has facilitated the molecular characterization of the ADD1 region. The biochemical characterization of this gene will help shed light on the molecular mechanism of plant meristem function.
Fig. Wild-type and mutant seedling development after germination and growth at 29oC for 6 days. (A) Wild-type Wassilewskija seedling with cotyledons and two true leaves. The apical region of an arrested development 1 (add1) seedling lacking true leaves (B). Wild-type Landsberg erecta seedling (C) with an epicotyl displaying the first 2 true leaves. Both leaves have developed leaf blades and trichomes. (D) arrested development 2 seedling with cotyledons, true primordia are not visible.
Force, A., W. A. Cresko, F. B. Pickett, S. R. Proulx, C. Amemiya, and M. Lynch. 2005. The origin of subfunctions and modular gene regulation. Genetics 170:433-446.
Force, A., Cresko, W.F. and Pickett, F.B. (2004). Informational accretion, gene duplication, and the mechanisms of genetic module parcellation. in Modularity in Development and Evolution: G. Schlosser and G. Wagner, Eds. University of Chicago Press.
Prince, V.E. and Pickett, F.B. (2002). Splitting Pairs: Diverging fates of duplicated genes. Nature Reviews: Genetics. 3: 827-837 .
Saulsberry, A., Martin, P.R., O'Brien, T., Sieburth, L.E. and Pickett, F.B. (2002). The induced sector Arabidopsis apical embryonic fate map. Development 129: 3403-3410.
Woodrick, R., Martin, P. R., Birman,I., and Pickett, F.B. (2000) The Arabidopsis Embryonic Shoot Fate Map . Development 127:813-820.
Force, A., Lynch, M., Pickett, F. B., Amores, A., Yan, Y-L and Postlethwait, J. (1999). Preservation of Duplicate Genes by Complementary, Degenerative Mutations. Genetics 151: 1531-1545
Leyser, H.M.O., Pickett, F.B., Dharmaseri, S., and Estelle, M. (1996). Mutations in the AXR3 Gene of Arabidopsis Result in Altered Auxin Response. The Plant Journal, 10: 403-413.
Pickett, F.B., Champagne, M.M. and Meeks-Wagner, D.R., (1996). Temperature-Sensitive Mutations that Arrest Arabidopsis Shoot Development. Development 122, 3799-3807.
Pickett, F.B. and Meeks-Wagner, D.R., (1995). Seeing Double: Appreciating Genetic Redundancy. The Plant Cell 7: 1347-1356.
Timpte, C., Lincoln, C., Pickett, F.B., Turner, J., and Estelle, M., (1995). The AXR1 and AUX1 Genes of Arabidopsis Function in Separate Auxin Responsive Pathways. The Plant Journal, 8: 561-569.