Loyola University Chicago

 Synthetic Evolution and  Developmental Genetics of Arabidopsis Thaliana

by Paul Roman Zurek

Department of Biology
Chicago, Illinois

Thursday, July  17, 2008, 10:00 a.m.
Life Science Building, Room 412

ABSTRACT

          The Duplication, Degeneration, Complementation (DDC) model concerns the evolutionary preservation of duplicated genes.  As originally described by Force, Lynch and Pickett, the DDC model suggests that newly duplicated, multifunctional genes can partition the disparate functions present in the ancestral gene between the two new paralogs through the accumulation of partial loss of function mutations.  These mutations must impact discretely mutable cis- regulatory elements, protein functional modules, splice sites or other regulatory or functional regions of a gene locus.  In the parlance of the DDC model these types of elements in a gene are called “sub-functions” of the gene. A series of mutational events ultimately results in the two paralogs having complementary sets of sub-functions. Thus in order for the whole suite of ancestral functions to be available to the organism, both of the paralogs have to be maintained within the genome.

          The modifications of expression and/or function required by the DDC process depend on mutational changes to the sub-functions, and therefore an evolutionary time scale and small population numbers are also required. A faster approach to subfunction partitioning may be caused by epigenetic regulation of subfunctions. Epigenic changes could cause rapid initial subfunctionalization, which could in turn predispose the DNA to more permanent mutational changes. The goal of this project was to create a synthetic duplication event using a 5’ cis regulatory region from a multifunctional gene, fused to one of three possible fluorescent proteins. Two of these constructs could then be introduced into the same plant background and a large number of progeny seedlings observed for any signs of epigenetic subfunctionalization simply by looking at the pattern of partitioning of expression patters of both of the constructs.  If rapid epigenetic subfunctionalization occurs, the state of methylation or level of chromatin condensation could be assessed for known cis regulatory regions, helping to determine if mechanisms exist to rapidly bring duplicate genes under independent selection regimes.

           Complementing my synthetic evolution project, I am also participating in the analysis of the genetic regulation of plant development.  The arrested development 1 (add1) gene is an Arabidopsis thaliana gene that was identified by mutation as a part of a screen for temperature sensitive developmental mutants. The mutant phenotype causes an arrest in epicotyl development and results in plants with a severely limited number of leaves. At permissive temperatures (21°C) epicotyl development is slowed down, but true leaves are produced as in wildtype plants. The few true leaves produced at high temperatures (29°C) have a needle-like appearance. This indicates that add1 might play a role in the shoot apical meristem because the meristem is required for both the initiation of leaves, as well as for establishing leaf blade dorsoventrality.

            To study this gene further, two reporter constructs were made using either the green fluorescent protein or the β-Glucoronidase (GUS), driven by the add1 promoter region. The fluorescent protein reporter was used because plants remain viable while this reporter is analyzed, while the GUS reporter was used because of the large amount of signal amplification inherent in this reporter. The expression was driven using the promoter region upstream of the add1 coding region. As expected, the reporter proteins were seen to accumulate both in the shoot apical meristem and true leaves. Additionally the reporter proteins also accumulated in other types of meristematic plant tissues, like the root apical meristem, axilary meristem and the cambium.

VITA

         Paul Roman Zurek attended St. Patrick High School before attending the Loyola University Chicago, College of Arts and Sciences. He graduated cum laude with honors after majoring in Biology and obtaining a minor in Chemistry. During his last year and a half as an undergraduate, he volunteered his time in the lab of Dr. F. Bryan Pickett, and then stayed in the lab as a graduate student. He has been admitted to the Duke University Program in Genetics and Genomics and will begin his doctoral studies in Fall 2008.

ACKNOWLEDGMENT

        I would like to thank both of my advisors, Dr. F. Bryan Pickett and Ms. Paula Martin for getting me through my time as an undergraduate and graduate student. I would like to thank my committee, Dr. William Rochlin and Dr. Kim Williamson. I also want to offer thanks to both Matt Kolinski and Kara Nordin, two undergraduate students that contributed heavily to both projects. I would also like to thank Dr. Terry Grande, Ms. Audrey Berry, Ms. Pam Bradley, and Mr. Joe Schleup. Next, I would like to thank Drs. Eric Lam of Rutgers University and Naohiro Kato of Louisiana State University for providing both the eYFP and eGFP vectors, as well as Dr. Eric Schroeter for providing the Cerulean florescent protein. Lastly I would like to thank my family for their support through the whole process.
       My work in the Pickett lab was supported by federal research grants to Dr. Pickett from the National Institutes of Health, the U.S. Department of Agriculture and the National Science Foundation.

COMMITTEE MEMBERS:

Dr. F. Bryan Pickett
Dr. M. William Rochlin
Dr. Kim Williamson