EVOLUTION AND ACTIVITY OF AN ERYTHROID CIS-REGULATORY MODULE IN MULTIPLE MAMMALIAN LINEAGES

Open Access
Author:
Abebe, Demesew F
Graduate Program:
Biochemistry, Microbiology, and Molecular Biology
Degree:
Master of Science
Document Type:
Master Thesis
Date of Defense:
April 10, 2009
Committee Members:
  • Ross Cameron Hardison, Thesis Advisor
Keywords:
  • boreoeutheria
  • Motif constraint
  • CIS-REGULATORY MODULE
  • GATA1
Abstract:
Evidence of evolutionary constraint in noncoding regions can be derived from alignments of multiple genomes, and this is a reliable guide to a small but important subset of Cis Regulatory Modules (CRMs). This thesis reports results from several tests conducted in one example of an erythroid CRM (present in human and absent in mice) called GATA1-HS+14. It contains four motifs that match the consensus GATA-1 binding site but none of these are conserved in mouse. Two of these motifs are also present in the reconstructed boreoeutherian ancestor. Our result indicates that HS+14 is an active enhancer in human but a neutral fragment in mouse. It is also an enhancer in the ancestor. In addition, motif C (the only motif conserved in multiple mammals) plays a positive role in enhancement while motifs A and B may play negative role in transcription. Although the level of enhancement varies among species, all tested mammalian DNA fragments that are homologous to HS+14, and having the conserved WGATAR motif, increased transcriptional activity in our enhancement assay. We have also statistical analyzed the region of interest in search of other binding site motifs that are associated with level of enhancement. Six hexamers, with a strong correlation to enhancement level, are identified. These motifs are potentially binding sites for other transcription factors, which may interact with GATA1 to increase transcriptional activity. Although further work is needed, this examination of naturally occurring sequence alterations (in contemporary and ancestral sequences) combined with site-directed mutagenesis of binding site motifs should provide a rich and robust history of evolutionary change and its consequences in mammalian enhancers.