Publication Date
2022
Document Type
Thesis
Committee Members
Courtney E.W. Sulentic, Ph.D. (Advisor); Ravi P. Sahu, Ph.D. (Committee Member); Mike Kemp, Ph.D. (Committee Member); Khalid Elased, Pharm.D., Ph.D. (Committee Member)
Degree Name
Master of Science (MS)
Abstract
The 3'IGHRR is thought to be responsible for the transcription of the immunoglobulin heavy chain (IGH) gene locus, which is essential for the production of antibodies. The human 3’IGHRR, which is duplicated in humans, contains the hs3, hs4, and hs1.2 enhancers. Additionally, the hs1.2 enhancer is polymorphic in humans and consists of a 53 bp invariant sequence that can be repeated one to four times. Previous experiments in a mouse and human B-cell line model have shown that the high affinity AhR ligand and environmental toxicant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) can induce hs1.2 enhancer activity in an AhR-dependent manner. The AhR is a transcription factor that can modulate gene transcription either directly through its transactivation domain or indirectly by interacting with other transcription factors. Further studies with a human B-cell line model (CL-01) that lacks AhR transactivation function demonstrated TCDD-induced activation of the hs1.2 enhancer that was not influenced by the number of the invariant sequence repeat. These studies suggest that a functional transactivation domain may not be necessary for the AhR to mediate TCDD-induced activation of the hs1.2 enhancer. The objective of this study is to assess whether a functional AhR transactivation domain enhances TCDD-induced hs1.2 enhancer activity and alters the transcriptional impact of an increased number of invariant sequences. For this experiment, wildtype CL-01 cells lacking AhR transactivation function and CRISPR/Cas9-edited CL-01 cells expressing an AhR with a functional transactivation domain were transfected via electroporation with luciferase reporter plasmids that contain one of the four human polymorphic hs1.2 enhancer alleles (α1A, α1B, α1C, or α1D corresponding to one, two, three, or four invariant sequence repeats, respectively) and treated with different concentrations of TCDD. Results demonstrated no difference in TCDD-induced activation of the hs1.2 reporter in cells with or without a functional AhR transactivation domain. Additionally, compared to previous results with a mouse model, these studies suggest a marked species difference in the sensitivity of the hs1.2 enhancer to AhR ligands. Given the importance of antibodies in maintaining immunocompetence and that our current knowledge of B-cell biology is primarily based on mouse models, this work underscores the gap in understanding of human antibody production and the ability to assess risk of exposure to environmental stressors and individual sensitivity to disease.
Page Count
74
Department or Program
Department of Pharmacology and Toxicology
Year Degree Awarded
2022
Copyright
Copyright 2022, all rights reserved. My ETD will be available under the "Fair Use" terms of copyright law.
ORCID ID
0000-0001-5563-5271
