KLF1 Effects on γ-Globin Regulation

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KLF1 Effects on γ-Globin Regulation Ashria Arora

β– and γ – Globin Genes Beta globin expressed in adulthood Gamma globin expressed fetal erythroblasts Transcription factors and repressor involved Mutation in the upstream promoter can cause: -beta thalasemia ~production of low to none beta globin ~production and expression of γ – globin to compensate for beta

KLF1 Krüppel-like transcription factors -Krüppel is a gap gene, encodes zinc fingers ~Cys/His zinc fingers -Family of 17 factors -Previous studies proven KLF1 regulates transcription of beta globin ~KLF1 null can cause beta thalasemia- patient with null

Research Question Is the overexpression of KLF1 a positive or negative regulator γ-globin genes?

Experiment Human fetal erythroid cell line along with zebrafish fetal cell line -control- zebrafish, human adult cell with no plasmid construct Plasmid constructs Luciferase assay

Methods Plasmid constructs -KLF1, LUC, and γCACCCLuc will be generated -KLF1 replicated inside human cell line to produce the overexpression of KLF1 (recobinant DNA) -Generating expression constructs, luciferase assay Luciferase assay- firefly enzyme Emission of light is measured to determine expression of γ-globin genes, CAACCLuc allows More light, mean transcription of γ-globin

Possible Results Zebrafish represents the expression w/o KLF1, human with normal levels of KLF1, and human with the overexpression Possible outcomes: Light emitted to be similar to control (human cell line, normal levels), overexprssion has no effect Light emitted to be less than control, therefore overexpression negatively regulates γ-globin Light emitted more than control levels, overexpression positively regulates