Title

Deregulated Survival Pathways Confer Erythropoietin-Independent Survival and Proliferation in Erythroleukemia Progenitor Cells

Document Type

Thesis

Degree Name

Master of Science (MS)

Department

Biological and Environmental Sciences

Date of Award

Fall 2020

Abstract

Abnormal erythropoietin (Epo) signaling in erythroid precursor cells is a characteristic of Acute Erythroid Leukemia (AEL). AEL is defined by the uncontrolled proliferation of maturation-arrested erythroid precursor cells. Since Epo signaling controls normal erythropoiesis, we hypothesize that dysregulated survival pathways confer Epo-independent survival and proliferation in erythroleukemia progenitor cells. To test this hypothesis, we developed two Epo-independent cell lines, UT-7/CI1 and UT-7/CI2 from the Epo-dependent erythroid progenitor cell line UT-7/Epo. Unlike parental UT-7/Epo cells, UT-7/CI1 and UT-7/CI2 cells survived in the absence and presence of Epo. In colony forming cell assay, UT-7/CI1 and UT-7/CI2 cells formed colonies resembling Colony Forming Unit-Erythroid (CFU-E), which are immature erythroid colonies normally dependent of Epo for survival. Furthermore, heme content was similar among UT-7/Epo, UT-7/CI1, and UT-7/CI2 cells. Epo was not detected in the culture media of UT-7/CI1 and UT-7/CI2 cells at 24, 48, and 72 hours, indicating they do not produce Epo as a means of autocrine signaling. Since UT-7/CI1 and UT-7/CI2 cells do not depend on Epo for survival, the signaling pathways of the signal transducer and activator of transcription (STAT) and mitogen-activated protein kinase (MEK) family, which are typically involved in Epo signaling, were investigated. Western blotting demonstrated an increase in STAT3 phosphorylation in UT-7/CI2 cells as well as an increase in p44/42 and p38 phosphorylation in UT-7/CI1 cells. The Janus Kinase (JAK) inhibitor Ruxolitinib induced apoptosis and Caspase3 cleavage in UT-7/Epo and UT-7/CI2 cells, but not UT-7/CI1 cells. However, The MEK1/2 inhibitor U0126 induced apoptosis and Caspase3 cleavage in UT-7/CI1 cells. Overall, the results from this study indicate that UT-7/CI1 cells are dependent on abnormal MEK1/2 signaling for Epo-independent survival whereas UT-7/CI2 cells are dependent on abnormal JAK signaling.

Advisor

Venu Cheriyath

Subject Categories

Cell and Developmental Biology | Life Sciences

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