Graduation Date

Fall 12-16-2022

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Programs

Cancer Research

First Advisor

Jennifer Black

Abstract

Protein kinase Cα (PKCα) is a member of the PKC family of serine/threonine kinases, which have been implicated in regulation of many cellular processes, including cell proliferation, differentiation, survival, and transformation. A large body of evidence from the Black laboratory and others support an anti-proliferative function of PKCα in normal epithelial tissues, including the intestinal mucosa and endometrial epithelium. PKCα is also tumor suppressive in epithelial cancers, such as colorectal cancer (CRC) and endometrial cancer (EC). However, a major obstacle to harnessing the tumor suppressive functions of PKCα to benefit patients is the widespread loss of PKCα expression in tumors. Thus, the overall goal of this study was to identify downstream effectors mediating the growth suppressive function of PKCα and understand the mechanism(s) underlying PKCα loss in epithelial cancers. Our work has broadened our knowledge of the effector signaling cascade activated by PKCα and fills the gap in understanding of the mechanisms mediating loss of PKCα expression in cancer.

Our studies have revealed crosstalk between PKCα signaling and the TGFβ pathway, a major antiproliferative and tumor suppressive signaling cascade in many tissue types. PKCα activation induces the expression of TGFβR1 mRNA and protein by triggering a RAS-RAF-MEK-ERK-Runx2 signaling axis. While PKCα induces Runx2 gene expression, ERK-dependent activating phosphorylation of Runx2 plays a major role in enhancing TGFβR1 transcription. Functional studies have revealed that disruption of PKCα reduces the sensitivity of intestinal cells to TGFβ signaling, indicated by abrogation of TGFβ-induced SMAD2 phosphorylation and G1 to S phase cell cycle arrest. PKCα-induced upregulation of p21 and p27 was also affected by inhibition of TGFβR1 activity. These findings are supported by a strong correlation between PKCα and TGFβR1 expression in CRC and EC tumors.

Analysis of the methylomes of endometrial tumors revealed de novo hypermethylation on the right CpG island (CGI) shore of the PRKCA gene compared with normal endometrial epithelium. A strong correlation between right CGI shore methylation and PKCα mRNA expression in CRC and EC cancer cell lines provided further support for the role of epigenetic modification in regulating PKCα expression in tumors. These findings, combined with the demonstration that the hypomethylating agent, decitabine, restored PKCα expression in CRC and EC cell lines and CRC patient-derived organoids, point to right shore methylation as an important mechanism mediating PKCα loss in colon and endometrial cancers.

In conclusion, this dissertation reports novel crosstalk between PKCα and TGFβ signaling in regulation of intestinal epithelial homeostasis and de novo hypermethylation on the right CGI shore of the PRKCA gene, which likely accounts for suppression of PKCα expression in many tumor types.

Comments

2022 Copyright, the authors

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