Examination of the Potential for ZO1 and ZEB2 to Mediate the Effects of AJAP1 in Epithelial Cells During Cardiovascular Development

Date of Award


Degree Type


Degree Name

Master of Science in Biomedical Sciences

First Advisor

Cathy Hatcher, PhD

Second Advisor

Ruth Borghaei, PhD

Third Advisor

Heather Montie, PhD


Formation of the epicardium by epithelial cells is impaired in Tbx5-deficient mouse hearts. These embryonic mice have reduced cardiac Ajap1 mRNA expression. Knockdown of endogenous AJAP1 (AJAP1 KD) mRNA expression in epithelial cells increases cell adhesion, reduces cell migration, and alters the transcript profile. Tight junction protein 1 (TJP1) and zinc finger E-box binding homeobox 2 (ZEB2) are two transcripts whose expression is altered by AJAP1 silencing. The goal of this study was to examine the role of AJAP1 in regulating protein expression of ZO1 and ZEB2 in epithelial cells. Based on our previous data, we hypothesize that ZO1 and ZEB2 mediate the effects of AJAP1 in epithelial cell behaviors required for epicardium development.

Methods: We used human mammary epithelial cells (HMEpiCs) as an in vitro model to examine the effects of AJAP1 KD on epithelial cell biology. We compared the expression of ZO1 and ZEB2 proteins in AJAP1 KD versus negative control HMEpiCs. Also, we examined ZO1 protein localization within untransfected HMEpiCs and embryonic mouse hearts via immunofluorescence.

Results: Knockdown of AJAP1 mRNA expression in HMEpiCs increased ZO1 protein expression but did not change ZEB2 protein expression. Immunofluorescence studies revealed ZO1 localized to the plasma membrane and appeared diffusely in the cytoplasm of HMEpiCs. ZO1 also appeared in the epicardial cells of embryonic mouse hearts.

Conclusion: Our lab showed that AJAP1 knockdown altered epithelial cell function and now we demonstrate it increases ZO1 protein expression. We also show ZO1 expression in the embryonic mouse epicardium. We hypothesize that ZO1 mediates the effects of AJAP1 in epithelial cell behaviors required for epicardium development.

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