Master of Science
The processes of cellular differentiation and apoptosis are critical for placental development. The galectin-16 gene (LGALS16) is associated with these processes in the placenta, but the mechanism by which it acts is poorly characterized. My bioinformatics analysis revealed that LGALS16 expression was limited to specific tissues including the placenta and identified multiple regulatory molecules (transcription factors and miRNAs), which could potentially control relevant transcriptional and post-transcriptional mechanisms. Considering the predominant expression of LGALS16 in placenta, I examined the expression, regulation, and function of LGALS16 in BeWo and JEG-3 cell lines representing in vitro models of trophoblast differentiation. In both models, LGALS16 was significantly upregulated in parallel with human chorionic gonadotropin beta (CGB) during trophoblastic differentiation induced by 8-Br-cAMP. Inhibition of p38 MAPK and Epac significantly altered LGALS16 expression during differentiation, while modulation of O-GlcNAc homeostasis failed to change LGALS16 and CGB expression. Lastly, CRISPR/Cas9 LGALS16 knockouts suppressed expression of CGB. These findings suggest that LGALS16 may play a role in trophoblastic differentiation through p38 MAPK and Epac signaling pathways.
Summary for Lay Audience
Galectins are a group of carbohydrate binding proteins with roles in cell growth, differentiation, and cell death. Of the galectins characterized in humans, galectins-13, -14, and -16 are highly expressed in the placenta, suggesting that they play a unique role in placental development. Furthermore, improper expression of these three galectins leads to disorders such as preeclampsia, a condition characterized by high blood pressure in pregnant mothers that can be fatal. Limited literature exists on galectin-16 (encoded by the LGALS16 gene) and thus it is the focus of the current study. Datasets from other studies were extracted and analyzed to determine the level of galectin-16 in different tissues. This revealed that galectin-16 expression is expressed at a high level in two tissues, the placenta and brain. I also used a program to align the upstream sequence of galectin-16 with sequences of binding sites for specific transcription factors. I determined predicted transcription factors by identifying binding site sequences which matched with 100% similarity to a part of the upstream sequence. Using placental culture models, I induced placental differentiation with cAMP and examined the effects on LGALS16 expression. Additionally, I treated cells with biochemical inhibitors and stimulators of transcription factors, cAMP signaling molecules, and enzymes responsible for adding and removing sugars to proteins. I analyzed how these inhibitors/stimulators affected galectin gene and protein expression, particularly LGALS16, and the biomarker of differentiation, chorionic gonadotropin. Finally, cell pools were genetically modified to knockout galectin-16 and then treated with cAMP to test whether cells could still undergo differentiation. In placental cell models, LGALS16 expression was significantly increased during placental differentiation. Inhibition of two cAMP signaling molecules led to significant changes in LGALS16 expression while no changes were observed in response to drugs changing glycosylation of intracellular proteins. Knocking out galectin-16 in placental cells changed the morphology of cells and inhibited gene expression of chorionic gonadotropin. These findings provide insights into the significance of galectin-16 as a critical regulator of placental differentiation which may serve as a tool to identify dysregulated placental differentiation in cancers and pregnancy disorders.
Kaminker, Jennifer, "Characterization of galectin-16 expression and function in placental cells" (2022). Electronic Thesis and Dissertation Repository. 8628.