Date of Award

1-1-2016

Document Type

Thesis

Degree Name

M.S.

Department

Biological Sciences

First Advisor

Joseph K. Angleson, Ph.D.

Keywords

AKAP150, Anterior Pituitary Cells, FRAP, Gonadotropes, LβT2 cells

Abstract

Cellular communication occurs as a result of changes in signaling pathways. A well-studied signaling pathway is through G protein coupled receptors (GPCRs). In gonadotropes, GPCR stimulation by GnRH leads to the activation of protein kinase A (PKA). Activated PKA can phosphorylate ion channels, potentially causing an influx of calcium, depolarization and secretion of hormones. A scaffolding protein known as AKAP150 anchors PKA near L-type calcium channels. In addition, AKAP150 anchors phosphatases, which provides temporal control during signaling events. It was recently shown that AKAP150 is mobile in neuronal dendrites, providing regulation to where the signaling cascade occurs in the cell. It is not yet known what AKAP150 binds to in gonadotropes or how it translocates. In order to investigate the mobility of this protein in pituitary cells, a variety of methods were used in this study. Time-lapse imaging and permeabilization assays suggested that AKAP150-GFP was mobile in LbetaT2 cells, but more sensitive methods were needed to study temporal and spatial resolution. Quantitative FRAP and FLIP analysis provided strong qualitative evidence that AKAP150-GFP was mobile in LbetaT2 cells and the protein moved laterally along the cell rim. Lastly, comparing rim intensity in LbetaT2 cells immunostained for endogenous AKAP150 and actin indicated the scaffolding protein could cluster with to actin.

Copyright Statement / License for Reuse

Creative Commons Attribution 4.0 License
This work is licensed under a Creative Commons Attribution 4.0 License.

Provenance

Received from ProQuest

Rights holder

Kristen E. Dew

File size

83 p.

File format

application/pdf

Language

en

Discipline

Cellular Biology, Molecular Biology

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