Ciliary and Non-Ciliary Protein Kinase A (PKA) Differentially Coordinate 3T3-L1 Adipogenesis
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Authors
Delios, Samantha
Date
2024-09-06
Type
thesis
Language
eng
Keyword
Adipogenesis , Primary Cilium , Adipose Tissue , Obesity
Alternative Title
Abstract
The primary cilium (PC) is a compact organelle that regulates cellular processes in virtually all cell types. In this context, research attests that adipocyte progenitor cell (APC) adipogenesis is regulated by PC-specific signaling systems (i.e. cAMP signaling). Recent work identified a role for ciliary expression of free fatty acid receptor 4 (FFAR4), and its ability to increase ciliary cAMP levels, in potentiating pro-adipogenic actions of low concentrations of insulin and dexamethasone in APCs, including those isolated from mouse or human adipose tissues (ATs) and the preadipocyte cell line (i.e. 3T3-L1 cells). We propose that promoting APC hyperplasia, via this PC-specific cAMP system, could reduce AT hypoxia and inflammation in obesity or type 2 diabetes (T2D), therefore, alleviating associated vascular dysfunctions.
While earlier work suggested that FFAR4 promoted adipogenesis by stimulating extra-ciliary exchange protein activated by cAMP (EPAC) activity, in contrast, our work hypothesizes that FFAR4-regulated ciliary cAMP promotes adipogenesis by selectively activating ciliary protein kinase A (PKA). Thus, we investigated the role of ciliary PKA activity in promoting FFAR4-dependent adipogenesis in 3T3-L1 preadipocytes, a cell model of APC adipogenesis. Specifically, we determined the impact of expressing a ciliary targeted construct [NPHP3(1-203)-YFP-PKI] that allowed ciliary expression of PKA inhibitory peptide (PKI) on PC, and non-PC-dependent, 3T3-L1 adipogenesis. As a control, a construct showcasing the inactive version of PKI [NPHP3-YFP-PKI(4A)] was expressed in these cells. Adipogenic responses were determined through PPARγ nuclear staining. Our findings are consistent with the idea that specific inhibition of ciliary PKA (i.e. PKI) significantly (p ≤ 0.0001) antagonized FFAR4-mediated pro-adipogenic responses in 3T3-L1 cells (i.e. PPARγ staining was notably absent). In marked contrast, expression of the inactive PKI variant did not alter the pro-adipogenic effects promoted by the FFAR4 agonist TUG-891 in these cells (i.e. PPARγ staining was not altered). Speaking to the specificity of these effects on ciliary FFAR4/PKA signaling, experiments in which adipogenesis was induced using a non-PC-dependent angiogenic cocktail (i.e., high concentrations of insulin, dexamethasone and isobutyl methyl-xanthine (IBMX)) was insensitive to ciliary PKI expression. We believe our work will help identify PC-specific PKA as a pharmacological target in regulating APC adipogenesis.
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Attribution 4.0 International
Queen's University's Thesis/Dissertation Non-Exclusive License for Deposit to QSpace and Library and Archives Canada
ProQuest PhD and Master's Theses International Dissemination Agreement
Intellectual Property Guidelines at Queen's University
Copying and Preserving Your Thesis
This publication is made available by the authority of the copyright owner solely for the purpose of private study and research and may not be copied or reproduced except as permitted by the copyright laws without written authority from the copyright owner.
Queen's University's Thesis/Dissertation Non-Exclusive License for Deposit to QSpace and Library and Archives Canada
ProQuest PhD and Master's Theses International Dissemination Agreement
Intellectual Property Guidelines at Queen's University
Copying and Preserving Your Thesis
This publication is made available by the authority of the copyright owner solely for the purpose of private study and research and may not be copied or reproduced except as permitted by the copyright laws without written authority from the copyright owner.