Endocytosis is the process in which a cell engulfs extracellular cargo by creating invaginations within its plasma membrane. The cargo that has entered the cell enters an endosome and then is delivered to either the trans-Golgi network for recycling to the plasma membrane or to the lysosome for its degradation (Trousdale & Kim, 2015). Retromer is a peripheral membrane protein complex that plays a key role in sorting of these cargo molecules (Collins, 2008). More specifically, retromer deliver cargo from the endosome to the trans-Golgi Network, the process which is called retrograde transport of cargo molecules.
Retromer dysfunction is strongly linked to neurodegenerative diseases such as Alzheimer’s and Parkinson’s Disease. However, recent Genome Wide Association Studies suggest that a mutation in retromer subunit VPS26a, has been linked to Type II Diabetes (Trousdale & Kim, 2015). A 2016 study published in The Faseb Journal attempts to characterize the role of retromer in adipocyte differentiation and insulin-stimulated uptake of glucose through transporter GLUT4 (Yang et al., 2016). The aim of this study is to further investigate the role of retromer in adipogenesis and to determine whether retromer plays a role at the transcriptional level or translational level.
In this study, retromer’s VPS35 subunit was knocked down in four mouse 3T3-L1 fibroblast cell lines using the CRISPR-Cas9 approach. These cell lines were differentiated into mature adipocytes and analyzed by Oil-Red O staining, Western Blotting and quantitative PCR.
The knockdown of retromer produced varying effects on adipocyte differentiation. In two of the knockdown cell lines, adipocyte differentiation was downregulated whereas adipocyte differentiation was upregulated in the other two cell lines. Although the results from Oil-Red O staining and Western Blot analyses complemented each other, results obtained from qPCR were not as straightforward and further analysis is needed to fully comprehend how retromer acts at the transcriptional level of cell differentiation.
Based on the results of this study, retromer is involved in adipogenesis at both the transcriptional and translational level, however it’s mechanism of action remains unclear as both cases of impaired differentiation and upregulated differentiation were observed. Further studies are necessary to determine retromer’s exact role in adipogenesis.
| Identifer | oai:union.ndltd.org:bu.edu/oai:open.bu.edu:2144/23747 |
| Date | 12 July 2017 |
| Creators | Chaudhry, Hira |
| Source Sets | Boston University |
| Language | en_US |
| Detected Language | English |
| Type | Thesis/Dissertation |
| Rights | Attribution 4.0 International, http://creativecommons.org/licenses/by/4.0/ |
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