The Quaking (qk) gene has been implicated in the development of oligodendroglial cells which are the primary source of myelin in the mammalian central nervous system (CNS). Qk encodes three alternatively spliced variants, QKI-5, QKI-6 and QKI-7, all of which are RNA binding proteins. Loss of QKI-6 and QKI-7 results in a dysmyelination phenotype that is present shortly after birth while loss of QKI-5 results in embryonic lethality. CG4 oligodendroglial cells were transfected with either pIRES2-QKI5 to up regulate QKI-5 expression or a QKI-5 specific siRNA to down regulate QKI-5. Cells were cultured for 6d in differentiation medium (DM) following which total RNA and protein was collected from the cell cultures, and coverslips with attached cells were processed for immunofluorescence. Increased QKI-5 expression following transfection with pIRES2-QKI5 resulted in increased Sirt2 and Plp mRNA expression, but did not affect SIRT2 and PLP protein expression. Down regulation of QKI-5 expression had no significant effect on mRNA or protein levels for QKI-6, QKI-7, Plp or Sirt2. Immunocytochemistry revealed that up regulation of QKI-5 resulted in significantly higher percentage of A2B5+ cells and a lower percentage of GalC+ cells, whereas siRNA treatment resulted in an increase in the percentage of GalC+ cells. Our results suggest QKI-5 regulates oligodendrocyte differentiation and modulates the transcription and availability of target mRNAs, such as Sirt2 and Plp, for translation. In order to gain a more complete understanding of the relationship between qk and both Sirt2 and Plp, future studies would include RNA coimmunoprecipitation, miRNA studies, and expanding the list of target genes to include various cell cycle components.
| Identifer | oai:union.ndltd.org:USASK/oai:ecommons.usask.ca:10388/ETD-2013-09-1205 |
| Date | 2013 September 1900 |
| Contributors | Nazarali, Adil J., Doucette, Ronald |
| Source Sets | University of Saskatchewan Library |
| Language | English |
| Detected Language | English |
| Type | text, thesis |
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