Expression of Wnt5a alternative promoters A and B during cancer progression and cellular differentiation

UNCG Author/Contributor (non-UNCG co-authors, if there are any, appear on document)
Chia-Chi Hsu (Creator)
Institution
The University of North Carolina at Greensboro (UNCG )
Web Site: http://library.uncg.edu/
Advisor
Karen Katula

Abstract: WNT5A is a secreted glycoprotein that plays an important role in cellular differentiation, cell homeostasis and development. It is also misregulated in numerous cancer cell types. The Wnt5a gene generates multiple transcripts from distinct promoters and alternative splicing, leading to different protein isoforms. Currently, little is known regarding the regulation of the Wnt5a alternative promoters. The goal of this study was to characterize the expression of Wnt5a alternative promoters A and B during differentiation and cellular transformation. TaqMan primer-probe sets, specific to promoter A and promoter B derived transcripts, were designed and characterized. The level of promoter A and B specific transcripts were determined in normal human osteoblasts and the osteosarcoma cell line, SaOS-2, as a model for cancer progression. The level of promoter A and B transcripts were nearly equal in osteoblasts cells. In contrast, there was a dramatic decrease in promoter B transcripts in osteosarcoma cells and an increase of 3.5 fold in promoter A transcripts, giving an A to B ratio of 2320 to 1. Wnt5a promoter A and promoter B luciferase reporter constructs were transfected into osteosarcoma cells. Promoter A and promoter B activities were found to be nearly equal, suggesting that the lower level of promoter B transcripts in osteosarcoma cells is not due to altered levels of transcription factors. Promoter A and promoter B specific transcripts were assayed in 3T3-L1 mouse fibroblasts, as a model for differentiation, in the following stages: exponential preadipocytes (EX PA), confluent preadipocytes (CON PA), two days after MDI treatment (D2 Post MDI) and in mature differentiated adipocytes (Diff AD). In EX PA and CON PA cells, both promoter A and promoter B transcripts increased followed by decreased transcript levels in D2 Post MDI cells. While promoter A transcripts slightly increased in Diff AD, promoter B transcript levels remained at a low level. Overall, these results suggest that Wnt5a promoter A and promoter B are differentially regulated.

Additional Information

Publication
Thesis
Language: English
Date: 2012
Keywords
WNT5A, Glycoprotein, Cells, Alternative promoter A, Alternative promoter B
Subjects
Glycoproteins $x Therapeutic use $x Research