Using A Proteomics Approach In The Epithelial Cell Line Mdck You Discover A New Protein That Binds Physically To The Ce 1 (585.26 KiB) Viewed 36 times
Using a proteomics approach in the epithelial cell line MDCK, you discover a new protein that binds physically to the cell polarity protein PatJ. You name it Partner Of PatJ -1, POP-1. You characterise POP-1 as a new member of the cell polarity complex that PatJ belongs to. Your initial studies show co-localisation of POP-1 with ZO-1 in MDCK cells in 2D cultures (see figure below). TOP VIEW SIDE VIEW POP-1 ZO-1 I POP-1/ZO-1 MERGE a. What other cell polarity proteins would you expect to find in the POP-1 complex? (2 marks) b. What junctional complex is ZO-1 part of? What other types of junctions are basally localised to Tight Junctions? (2 marks) c. Would you expect POP-1 to inhibit or cooperate with the Scribble complex to regulate Apicobasal cell polarity? Why?
Looking through cancer mutation databases you notice that a point mutation at amino acid 233 (L->P) of POP-1 occurs in more than 30% of human renal cell carcinomas. You use CRISPR editing to introduce the equivalent point mutation in MDCK cells, POP-1 L233P. You compare the localisation of PATJ by immunofluorescence (aPATJ) in POP- 7 wildtype (WT) and POP-1 L233P MDCK cells (see Figure). aPATJ POP-1 WT aPATJ POP-1 L233P d. What effect would you predict the L233P mutation would have on the cell polarity function of PATJ? Why? (2 marks) e. What effect do you think the POP-1 L233P mutation would have on the localisation of the Discs Large (Dlg) proteins in MDCK cells? Why? (2 Marks)
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