Limits...
The C-terminal region Mesd peptide mimics full-length Mesd and acts as an inhibitor of Wnt/β-catenin signaling in cancer cells.

Lin C, Lu W, Zhang W, Londoño-Joshi AI, Buchsbaum DJ, Bu G, Li Y - PLoS ONE (2013)

Bottom Line: In our previous study, we found that the C-terminal region of Mesd, which is absent in sequences from invertebrates, is necessary and sufficient for binding to mature LRP6 on the cell surface.We found that Mesd C-terminal region-derived peptides block Mesd binding to LRP5 at the cell surface too.Finally, we found that treatment of the full-length Mesd protein and its C-terminal region peptide significantly increased chemotherapy agent Adriamycin-induced cytotoxicity in HS578T and PC-3 cells.

View Article: PubMed Central - PubMed

Affiliation: Drug Discovery Division, Southern Research Institute, Birmingham, Alabama, United States of America.

ABSTRACT
While Mesd was discovered as a specialized molecular endoplasmic reticulum chaperone for the Wnt co-receptors LRP5 and LRP6, recombinant Mesd protein is able to bind to mature LRP5 and LRP6 on the cell surface and acts as a universal antagonist of LRP5/6 modulators. In our previous study, we found that the C-terminal region of Mesd, which is absent in sequences from invertebrates, is necessary and sufficient for binding to mature LRP6 on the cell surface. In the present studies, we further characterized the interaction between the C-terminal region Mesd peptide and LRP5/6. We found that Mesd C-terminal region-derived peptides block Mesd binding to LRP5 at the cell surface too. We also showed that there are two LRP5/6 binding sites within Mesd C-terminal region which contain several positively charged residues. Moreover, we demonstrated that the Mesd C-terminal region peptide, like the full-length Mesd protein, blocked Wnt 3A- and Rspodin1-induced Wnt/β-catenin signaling in LRP5- and LRP6- expressing cells, suppressed Wnt/β-catenin signaling in human breast HS578T cells and prostate cancer PC-3 cells, and inhibited cancer cell proliferation, although the full-length Mesd protein is more potent than its peptide. Finally, we found that treatment of the full-length Mesd protein and its C-terminal region peptide significantly increased chemotherapy agent Adriamycin-induced cytotoxicity in HS578T and PC-3 cells. Together, our results suggest that Mesd C-terminal region constitutes the major LRP5/6-binding domain, and that Mesd protein and its C-terminal region peptide have a potential therapeutic value in cancer.

Show MeSH

Related in: MedlinePlus

Mesd C-terminal region peptides block Mesd binding to LRP6 and LRP5 at the cell surface.(A) Schematic representation of mouse Mesd and its C-terminal region peptides and amino acid sequences of mouse Mesd C-terminal region peptides. (B) 125I-Mesd (5 nM) binding to LRP6-transduced HT1080 cells and the corresponding control cells was carried out for 4 h at 4°C in the absence or presence of 500 nM mouse Mesd or 500 nM mouse Mesd C-terminal region peptide. Values are the average of triple determinations with the s.d. indicated by error bars. (C) 125I-Mesd (5 nM) binding to LRP6-transduced HT1080 cells or LRP5-trasfected ldl-7 cells was carried out for 4 h at 4°C in the absence or presence of various mouse Mesd C-terminal region peptides at the indicated concentrations. Values are the average of triple determinations with the s.d. indicated by error bars. **P<0.01 compared to the LRP6 or LRP5 cells in the absence of Mesd and its peptides.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3585277&req=5

pone-0058102-g001: Mesd C-terminal region peptides block Mesd binding to LRP6 and LRP5 at the cell surface.(A) Schematic representation of mouse Mesd and its C-terminal region peptides and amino acid sequences of mouse Mesd C-terminal region peptides. (B) 125I-Mesd (5 nM) binding to LRP6-transduced HT1080 cells and the corresponding control cells was carried out for 4 h at 4°C in the absence or presence of 500 nM mouse Mesd or 500 nM mouse Mesd C-terminal region peptide. Values are the average of triple determinations with the s.d. indicated by error bars. (C) 125I-Mesd (5 nM) binding to LRP6-transduced HT1080 cells or LRP5-trasfected ldl-7 cells was carried out for 4 h at 4°C in the absence or presence of various mouse Mesd C-terminal region peptides at the indicated concentrations. Values are the average of triple determinations with the s.d. indicated by error bars. **P<0.01 compared to the LRP6 or LRP5 cells in the absence of Mesd and its peptides.

Mentions: In our previous study, we have shown that the Mesd C-terminal region peptide mMesd (150–195) blocks the full-length Mesd protein binding to mature LRP6 at the cell surface [12]. However, whether the Mesd C-terminal region peptide is able to block Mesd protein binding to mature LRP5 at cell surface is unclear. The most part of the C-terminal region of Mesd is absent in sequences from invertebrates. As the first 5 amino acid residues at the N-terminus and the last 4 amino acid residues at the C-terminus of mMesd (150–195) are conserved across different species, we prepared a shorter mouse Mesd C-terminal region peptide, mMesd (155–191), which lacks these 9 amino acid residues (Figure 1A). We then performed 125I-Mesd binding assay with LRP6- and LRP5-expressing cells. We found that mMesd (155–191), like mMesd (150–195), blocked Mesd protein binding to LRP6 as well as LRP5 at the cell surface (Figure 1B), indicating that the Mesd C-terminal region is also important for Mesd binding to LRP5.


The C-terminal region Mesd peptide mimics full-length Mesd and acts as an inhibitor of Wnt/β-catenin signaling in cancer cells.

Lin C, Lu W, Zhang W, Londoño-Joshi AI, Buchsbaum DJ, Bu G, Li Y - PLoS ONE (2013)

Mesd C-terminal region peptides block Mesd binding to LRP6 and LRP5 at the cell surface.(A) Schematic representation of mouse Mesd and its C-terminal region peptides and amino acid sequences of mouse Mesd C-terminal region peptides. (B) 125I-Mesd (5 nM) binding to LRP6-transduced HT1080 cells and the corresponding control cells was carried out for 4 h at 4°C in the absence or presence of 500 nM mouse Mesd or 500 nM mouse Mesd C-terminal region peptide. Values are the average of triple determinations with the s.d. indicated by error bars. (C) 125I-Mesd (5 nM) binding to LRP6-transduced HT1080 cells or LRP5-trasfected ldl-7 cells was carried out for 4 h at 4°C in the absence or presence of various mouse Mesd C-terminal region peptides at the indicated concentrations. Values are the average of triple determinations with the s.d. indicated by error bars. **P<0.01 compared to the LRP6 or LRP5 cells in the absence of Mesd and its peptides.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3585277&req=5

pone-0058102-g001: Mesd C-terminal region peptides block Mesd binding to LRP6 and LRP5 at the cell surface.(A) Schematic representation of mouse Mesd and its C-terminal region peptides and amino acid sequences of mouse Mesd C-terminal region peptides. (B) 125I-Mesd (5 nM) binding to LRP6-transduced HT1080 cells and the corresponding control cells was carried out for 4 h at 4°C in the absence or presence of 500 nM mouse Mesd or 500 nM mouse Mesd C-terminal region peptide. Values are the average of triple determinations with the s.d. indicated by error bars. (C) 125I-Mesd (5 nM) binding to LRP6-transduced HT1080 cells or LRP5-trasfected ldl-7 cells was carried out for 4 h at 4°C in the absence or presence of various mouse Mesd C-terminal region peptides at the indicated concentrations. Values are the average of triple determinations with the s.d. indicated by error bars. **P<0.01 compared to the LRP6 or LRP5 cells in the absence of Mesd and its peptides.
Mentions: In our previous study, we have shown that the Mesd C-terminal region peptide mMesd (150–195) blocks the full-length Mesd protein binding to mature LRP6 at the cell surface [12]. However, whether the Mesd C-terminal region peptide is able to block Mesd protein binding to mature LRP5 at cell surface is unclear. The most part of the C-terminal region of Mesd is absent in sequences from invertebrates. As the first 5 amino acid residues at the N-terminus and the last 4 amino acid residues at the C-terminus of mMesd (150–195) are conserved across different species, we prepared a shorter mouse Mesd C-terminal region peptide, mMesd (155–191), which lacks these 9 amino acid residues (Figure 1A). We then performed 125I-Mesd binding assay with LRP6- and LRP5-expressing cells. We found that mMesd (155–191), like mMesd (150–195), blocked Mesd protein binding to LRP6 as well as LRP5 at the cell surface (Figure 1B), indicating that the Mesd C-terminal region is also important for Mesd binding to LRP5.

Bottom Line: In our previous study, we found that the C-terminal region of Mesd, which is absent in sequences from invertebrates, is necessary and sufficient for binding to mature LRP6 on the cell surface.We found that Mesd C-terminal region-derived peptides block Mesd binding to LRP5 at the cell surface too.Finally, we found that treatment of the full-length Mesd protein and its C-terminal region peptide significantly increased chemotherapy agent Adriamycin-induced cytotoxicity in HS578T and PC-3 cells.

View Article: PubMed Central - PubMed

Affiliation: Drug Discovery Division, Southern Research Institute, Birmingham, Alabama, United States of America.

ABSTRACT
While Mesd was discovered as a specialized molecular endoplasmic reticulum chaperone for the Wnt co-receptors LRP5 and LRP6, recombinant Mesd protein is able to bind to mature LRP5 and LRP6 on the cell surface and acts as a universal antagonist of LRP5/6 modulators. In our previous study, we found that the C-terminal region of Mesd, which is absent in sequences from invertebrates, is necessary and sufficient for binding to mature LRP6 on the cell surface. In the present studies, we further characterized the interaction between the C-terminal region Mesd peptide and LRP5/6. We found that Mesd C-terminal region-derived peptides block Mesd binding to LRP5 at the cell surface too. We also showed that there are two LRP5/6 binding sites within Mesd C-terminal region which contain several positively charged residues. Moreover, we demonstrated that the Mesd C-terminal region peptide, like the full-length Mesd protein, blocked Wnt 3A- and Rspodin1-induced Wnt/β-catenin signaling in LRP5- and LRP6- expressing cells, suppressed Wnt/β-catenin signaling in human breast HS578T cells and prostate cancer PC-3 cells, and inhibited cancer cell proliferation, although the full-length Mesd protein is more potent than its peptide. Finally, we found that treatment of the full-length Mesd protein and its C-terminal region peptide significantly increased chemotherapy agent Adriamycin-induced cytotoxicity in HS578T and PC-3 cells. Together, our results suggest that Mesd C-terminal region constitutes the major LRP5/6-binding domain, and that Mesd protein and its C-terminal region peptide have a potential therapeutic value in cancer.

Show MeSH
Related in: MedlinePlus