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Updated: May 5, 2026
Peptide-based Identification of Functional Motifs and their Binding Partners
Published on: June 30, 2013
Exploring the feasibility of protein phosphatase 1-docking motif-mimetic cell-penetrating peptides for modulating
Renato M Rodrigues1, Juliana Felgueiras1,2, Sarah Jones3
1Fardilha's Lab, Department of Medical Sciences, Institute of Biomedicine-iBiMED, University of Aveiro, Aveiro, Portugal.
Background:
Once considered "undruggable," protein phosphatases are now recognized as potential therapeutic targets. The serine and threonine-protein phosphatase 1 regulates key cellular processes and enhances androgen receptor activity in prostate cancer, even under castration-resistant conditions, suggesting a role in disease progression.
Methods:
LNCaP and PC3 cells were treated with peptides mimicking protein phosphatase 1-docking motifs in androgen receptor, alongside known bioportides (MSS1 and mitoparan). Cellular uptake was assessed by confocal microscopy and fluorescence assays. Viability was measured with PrestoBlue, and androgen receptor and Prostate-Specific Antigen expression was analyzed by quantitative reverse transcription-polymerase chain reaction and Western blot.
Results:
Androgen receptor sequence contains 3 protein phosphatase 1-docking motifs: KVFF (binding site 1), HVVKW (binding site 2), and KPIYF (binding site 3). Binding site 1 and binding site 2 peptides were modified for better solubility, while binding site 3 was combined with the Tat sequence to enhance cellular uptake. Fluorophore-conjugated peptides successfully entered cells, with androgen receptor-binding site 3 showing the highest internalization in LNCaP cells (P = .0495). Treatment with the 3 androgen receptor-binding site peptides individually reduced cell viability in LNCaP and PC3 cells (P = .0352 and P = .0298, respectively). Combining androgen receptor-binding site peptides statistically reduced cell viability, particularly with all 3 peptides together (LNCaP: 68%, P = .0369; PC3: 80%, P = .0369). No statistically significant changes in androgen receptor or prostate-specific antigen expression were observed.
Conclusion:
Bioportides targeting protein phosphatase 1-docking motifs, especially when combined, decrease prostate cancer cell viability, and additional protein phosphatase 1-interfering peptides such as MSS1 and mitoparan display potent cytotoxic effects. The absence of changes in androgen receptor and prostate-specific antigen expression highlights the need to further investigate their mechanisms of action.
Insights
Novel bioportides targeting protein phosphatase 1 (PP1) docking motifs in androgen receptor significantly reduced prostate cancer cell viability. Combined peptide treatment showed potent cytotoxic effects, suggesting a new therapeutic strategy for prostate cancer.
Area of Science:
- Biochemistry
- Molecular Biology
- Oncology
Background:
- Protein phosphatases, once deemed undruggable, are emerging as critical therapeutic targets.
- Serine and threonine-protein phosphatase 1 (PP1) plays a key role in cellular processes and enhances androgen receptor (AR) activity in prostate cancer, even under castration-resistant conditions.
Purpose of the Study:
- To investigate the therapeutic potential of targeting protein phosphatase 1-docking motifs on the androgen receptor in prostate cancer.
- To evaluate the efficacy of novel bioportides designed to inhibit PP1 activity in prostate cancer cell lines.
Main Methods:
- LNCaP and PC3 prostate cancer cells were treated with peptides mimicking AR-PP1 docking motifs and known bioportides (MSS1, mitoparan).
- Cellular uptake was confirmed via confocal microscopy and fluorescence assays.
- Cell viability was assessed using PrestoBlue, and AR and Prostate-Specific Antigen (PSA) expression levels were analyzed using qRT-PCR and Western blot.
Main Results:
- The AR sequence contains three PP1-docking motifs (KVFF, HVVKW, KPIYF).
- Modified peptides, particularly the AR-binding site 3 peptide combined with Tat sequence, demonstrated enhanced cellular uptake.
- Individual and combined peptide treatments significantly reduced prostate cancer cell viability in both LNCaP and PC3 cell lines, with the combination of all three peptides showing the most potent effect (68% viability in LNCaP, 80% in PC3).
- No significant changes in AR or PSA expression were observed post-treatment.
Conclusions:
- Bioportides targeting PP1-docking motifs on AR, especially when used in combination, effectively decrease prostate cancer cell viability.
- Additional PP1-interfering peptides like MSS1 and mitoparan exhibit significant cytotoxic effects.
- The lack of change in AR and PSA expression warrants further investigation into the precise mechanisms of action for these novel therapeutic agents.
