Intracellular targets of RGDS peptide in melanoma cells.

Abstract

BACKGROUND:

RGD-motif acts as a specific integrins-ligand and regulates a variety of cell-functions via extracellular action affecting cell-adhesion properties. However, increasing evidence identifies additional RGDS-functions at intracellular level. Previous reports show RGDS-internalization in endothelial cells, cardiomyocytes and lymphocytes, indicating intracellular targets such as caspase-8 and caspase-9, and suggest RGDS specific activity at cytoplasmic level. Given the role RGDS-peptides play in controlling proliferation and apoptosis in several cell types, investigating intracellular targets of RGDS in melanoma cells may un-reveal novel molecular targets and key pathways, potentially useful for a more effective approach to melanoma treatment.

RESULTS:

In the present study we show for the first time that RGDS-peptide is internalized in melanoma cells in a time-dependent way and exerts strong anti-proliferative and pro-apoptotic effects independently from its extracellular anti-adhesive action. RGES control-peptide did not show biological effects, as expected; nevertheless it is internalized, although with slower kinetics. Survivin, a known cell-cycle and survival-regulator is highly expressed in melanoma cells. Co-immunoprecipitation assays in cell lysates and overlay assays with the purified proteins showed that RGDS interacts with survivin, as well as with procaspase-3, -8 and -9. RGDS-peptide binding to survivin was found to be specific, at high affinity (Kd 27.5 muM) and located at the survivin C-terminus. RGDS-survivin interaction appeared to play a key role, since RGDS lost its anti-mitogenic effect in survivin-deprived cells with a specific siRNA.

CONCLUSIONS:

RGDS inhibits melanoma growth with an adhesion-independent mechanism; it is internalized in melanoma cells and specifically interacts with survivin. The present data may indicate a novel role of RGDS-containing peptides physiologically released from the extracellular matrix and may suggest a possible novel anti-proliferation strategy in melanoma.

• Authors: Maria Simona Aguzzi, Paola Fortugno, Claudia Giampietri, Gianluca Ragone, Maurizio C Capogrossi, Antonio Facchiano
• Journal: Molecular cancer (Mol Cancer) Vol. 9 Pg. 84 (Apr 22 2010) ISSN: 1476-4598 [Electronic] England
• PMID: 20412563 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• BIRC5 protein, human
• Inhibitor of Apoptosis Proteins
• Microtubule-Associated Proteins
• Oligopeptides
• RNA, Small Interfering
• Survivin
• arginyl-glycyl-aspartyl-serine
• Caspase 3
• Caspase 8
• Caspase 9

Topics

• Apoptosis (physiology)
• Blotting, Western
• Caspase 3 (metabolism)
• Caspase 8 (metabolism)
• Caspase 9 (metabolism)
• Cell Line, Tumor
• Cell Proliferation
• Cell Separation
• Enzyme Activation (physiology)
• Flow Cytometry
• Humans
• Immunoprecipitation
• Inhibitor of Apoptosis Proteins
• Melanoma (metabolism)
• Microscopy, Confocal
• Microtubule-Associated Proteins (metabolism)
• Oligopeptides (metabolism)
• RNA Interference
• RNA, Small Interfering
• Survivin