PEGylation of lysine residues reduces the pro-migratory activity of IGF-I

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• 作者：Manaswini Sivaramakrishnan ; Abhishek S. Kashyap ; Beat Amrein ; Stefanie Saenger ; Sonja Meier ; Christian Staudenmaier ; Zee Upton ; Friedrich Metzger
• 关键词：Insulin-like growth factor 1 ; IGF-I ; PEG-IGF-I ; Migration ; Viability
• 刊名：Biochimica et Biophysica Acta (BBA)/General Subjects
• 出版年：2013
• 出版时间：October, 2013
• 年：2013
• 卷：1830
• 期：10
• 页码：4734-4742
• 全文大小：620 K

文摘

Background

The insulin-like growth factor (IGF) system is composed of ligands and receptors which regulate cell proliferation, survival, differentiation and migration. Some of these functions involve regulation by the extracellular milieu, including binding proteins and other extracellular matrix proteins. However, the functions and exact nature of these interactions remain incomplete.

Methods

IGF-I variants PEGylated at lysines K27, K65 and K68, were assessed for binding to IGFBPs using BIAcore, and for phosphorylation of the IGF-IR. Furthermore, functional consequences of PEGylation were investigated using cell viability and migration assays. In addition, downstream signaling pathways were analyzed using phospho-AKT and phospho-ERK1/2 assays.

Results

IGF-I PEGylated at lysines 27 (PEG-K27), 65 (PEG-K65) or 68 (PEG-K68) was employed. Receptor phosphorylation was similarly reduced 2-fold with PEG-K65 and PEG-K68 in 3T3 fibroblasts and MCF-7 breast cancer cells, whereas PEG-K27 showed a more than 10- and 3-fold lower activation for 3T3 and MCF-7 cells, respectively. In addition, all PEG-IGF-I variants had a 10-fold reduced association rate to IGF binding proteins (IGFBPs). Functionally, all PEG variants lost their ability to induce cell migration in the presence of IGFBP-3/vitronectin (VN) complexes, whereas cell viability was fully preserved. Analysis of downstream signaling revealed that AKT was preferentially affected upon treatment with PEG-IGF-I variants whereas MAPK signaling was unaffected by PEGylation.

Conclusion

PEGylation of IGF-I has an impact on cell migration but not on cell viability.

General significance

PEG-IGF-I may differentially modulate IGF-I mediated functions that are dependent on receptor interaction as well as key extracellular proteins such as VN and IGFBPs.