Cooperative regulation of the invasive and metastatic phenotypes by different domains of the type I insulin-like growth factor receptor beta subunit.
The receptor for the type 1 insulin-like growth factor (IGF-I) regulates multiple cellular functions impacting on the metastatic phenotype of tumor cells, including cellular proliferation, anchorage-independent growth, survival, migration, synthesis of the 72-kDa type IV collagenase and invasion. We have used site-directed mutagenesis to generate domain-specific mutants of the receptor beta subunit to analyze the role of specific tyrosines in the regulation of the invasive/metastatic phenotype. Poorly invasive M-27 carcinoma cells expressing low receptor numbers were transfected with a plasmid vector expressing IGF-I receptor cDNA in which single or multiple tyrosine codons in the kinase domain, namely Tyr-1131, Tyr-1135, and Tyr-1136 or the C-terminal tyrosines 1250 and 1251 were substituted with phenylalanine. Changes in the invasive and metastatic properties were analyzed relative to M-27 cells expressing the wild type receptor. We found that cells expressing the Y1131F,Y1135F,Y1136F or Y1135F receptor mutants lost all IGF-IR-dependent functions and their phenotypes were indistinguishable from, or suppressed relative to, the parent line. The Y1250F,Y1251F substitution abolished anchorage-independent growth, cell spreading, and the anti-apoptotic effect of IGF-I whereas all other IGF-IR-dependent phenotypes were either unperturbed (i.e. mitogenicity) or only partially reduced (migration and invasion). The results identify three types of receptor-dependent functions in this model: those dependent only on an intact kinase domain (DNA synthesis), those dependent equally on kinase domain and Tyr-1250/1251 signaling (e.g. apoptosis, soft agar cloning) and those dependent on kinase domain and enhanced through Tyr-1250/1251 signaling (migration, invasion). They suggest that signals derived from both regions of the receptor cooperate to enhance tumor metastasis.
Brodt
P
P
Department of Surgery, McGill University Health Center, Royal Victoria Hospital, 687 Pine Ave W., Montreal, Quebec H3A 1A1, Canada. pnina.brodt@muhc.mcgill.ca
Fallavollita
L
L
Khatib
A M
AM
Samani
A A
AA
Zhang
D
D
D002465:Cell Movement; D003001:Cloning, Molecular; D004252:DNA Mutational Analysis; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D015972:Gene Expression Regulation, Neoplastic; D005799:Genes, Dominant; D006801:Humans; D007334:Insulin-Like Growth Factor I / Q000378:metabolism; D007700:Kinetics; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D009361:Neoplasm Invasiveness; D009362:Neoplasm Metastasis; D010641:Phenotype; D017434:Protein Structure, Tertiary; D017526:Receptor, IGF Type 1 / Q000737:chemistry* / Q000378:metabolism; D015398:Signal Transduction; D013997:Time Factors; D014162:Transfection; D014407:Tumor Cells, Cultured; D014443:Tyrosine / Q000737:chemistry
D018076:DNA, Complementary; D014443:Tyrosine; D007334:Insulin-Like Growth Factor I; D017526:Receptor, IGF Type 1; D020778:Matrix Metalloproteinase 2