You have isolated an epithelial cell line which is unrespons

You have isolated an epithelial cell line which is unresponsive to EGF (normal epithelial derived cell lines require EGF in the culture media for growth). You suspect that EGF receptor signaling in this cell line is impaired, and you designate this a mutant cell line. As an assay for EGFR signaling, you have stably transfected this mutant cell line and a normal cell line with a luciferase-based reporter construct that is responsive to the transcription factor Elk-1. You want to determine what component of the signaling pathway is impaired.

The luciferase activity is measured in both the mutant cell line and the normal cell line following treatment with (+) or without(-) EGF in the culture media.

Briefly state what the results tell you and indicate whether the impaired component in the mutant cell line lies upstream or downstream in the EGF signaling cascade, or if the component indicated is in fact the impaired component.

Luciferase Activity A.U. +10%) A.U. arbitrary units EGF EGF Mutant 9 10 Norma 8 182

Solution

Answer:

The result shows that:

In normal cell line, on treatment of EGF there is more luciferase activity than untreated cell line. As luciferase-based reporter construct is responsive to the transcription factor Elk-1 and Elk-1 is related to EGFR signaling transcription factor, so in normal cell line there is efficient EGFR signaling occurs.

In mutant cell line, the presence or absence of EGF does not significantly affect the luciferase activity. That means the EGF is not coming to the signaling pathway. So there is problem in the upstream of the EGF signaling cascade. The impaired component may be EGFR (EGF Receptor).