The insulin receptor is a receptor tyrosine kinase that engages the PI3 kinase pathway to regulate a FOXO transcription factor. A student uses qRT-PCR to determine the expression of a direct FOXO target gene (GeneX) in a mammalian cell line under different conditions and makes the following observations.

A. Treating the cells with a PTEN inhibitor increases GeneX expression.
B. A cell line with an AKT (S308A) mutation has increased GeneX expression.
C. Change in GeneX expression due to a ligand-binding defective insulin receptor is partly reversed by a PTEN inhibitor.
D. Phosphorylation of FOXO by PDK1 creates a phosphoserine binding site for 14-3-3 protein, reducing GeneX expression.

Which one of the following options represents all correct statements?

Correct option is C

Statement A: -"Treating the cells with a PTEN inhibitor increases GeneX expression."

• PTEN is a phosphatase that dephosphorylates PIP3, a second messenger that activates PI3K/AKT signaling.

• Inhibiting PTEN leads to increased AKT activity, which, in turn, phosphorylates FOXO and sequesters it in the cytoplasm.

• This reduces FOXO activity, which typically leads to decreased GeneX expression (since FOXO usually activates GeneX transcription).

• Therefore, Statement A is incorrect. Inhibiting PTEN would likely decrease GeneX expression, not increase it.

Statement B: - "A cell line with an AKT (S308A) mutation has increased GeneX expression."

• AKT is a kinase that is activated by PI3K signaling. The S308A mutation likely reduces AKT activation or blocks its function.

• This reduction in AKT activity means less phosphorylation of FOXO, which results in greater FOXO activity.

• FOXO is a transcription factor that increases GeneX expression, so GeneX expression would increase in this mutant.

• Statement B is correct.

Statement C: - "Change in GeneX expression due to a ligand-binding defective insulin receptor is partly reversed by a PTEN inhibitor."

• A ligand-binding defective insulin receptor would reduce PI3K/AKT signaling, leading to increased FOXO activity and increased GeneX expression.

• PTEN inhibition would increase PI3K/AKT signaling, which would reverse the effects of the defective insulin receptor by inhibiting FOXO activity and reducing GeneX expression.

• Therefore, Statement C is correct, as PTEN inhibition compensates for reduced PI3K signaling, leading to a partial reversal of changes in GeneX expression caused by the defective insulin receptor.

Statement D: -"Phosphorylation of FOXO by PDK1 creates a phosphoserine binding site for 14-3-3 protein, reducing GeneX expression."

• PDK1 is involved in activating AKT, which subsequently phosphorylates FOXO at specific residues.

• When FOXO is phosphorylated, it binds 14-3-3 proteins, which sequester FOXO in the cytoplasm and prevent it from activating GeneX expression.

• However, PDK1's role is to activate AKT and does not directly involve FOXO phosphorylation in the described mechanism. The AKT/FOXO pathway mainly governs GeneX expression.

• Statement D is misleading or incorrect because it misrepresents the role of PDK1 and FOXO phosphorylation in this context.