Cross-talks via mTORC2 can explain enhanced activation in response to insulin in diabetic patients

The molecular mechanisms of insulin resistance in Type 2 diabetes have been extensively studied in primary human adipocytes, and mathematical modelling has clarified the central role of attenuation of mammalian target of rapamycin (mTOR) complex 1 (mTORC1) activity in the diabetic state. Attenuation of mTORC1 in diabetes quells insulin-signalling network-wide, except for the mTOR in complex 2 (mTORC2)-catalysed phosphorylation of protein kinase B (PKB) at Ser473 (PKB-S473P), which is increased. This unique increase could potentially be explained by feedback and interbranch cross-talk signals. To examine if such mechanisms operate in adipocytes, we herein analysed data from an unbiased phosphoproteomic screen in 3T3-L1 adipocytes. Using a mathematical modelling approach, we showed that a negative signal from mTORC1-p70 S6 kinase (S6K) to rictor–mTORC2 in combination with a positive signal from PKB to SIN1–mTORC2 are compatible with the experimental data. This combined cross-branch signalling predicted an increased PKB-S473P in response to attenuation of mTORC1 – a distinguishing feature of the insulin resistant state in human adipocytes. This aspect of insulin signalling was then verified for our comprehensive model of insulin signalling in human adipocytes. Introduction of the cross-branch signals was compatible with all data for insulin signalling in human adipocytes, and the resulting model can explain all data network-wide, including the increased PKB-S473P in the diabetic state. Our approach was to first identify potential mechanisms in data from a phosphoproteomic screen in a cell line, and then verify such mechanisms in primary human cells, which demonstrates how an unbiased approach can support a direct knowledge-based study. * IR, : insulin receptor; IRS1, : IR substrate-1; mTOR, : mammalian target of rapamycin; mTORC1, : mTORC complex 1; mTORC2, : mTORC complex 2; PKB, : protein kinase B; PKB-S473P, : phosphorylation of PKB at Ser473; S6K, : p70 S6 kinase; TSC2, : tuberous sclerosis complex 2