Ras & the MAP Kinase Pathway

Ras: a monomeric (one subunit) G-protein that stays at the plasma membrane.

MAP: Mitogen Activated Proteins (MAPs), many different protein that are activated by phosphorylation and can promote cell division.

Ras-GTP binds MAP-K-K-K to activate it. MAP-K-K-K, MAP-K-K, and MAP-K are different kinases. Multiple steps amplify the signal: 1 Ras, many MAPs.

?

?

?

?

?

?

?

?

?

?

?

?

?

?

?

[3] G-Protein-Coupled Receptors (GPCP)

G-protein coupled Receptors are 7-pass TM Domain Proteins. They have the extracellular domain, which binds ligands; and the intracellular domain, which associates with trimeric G proteins that have three subunits: α, β, and γ.

?

Trimeric G proteins:

α and γ are lipid linked proteins, so Trimeric G proteins always stay at the plasma membrane, they can diffuse around and interact with others within the plasma membrane; α can bind GTP and sequester the βγ complex - making it unable to bind other things.

?

Trimeric G-protein Activities

In the absence of signals, the α subunit has a GDP bound and the G protein is inactive. In some cases, the inactive G protein is associated with the inactive receptor, while in other cases, the G protein only binds after the receptor is activated.

The ligand-bound activated receptor induces the conformational change that triggers the GDP to be released by the α subunit. Then GTP, abundant in the cytosol, can readily bind in place of the GDP. GTP binding causes conformational changes in the G-protein, activates both the α subunit and the βγ complex. In some cases, as shown in the right picture, the activated α subunit disassociates from the activated βγ complex while two activated components may stay together. Both components can regulate activities of target proteins in the plasma membrane.

Two forms: GDP-αβγ, inactive, α & βγ complex stay together;

GTP-α ?+ ?βγ complex: both active as targets.

GDP-αβγ is activated by a GEF (in this case GEF is the activated ligand-bound GPCR) to become GTP-α + βγ complex. Different trimeric G-proteins can activate different things at the plasma membrane, such as ion channels (left below), adenylyl cyclase (middle below), or phospholipase C (right below), etc.

All increase the concentration of a 2nd?messenger (ion, cAMP, IP3...) in the cell.

?

?

?

?

?

?

Eventually, the activated α subunit hydrolyzes the GTP to GDP, which inactivates the subunit. This step is often accelerated by the binding of GAP (GTPase-accelerating). The inactive GDP-bound α subunit now reforms an inactive G protein with a βγ complex, turning off other downstream events.

Ligand-bound?GPCR acts catalytically, GPCR can activate more than one downstream trimeric G proteins. As long as the signaling receptor remain stimulated, it can continue to activate G proteins.

?

?

?

?

?

?

?

?

?

?

?

?

?

?

?

?

?

?

?

?

?

?

?

?

?

?

However, upon prolong stimulation, the receptor eventually inactivates, even an activating ligand remains bound. In this case, a G-Protein-Coupled Receptor Kinase (GRK) phosphorylates the cytosol portion of the activated receptor. Once the receptor is phosphorylated, it binds an Arrestin (Arr) protein that inactivates the receptor by preventing its interaction with G proteins, steps are shown in the right pictures.

Arrestin proteins also act as adaptor proteins that can recruit the phosphorylated receptor to clathrin-coated pits (recall p. 112-113) from where the receptors are endocytosed - then be degraded in lysosomes or activate new signaling pathways.