Cells often use the same signaling systems for different inputs and purposes. Scaffolds come to the rescue.
Eukaryotic cells are huge, at least compared with their progenitors, bacteria. Thanks to their mitochondria and other organizational apparatus, the typical eukaryotic cell has about 100,000 times the volume of a bacterium. These cells are virtual cities- partially organized by their membrane organelles, but there is a much more going on, with tremendous complexity to manage. One issue that was puzzling over the decades was how signals were kept straight. Eukaryotic cells use a variety of signaling systems, proto-typicaly starting with a receptor at the cell surface, linking to a kinase (or series of kinases) that then amplifies and broadcasts the signal inside the cell, ending up with the target phosphorylated proteins entering the nucleus and changing the transcription program of the cell.
While our genome does have roughly 500 kinases, and one to two thousand receptors, a few of them (especially some kinases and their partners, which form "intracellular signaling systems") tend to crop up frequently in different systems and cell types, like the MAP kinase cascade, associated with growth and stress responses, and the AKT kinase, associated with nutrient sensing and growth responses. Not only do many different receptors turn these cellular signaling hubs on, but their effects can often be different as well, even from unrelated signals hitting the same cell.
If all these proteins were diffusable all over the cell, such specificity of signaling would be impossible. But it turns out that they are usually tethered in particular ways, by organizational helpers called scaffold proteins. These scaffolds may localize the signaling to some small volume within the larger cell, such as a membrane "raft" domain. They may also bind multiple actors of the same signaling cascade, bringing several proteins (kinases and targets) together to make signaling both efficient and (sterically) insulated from outside interference. And, in a recent paper, they can also tweak their binding targets allosterically to insulate them from outside interference.
What is allostery vs stery? If one protein (A) binds another (B) such that a phosphorylation or other site is physically hidden from other proteins, such as a kinase (C) that would activate it, that site is said to be sterically hidden- that is, by geometry alone. On the other hand, if that site remains free and accessible, but the binding of A re-arranges protein B such that it no longer binds C very well, blocking the kinase event despite the site of phosphorylation being available, then A has allosterically regulated B. It has altered the shape of B in some subtle way that alters its behavior. While steric effects are dominant and occur everywhere in protein interactions and regulation, allostery comes up pretty frequently as well, proteins being very flexible gymnasts.
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| GSK3 is part of insulin signaling. It is turned off by phosphorylation, which affects a large number of downstream functions, such as turning on glycogen synthase. |
The current case turns on the kinase GSK3, which, according to Wikipedia... "has been identified as a protein kinase for over 100 different proteins in a variety of different pathways. ... GSK-3 has been the subject of much research since it has been implicated in ... diseases, including type 2 diabetes, Alzheimer's disease, inflammation, cancer, addiction and bipolar disorder." GSK3 was named for its kinase activity targeting glycogen synthase, which inactivates the synthase, thus shutting down production of glycogen, which is a way to store sugar for later use. Connected with this homeostatic role, the hormone insulin turns GSK3 off by phosphorylation by a pathway downstream of the membrane-resident insulin receptor called the PI3 kinase / protein kinase B pathway. Insulin thus indirectly increases glycogen synthesis, mopping up excess blood sugar. The circuit reads: insulin --> kinases --| GSK3 --| glycogen synthase --> more glycogen.
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| GSK3 also functions in this totally different pathway, downstream of WNT and Frizzled. Here, GSK3 phosphorylates beta-catenin and turns it off, most of the time. WNT (like insulin) turns GSK3 off, which allows beta-catenin to accumulate and do its gene regulation in the nucleus. Cross-talk between these pathways would be very inopportune, and is prevented by the various functions of Axin, a scaffold protein. |
Another well-studied role of GSK3 is in a developmental signal, called WNT, which promotes developmental decisions of cells during embryogenisis, wound repair, and cancer, cell migration, proliferation, etc. GSK3 is central here for the phosphorylation of beta-catenin, which is a transcription regulator, among other things, and when active migrates to the nucleus to turn its target genes on. But when phosphorylated, beta-catenin is diverted to the proteosome and destroyed, instead. This is the usual state of affairs, with WNT inactive, GSK3 active, and beta-catenin getting constantly made and then immediately disposed of. This complex is called a "destruction" complex. But an incoming WNT signal, typically from neighboring cells carrying out some developmental program, alters the activity of a key scaffold in this pathway, Axin, which is destroyed and replaced by Dishevelled, which turns GSK3 off.
How is GSK3 kept on all the time for the developmental purposes of the WNT pathway, while allowing cells to still be responsive to insulin and other signals that also use GSK3 for their intracellular transmission? The current authors found that the Axin scaffold has a special property of allosterically preventing the phosphorylation of its bound GSK3 by other upstream signaling systems. They even re-engineered Axin to an extremely minimal 26 amino acid portion that binds GSK3, and this still performed the inhibition, showing that the binding doesn't sterically block phosphorylation by insulin signals, but blocks allosterically.
That is great, but what about the downstream connections? Keeping GSK3 on is great, but doesn't that make a mess of the other pathways it participates in? This is where scaffolds have a second job, which is to bring upstream and downstream components together, to keep the whole signal flow isolated. Axin also binds beta-catenin, the GSK3 substrate in WNT signaling, keeping everything segregated and straight.
Scaffold proteins may not "do" anything, as enzymes or signaling proteins in their own right, but they have critical functions as "conveners" of specific, channeled communication pathways, and allow the re-use of powerful signaling modules, over evolutionary time, in new circuits and functions, even in the same cells.
