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A bipolar clamp mechanism for activation of Jak-family protein tyrosine kinases.

Barua D, Faeder JR, Haugh JM - PLoS Comput. Biol. (2009)

Bottom Line: Building on a rule-based kinetic modeling approach that considers the concerted nature and combinatorial complexity of modular protein domain interactions, we examine these mechanisms in detail, focusing on the growth hormone (GH) receptor/Jak2/SH2-Bbeta system.The modeling results suggest that, whereas Jak2-(SH2-Bbeta)(2)-Jak2 heterotetramers are scarcely expected to affect Jak2 phosphorylation, SH2-Bbeta and dimerized receptors synergistically promote Jak2 trans-activation in the context of intracellular signaling.Analysis of the results revealed a unique mechanism whereby SH2-B and receptor dimers constitute a bipolar 'clamp' that stabilizes the active configuration of two Jak2 molecules in the same macro-complex.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemical and Biomolecular Engineering, North Carolina State University, Raleigh, North Carolina, USA.

ABSTRACT
Most cell surface receptors for growth factors and cytokines dimerize in order to mediate signal transduction. For many such receptors, the Janus kinase (Jak) family of non-receptor protein tyrosine kinases are recruited in pairs and juxtaposed by dimerized receptor complexes in order to activate one another by trans-phosphorylation. An alternative mechanism for Jak trans-phosphorylation has been proposed in which the phosphorylated kinase interacts with the Src homology 2 (SH2) domain of SH2-B, a unique adaptor protein with the capacity to homo-dimerize. Building on a rule-based kinetic modeling approach that considers the concerted nature and combinatorial complexity of modular protein domain interactions, we examine these mechanisms in detail, focusing on the growth hormone (GH) receptor/Jak2/SH2-Bbeta system. The modeling results suggest that, whereas Jak2-(SH2-Bbeta)(2)-Jak2 heterotetramers are scarcely expected to affect Jak2 phosphorylation, SH2-Bbeta and dimerized receptors synergistically promote Jak2 trans-activation in the context of intracellular signaling. Analysis of the results revealed a unique mechanism whereby SH2-B and receptor dimers constitute a bipolar 'clamp' that stabilizes the active configuration of two Jak2 molecules in the same macro-complex.

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SH2-Bβ dimerization coordinates the formation of macro-complexes containing two Jak2 molecules bound to GH-dimerized receptors.Steady-state calculations were performed using the Simplified Cellular Model and the same parameter values as in Figure 3, except with 10 nM GH stimulation and varying SH2-Bβ concentration. (A) Receptor-bound, phosphorylated Jak2 (Y2∼P), for various values of the SH2-Bβ dimerization affinity. The extreme cases of KD,SS equal to zero and infinity correspond to irreversible and no dimerization, respectively; intermediate KD,SS values are 10 nM, 100 nM, 1 µM, and 10 µM. (B) Analysis of receptor/Jak2 complexes, with KD,SS = 100 nM. SH2-Bβ dimerization coordinates the binding of two Jak2 molecules to dimerized receptors, while affecting overall receptor/Jak2 binding only modestly. Complexes containing more than two Jak2 molecules (e.g., J(RLR)JS2J) are rare.
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pcbi-1000364-g004: SH2-Bβ dimerization coordinates the formation of macro-complexes containing two Jak2 molecules bound to GH-dimerized receptors.Steady-state calculations were performed using the Simplified Cellular Model and the same parameter values as in Figure 3, except with 10 nM GH stimulation and varying SH2-Bβ concentration. (A) Receptor-bound, phosphorylated Jak2 (Y2∼P), for various values of the SH2-Bβ dimerization affinity. The extreme cases of KD,SS equal to zero and infinity correspond to irreversible and no dimerization, respectively; intermediate KD,SS values are 10 nM, 100 nM, 1 µM, and 10 µM. (B) Analysis of receptor/Jak2 complexes, with KD,SS = 100 nM. SH2-Bβ dimerization coordinates the binding of two Jak2 molecules to dimerized receptors, while affecting overall receptor/Jak2 binding only modestly. Complexes containing more than two Jak2 molecules (e.g., J(RLR)JS2J) are rare.

Mentions: To further characterize this hypothetical mechanism, the intracellular concentration and dimerization affinity of SH2-Bβ were varied for a constant GH concentration of 10 nM (Figure 4). Although a broad range of SH2-Bβ concentrations was tested in order to evaluate the full spectrum of behaviors, it is noted that the endogenous SH2-Bβ expression level is not expected to be above the nanomolar range. Given a constant Jak2/SH2-Bβ affinity (KD,JS = 100 nM), the SH2-Bβ concentration should be of a similar magnitude or somewhat higher for near maximal enhancement of Jak2 phosphorylation; extremely high SH2-Bβ concentrations, similar in magnitude to χr (100 µM; see Methods) are needed to antagonize the formation of the stable macro-complex, leading instead to formation of less stable, nine-member S2J(RLR)JS2 complexes (Figure 4A). Analysis of the GH receptor/Jak2 complexes formed reveals that, as expected, SH2-Bβ stabilizes complexes with two Jak2 molecules while increasing the total Jak2 recruitment only modestly (Figure 4B). In the absence of SH2-Bβ, approximately half of all GH receptors are Jak2-bound, and this constitutive binding accounts for a significant fraction of the total at all SH2-Bβ concentrations.


A bipolar clamp mechanism for activation of Jak-family protein tyrosine kinases.

Barua D, Faeder JR, Haugh JM - PLoS Comput. Biol. (2009)

SH2-Bβ dimerization coordinates the formation of macro-complexes containing two Jak2 molecules bound to GH-dimerized receptors.Steady-state calculations were performed using the Simplified Cellular Model and the same parameter values as in Figure 3, except with 10 nM GH stimulation and varying SH2-Bβ concentration. (A) Receptor-bound, phosphorylated Jak2 (Y2∼P), for various values of the SH2-Bβ dimerization affinity. The extreme cases of KD,SS equal to zero and infinity correspond to irreversible and no dimerization, respectively; intermediate KD,SS values are 10 nM, 100 nM, 1 µM, and 10 µM. (B) Analysis of receptor/Jak2 complexes, with KD,SS = 100 nM. SH2-Bβ dimerization coordinates the binding of two Jak2 molecules to dimerized receptors, while affecting overall receptor/Jak2 binding only modestly. Complexes containing more than two Jak2 molecules (e.g., J(RLR)JS2J) are rare.
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2667146&req=5

pcbi-1000364-g004: SH2-Bβ dimerization coordinates the formation of macro-complexes containing two Jak2 molecules bound to GH-dimerized receptors.Steady-state calculations were performed using the Simplified Cellular Model and the same parameter values as in Figure 3, except with 10 nM GH stimulation and varying SH2-Bβ concentration. (A) Receptor-bound, phosphorylated Jak2 (Y2∼P), for various values of the SH2-Bβ dimerization affinity. The extreme cases of KD,SS equal to zero and infinity correspond to irreversible and no dimerization, respectively; intermediate KD,SS values are 10 nM, 100 nM, 1 µM, and 10 µM. (B) Analysis of receptor/Jak2 complexes, with KD,SS = 100 nM. SH2-Bβ dimerization coordinates the binding of two Jak2 molecules to dimerized receptors, while affecting overall receptor/Jak2 binding only modestly. Complexes containing more than two Jak2 molecules (e.g., J(RLR)JS2J) are rare.
Mentions: To further characterize this hypothetical mechanism, the intracellular concentration and dimerization affinity of SH2-Bβ were varied for a constant GH concentration of 10 nM (Figure 4). Although a broad range of SH2-Bβ concentrations was tested in order to evaluate the full spectrum of behaviors, it is noted that the endogenous SH2-Bβ expression level is not expected to be above the nanomolar range. Given a constant Jak2/SH2-Bβ affinity (KD,JS = 100 nM), the SH2-Bβ concentration should be of a similar magnitude or somewhat higher for near maximal enhancement of Jak2 phosphorylation; extremely high SH2-Bβ concentrations, similar in magnitude to χr (100 µM; see Methods) are needed to antagonize the formation of the stable macro-complex, leading instead to formation of less stable, nine-member S2J(RLR)JS2 complexes (Figure 4A). Analysis of the GH receptor/Jak2 complexes formed reveals that, as expected, SH2-Bβ stabilizes complexes with two Jak2 molecules while increasing the total Jak2 recruitment only modestly (Figure 4B). In the absence of SH2-Bβ, approximately half of all GH receptors are Jak2-bound, and this constitutive binding accounts for a significant fraction of the total at all SH2-Bβ concentrations.

Bottom Line: Building on a rule-based kinetic modeling approach that considers the concerted nature and combinatorial complexity of modular protein domain interactions, we examine these mechanisms in detail, focusing on the growth hormone (GH) receptor/Jak2/SH2-Bbeta system.The modeling results suggest that, whereas Jak2-(SH2-Bbeta)(2)-Jak2 heterotetramers are scarcely expected to affect Jak2 phosphorylation, SH2-Bbeta and dimerized receptors synergistically promote Jak2 trans-activation in the context of intracellular signaling.Analysis of the results revealed a unique mechanism whereby SH2-B and receptor dimers constitute a bipolar 'clamp' that stabilizes the active configuration of two Jak2 molecules in the same macro-complex.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemical and Biomolecular Engineering, North Carolina State University, Raleigh, North Carolina, USA.

ABSTRACT
Most cell surface receptors for growth factors and cytokines dimerize in order to mediate signal transduction. For many such receptors, the Janus kinase (Jak) family of non-receptor protein tyrosine kinases are recruited in pairs and juxtaposed by dimerized receptor complexes in order to activate one another by trans-phosphorylation. An alternative mechanism for Jak trans-phosphorylation has been proposed in which the phosphorylated kinase interacts with the Src homology 2 (SH2) domain of SH2-B, a unique adaptor protein with the capacity to homo-dimerize. Building on a rule-based kinetic modeling approach that considers the concerted nature and combinatorial complexity of modular protein domain interactions, we examine these mechanisms in detail, focusing on the growth hormone (GH) receptor/Jak2/SH2-Bbeta system. The modeling results suggest that, whereas Jak2-(SH2-Bbeta)(2)-Jak2 heterotetramers are scarcely expected to affect Jak2 phosphorylation, SH2-Bbeta and dimerized receptors synergistically promote Jak2 trans-activation in the context of intracellular signaling. Analysis of the results revealed a unique mechanism whereby SH2-B and receptor dimers constitute a bipolar 'clamp' that stabilizes the active configuration of two Jak2 molecules in the same macro-complex.

Show MeSH
Related in: MedlinePlus