Chemically Controlled Protein Assembly: Techniques and Applications

Fegan, Adrian; White, Brian R.; Carlson, Jonathan C.; Wagner, Carston R.

doi:10.1021/cr8002888

Cited by 274 publications

(235 citation statements)

References 159 publications

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“…Interestingly, the response is biphasic because activation is reduced at the highest concentration. This behavior, however, is consistent with the properties of bifunctional ligands because at sufficiently high concentrations, the formation of binary ligand-protein complexes is favored over the ternary complex (32,37). It is also notable that the half-maximal activation by the FK506 conjugate occurred at concentrations (EC 50 ~3 nM) significantly lower than the intrinsic receptor affinity ( ~90 nM).…”

Section: Efficacy Switch From Antagonist To Agonist Through Extrinsicsupporting

confidence: 68%

“…These data argue that the enhanced activation in the presence of the fusion is due to extrinsic recruitment by the bifunctional ligand. If this is indeed the case, it can be anticipated that the effect should be disrupted by excess unconjugated FK506 (32). In support of this prediction, increasing concentrations of free FK506 reduced the activity of the bifunctional ligand ( Figure 4C) until it was essentially indistinguishable from the behavior of the ligand lacking FK506 (except for a small increase in basal activity at the highest concentration of FK506).…”

Section: Design Of Bifunctional Ligandsmentioning

confidence: 71%

See 1 more Smart Citation

Targeting Prostate Cancer with Bifunctional Modulators of the Androgen Receptor

Mapp¹,

Sturlis²,

Carolan³

et al. 2015

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show abstract

Section: Efficacy Switch From Antagonist To Agonist Through Extrinsicsupporting

confidence: 68%

Section: Design Of Bifunctional Ligandsmentioning

confidence: 71%

Targeting Prostate Cancer with Bifunctional Modulators of the Androgen Receptor

Mapp¹,

Sturlis²,

Carolan³

et al. 2015

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show abstract

“…Aurora B activity at kinetochores was manipulated using rapamycin-inducible dimerization (45)(46)(47)(48) in a stable cell line expressing Mis12-GFP-FKBP, mCherry-INbox-FRB, and shRNA against endogenous FKBP. FKBP and FRB are dimerization domains that bind rapamycin.…”

Section: Methodsmentioning

confidence: 99%

Minimal model for collective kinetochore–microtubule dynamics

Banigan

Chiou

Ballister

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Chromosome segregation during cell division depends on interactions of kinetochores with dynamic microtubules (MTs). In many eukaryotes, each kinetochore binds multiple MTs, but the collective behavior of these coupled MTs is not well understood. We present a minimal model for collective kinetochore–MT dynamics, based on in vitro measurements of individual MTs and their dependence on force and kinetochore phosphorylation by Aurora B kinase. For a system of multiple MTs connected to the same kinetochore, the force–velocity relation has a bistable regime with two possible steady-state velocities: rapid shortening or slow growth. Bistability, combined with the difference between the growing and shrinking speeds, leads to center-of-mass and breathing oscillations in bioriented sister kinetochore pairs. Kinetochore phosphorylation shifts the bistable region to higher tensions, so that only the rapidly shortening state is stable at low tension. Thus, phosphorylation leads to error correction for kinetochores that are not under tension. We challenged the model with new experiments, using chemically induced dimerization to enhance Aurora B activity at metaphase kinetochores. The model suggests that the experimentally observed disordering of the metaphase plate occurs because phosphorylation increases kinetochore speeds by biasing MTs to shrink. Our minimal model qualitatively captures certain characteristic features of kinetochore dynamics, illustrates how biochemical signals such as phosphorylation may regulate the dynamics, and provides a theoretical framework for understanding other factors that control the dynamics in vivo.

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“…The analysis here represents a more general framework extensible to other rapamycin-activatable signaling molecules (25,50,51), as well as to other cell types and multicellular systems. Furthermore, the effects of gradients of other proteins engineered to be sensitive to small membrane-permeable molecules, such as ATP analogs (52) and imidazole (53), could also be analyzed to refine our understanding of the mechanisms of cell responses to graded intracellular signaling activity.…”

Section: Discussionmentioning

confidence: 99%

Synthetic spatially graded Rac activation drives cell polarization and movement

Lin

Holmes

Wang

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

101

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Migrating cells possess intracellular gradients of active Rho GTPases, which serve as central hubs in transducing signals from extracellular receptors to cytoskeletal and adhesive machinery. However, it is unknown whether shallow exogenously induced intracellular gradients of Rho GTPases are sufficient to drive cell polarity and motility. Here, we use microfluidic control to generate gradients of a small molecule and thereby directly induce linear gradients of active, endogenous Rac without activation of chemotactic receptors. Gradients as low as 15% were sufficient not only to trigger cell migration up the chemical gradient but to induce both cell polarization and repolarization. Cellular response times were inversely proportional to the steepness of Rac inducer gradient in agreement with a mathematical model, suggesting a function for chemoattractant gradient amplification upstream of Rac. Increases in activated Rac levels beyond a well-defined threshold augmented polarization and decreased sensitivity to the imposed gradient. The threshold was governed by initial cell polarity and PI3K activity, supporting a role for both in defining responsiveness to Rac activation. Our results reveal that Rac can serve as a starting point in defining cell polarity. Furthermore, our methodology may serve as a template to investigate processes regulated by intracellular signaling gradients.cell signaling | synthetic biology | chemotaxis | signal transduction

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Chemically Controlled Protein Assembly: Techniques and Applications

Cited by 274 publications

References 159 publications

Targeting Prostate Cancer with Bifunctional Modulators of the Androgen Receptor

Targeting Prostate Cancer with Bifunctional Modulators of the Androgen Receptor

Minimal model for collective kinetochore–microtubule dynamics

Synthetic spatially graded Rac activation drives cell polarization and movement

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