Ionizable networks mediate pH-dependent allostery in SH2 signaling proteins DOI Creative Commons
Papa Kobina Van Dyck, Luke Piszkin,

Elijah Gorski

и другие.

bioRxiv (Cold Spring Harbor Laboratory), Год журнала: 2024, Номер unknown

Опубликована: Авг. 21, 2024

Introduction Transient intracellular pH dynamics 1 regulate mammalian proliferation 2,3 , migration 4 and differentiation 5 . However, for many pH-dependent cell processes, the molecular mediators are unknown 6 Prior work identified histidine residues as switches in pH-sensitive proteins, but how other ionizable contribute to protein allostery is understudied. Here, we develop an silico computational pipeline identify putative proteins their mechanisms. We first apply this SHP2, a known signaling with uncharacterized mechanism. show wild-type SHP2 phosphatase activity vitro cells, mutation of H116 E252 non-titratable alanine abolishes function. also that c-Src previously unrecognized kinase, network again activity. Constant simulations support conserved allosteric mechanism binding inhibitory SH2 domains functional catalytic c-Src. our across domain-containing evolutionarily pH-sensing networks. Our results reveal regulator providing insight into normal biology diseases where pHi dysregulated, such cancer.

Язык: Английский

Integrating bioelectronics with cell-based synthetic biology DOI
Jonathan Rivnay, Ritu Raman, Jacob T. Robinson

и другие.

Nature Reviews Bioengineering, Год журнала: 2025, Номер unknown

Опубликована: Янв. 7, 2025

Язык: Английский

Процитировано

5
CD44 and Ezrin restrict EGF receptor mobility to generate a novel spatial arrangement of cytoskeletal signaling modules driving bleb–based migration DOI Creative Commons
Ankita Jha, Ankit Chandra, Payam E. Farahani

и другие.

bioRxiv (Cold Spring Harbor Laboratory), Год журнала: 2025, Номер unknown

Опубликована: Янв. 1, 2025

Cells under high confinement form highly polarized hydrostatic pressure-driven, stable leader blebs that enable efficient migration in low adhesion, environments. Here we investigated the basis of bleb morphology metastatic melanoma cells migrating non-adhesive confinement. Using high-resolution time-lapse imaging and specific molecular perturbations, found EGF signaling via PI3K stabilizes maintains a bleb. Protein activity biosensors revealed unique EGFR/PI3K gradient decreasing from rear-to-front, promoting PIP3 Rac1-GTP accumulation at rear, with its antagonists PIP2 RhoA-GTP concentrated tip, opposite to front-to-rear organization these modules integrin-mediated mesenchymal migration. Optogenetic experiments showed disrupting this caused retraction, underscoring role stability. Mathematical modeling identified mechanism where, as initiates, CD44 ERM proteins restrict EGFR mobility membrane-apposed cortical actin meshwork establishing rear-to-front EGFR-PI3K-Rac gradient. Thus, our study reveals biophysical underpinnings cell polarity bleb-based confinement, underscores how alternative spatial arrangements can mediate different modes according local microenvironment.

Язык: Английский

Процитировано

2
In vivomeasurements of receptor tyrosine kinase activity reveal feedback regulation of a developmental gradient DOI Creative Commons
Emily K. Ho, Rebecca Kim, Alison G. Simpkins

и другие.

bioRxiv (Cold Spring Harbor Laboratory), Год журнала: 2025, Номер unknown

Опубликована: Янв. 7, 2025

A lack of tools for detecting receptor activity in vivo has limited our ability to fully explore receptor-level control developmental patterning. Here, we extend a new class biosensors tyrosine kinase (RTK) activity, the pYtag system, visualize endogenous RTK Drosophila . We build three RTKs that function across stages and tissues. By characterizing Torso::pYtag during terminal patterning early embryo, find Torso differs from downstream ERK two surprising ways: is narrowly restricted poles but produces broader gradient ERK, decreases over time while sustained. This decrease driven by pathway-dependent negative feedback. Our results suggest an updated model where narrow domain tuned amplitude feedback, locally activates signaling effectors which diffuse through syncytial embryo form gradient. Altogether, this work highlights usefulness pYtags investigating regulation

Язык: Английский

Процитировано

0
Live imaging of paracrine signaling: Advances in visualization and tracking techniques DOI Creative Commons

Eriko Deguchi,

Michiyuki Matsuda, Kenta Terai

и другие.

Cell Structure and Function, Год журнала: 2025, Номер 50(1), С. 1 - 14

Опубликована: Янв. 1, 2025

Live imaging techniques have revolutionized our understanding of paracrine signaling, a crucial form cell-to-cell communication in biological processes. This review examines recent advances visualizing and tracking factors through four key stages: secretion from producing cells, diffusion extracellular space, binding to target activation intracellular signaling within cells. Paracrine factor can be directly visualized by fluorescent protein tagging ligand, or indirectly the cleavage transmembrane pro-ligands plasma membrane fusion endosomes comprising factors. Diffusion has been studied using such as fluorescence correlation spectroscopy (FCS), recovery after photobleaching (FRAP), decay photoactivation (FDAP), single-molecule tracking. Binding cells various biosensors, including GPCR-activation-based (GRAB) sensors Förster resonance energy transfer (FRET) probes for receptor tyrosine kinases. Finally, is monitored biosensors second messengers, transcription factors, so on. In addition tools, also highlights emerging optogenetic chemogenetic tools triggering release which essential associating outcomes during bioimaging signaling.Key words: live imaging, optogenetics, chemogenetics.

Язык: Английский

Процитировано

0
Principles and Design of Molecular Tools for Sensing and Perturbing Cell Surface Receptor Activity DOI
Jennifer Sescil, Steven M. Havens, Wenjing Wang

и другие.

Chemical Reviews, Год журнала: 2025, Номер unknown

Опубликована: Фев. 25, 2025

Cell-surface receptors are vital for controlling numerous cellular processes with their dysregulation being linked to disease states. Therefore, it is necessary develop tools study and the signaling pathways they control. This Review broadly describes molecular approaches that enable 1) visualization of determine localization distribution; 2) sensing receptor activation permanent readouts as well in real time; 3) perturbing activity mimicking receptor-controlled learn more about these processes. Together, have provided valuable insight into fundamental biology helped characterize therapeutics target receptors.

Язык: Английский

Процитировано

0
Design and optimization of a kinase-controlled allosteric switch DOI Creative Commons
Qi Cao, Jared E. Toettcher

bioRxiv (Cold Spring Harbor Laboratory), Год журнала: 2025, Номер unknown

Опубликована: Апрель 15, 2025

Abstract Post-translational control enables rapid and precise regulation of cell behavior. Despite these advantages, general strategies to build phosphorylation-based synthetic circuits are limited. Here, we reasoned that engineered allostery, a technique has been applied design light- chemically-gated protein switches, could also be used engineer phosphorylation-controlled switches (phospho-switches). Using an allosterically controllable Gal4 transcription factor as scaffold, show classic kinase FRET biosensor architecture can starting point for phospho-switch design. We optimize all features the develop ERK-controlled with 20-fold phosphorylation-dependent change in transcriptional output. Our switch outperforms c-fos promoter, ERK-responsive biosensor, selectivity sensitivity. further our generalized other input kinases controlled targets. This work provides platform new generation kinase-responsive tools biosensing biology applications.

Язык: Английский

Процитировано

0
An optogenetic toolkit for robust activation of FGF, BMP, & Nodal signaling in zebrafish DOI Creative Commons
Leanne E. Iannucci, Varghese Thomas,

Micaela R. Murphy

и другие.

bioRxiv (Cold Spring Harbor Laboratory), Год журнала: 2025, Номер unknown

Опубликована: Апрель 19, 2025

ABSTRACT Cell signaling regulates a wide range of biological processes including development, homeostasis, and disease. Accessible technologies to precisely manipulate have important applications in basic translational research. Here, we introduce an optogenetic toolkit comprised 1) zebrafish-optimized FGF activator, 2) single-transcript Nodal 3) previously established BMP activator. We thoroughly characterize this suite tools zebrafish embryos show that they provide tunable, light-dependent spatiotemporal control vivo . In response blue light (∼455 nm), receptor kinase domains fused light-dimerizing LOV enable robust activation with minimal ectopic activity the dark or at wavelengths over 495 nm. Optogenetic by each tool is pathway-specific results increased expression known target genes. Signaling activated rapid on/off kinetics, strength depends on irradiance. Finally, demonstrate spatially localized our optimized Together, establish as potent experimental platform rapidly, directly, adjustably activate FGF, BMP, embryos.

Язык: Английский

Процитировано

0
AlkTango reveals a role for Jeb/Alk signaling in the Drosophila heart DOI Creative Commons
Georg Wolfstetter, Tafheem Masudi, Ezgi Uçkun

и другие.

Cell Communication and Signaling, Год журнала: 2025, Номер 23(1)

Опубликована: Май 17, 2025

Anaplastic lymphoma kinase (Alk) signaling is important in a variety of biological contexts such as cell type specification, regulation metabolic and endocrine programs, behavior, cancer. In this work, we generated Tango GPCR assay-based, dimerization-sensitive Alk activity reporter (AlkTango) followed receptor activation throughout Drosophila development. AlkTango reports embryonic larval tissues previously linked to signaling. Remarkably, was active the heart larvae adult flies. We show that cardiomyocytes express from late stages adulthood, while jeb expression pericardial cells coincided with activity. Perturbation cardiac leads decreased survival well lower fitness increased lethality response heat stress. keeping role for Alk, measurements reveal arrythmia irregular muscle contraction upon ligand stimulation. Finally, induces hyperplasia accessory wing hearts

Язык: Английский

Процитировано

0
Engineered chimeric receptors for dissecting interferon signaling DOI
Aaron E. Lin, Emily V. Mesev, Jared E. Toettcher

и другие.

Journal of Virology, Год журнала: 2024, Номер unknown

Опубликована: Сен. 18, 2024

ABSTRACT Though interferons (IFNs) were once heralded as panaceas to numerous diseases, how cells decode varying IFN stimuli and subsequently produce (in)appropriate signaling remain unclear. Our labs recently engineered novel erythropoietin receptor-IFN chimeric receptors, we highlight their utility in two cases uncovering differential genetic determinants of type I (IFN-α/β) III (IFN-λ) signaling. These other types synthetic (cytokine) receptors could be expanded real-time dynamics vivo studies.

Язык: Английский

Процитировано

1
Ionizable networks mediate pH-dependent allostery in SH2 signaling proteins DOI Creative Commons
Papa Kobina Van Dyck, Luke Piszkin,

Elijah Gorski

и другие.

bioRxiv (Cold Spring Harbor Laboratory), Год журнала: 2024, Номер unknown

Опубликована: Авг. 21, 2024

Introduction Transient intracellular pH dynamics 1 regulate mammalian proliferation 2,3 , migration 4 and differentiation 5 . However, for many pH-dependent cell processes, the molecular mediators are unknown 6 Prior work identified histidine residues as switches in pH-sensitive proteins, but how other ionizable contribute to protein allostery is understudied. Here, we develop an silico computational pipeline identify putative proteins their mechanisms. We first apply this SHP2, a known signaling with uncharacterized mechanism. show wild-type SHP2 phosphatase activity vitro cells, mutation of H116 E252 non-titratable alanine abolishes function. also that c-Src previously unrecognized kinase, network again activity. Constant simulations support conserved allosteric mechanism binding inhibitory SH2 domains functional catalytic c-Src. our across domain-containing evolutionarily pH-sensing networks. Our results reveal regulator providing insight into normal biology diseases where pHi dysregulated, such cancer.

Язык: Английский

Процитировано

0