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1.
Cell ; 186(21): 4710-4727.e35, 2023 10 12.
Artigo em Inglês | MEDLINE | ID: mdl-37774705

RESUMO

Polarized cells rely on a polarized cytoskeleton to function. Yet, how cortical polarity cues induce cytoskeleton polarization remains elusive. Here, we capitalized on recently established designed 2D protein arrays to ectopically engineer cortical polarity of virtually any protein of interest during mitosis in various cell types. This enables direct manipulation of polarity signaling and the identification of the cortical cues sufficient for cytoskeleton polarization. Using this assay, we dissected the logic of the Par complex pathway, a key regulator of cytoskeleton polarity during asymmetric cell division. We show that cortical clustering of any Par complex subunit is sufficient to trigger complex assembly and that the primary kinetic barrier to complex assembly is the relief of Par6 autoinhibition. Further, we found that inducing cortical Par complex polarity induces two hallmarks of asymmetric cell division in unpolarized mammalian cells: spindle orientation, occurring via Par3, and central spindle asymmetry, depending on aPKC activity.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Polaridade Celular , Técnicas Citológicas , Mitose , Animais , Citoesqueleto/metabolismo , Mamíferos/metabolismo , Microtúbulos/metabolismo , Proteína Quinase C/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo
2.
Nature ; 626(7998): 435-442, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38109936

RESUMO

Many peptide hormones form an α-helix on binding their receptors1-4, and sensitive methods for their detection could contribute to better clinical management of disease5. De novo protein design can now generate binders with high affinity and specificity to structured proteins6,7. However, the design of interactions between proteins and short peptides with helical propensity is an unmet challenge. Here we describe parametric generation and deep learning-based methods for designing proteins to address this challenge. We show that by extending RFdiffusion8 to enable binder design to flexible targets, and to refining input structure models by successive noising and denoising (partial diffusion), picomolar-affinity binders can be generated to helical peptide targets by either refining designs generated with other methods, or completely de novo starting from random noise distributions without any subsequent experimental optimization. The RFdiffusion designs enable the enrichment and subsequent detection of parathyroid hormone and glucagon by mass spectrometry, and the construction of bioluminescence-based protein biosensors. The ability to design binders to conformationally variable targets, and to optimize by partial diffusion both natural and designed proteins, should be broadly useful.


Assuntos
Desenho Assistido por Computador , Aprendizado Profundo , Peptídeos , Proteínas , Técnicas Biossensoriais , Difusão , Glucagon/química , Glucagon/metabolismo , Medições Luminescentes , Espectrometria de Massas , Hormônio Paratireóideo/química , Hormônio Paratireóideo/metabolismo , Peptídeos/química , Peptídeos/metabolismo , Estrutura Secundária de Proteína , Proteínas/química , Proteínas/metabolismo , Especificidade por Substrato , Modelos Moleculares
3.
Nature ; 623(7988): 842-852, 2023 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-37853127

RESUMO

Optimum protein function and biochemical activity critically depends on water availability because solvent thermodynamics drive protein folding and macromolecular interactions1. Reciprocally, macromolecules restrict the movement of 'structured' water molecules within their hydration layers, reducing the available 'free' bulk solvent and therefore the total thermodynamic potential energy of water, or water potential. Here, within concentrated macromolecular solutions such as the cytosol, we found that modest changes in temperature greatly affect the water potential, and are counteracted by opposing changes in osmotic strength. This duality of temperature and osmotic strength enables simple manipulations of solvent thermodynamics to prevent cell death after extreme cold or heat shock. Physiologically, cells must sustain their activity against fluctuating temperature, pressure and osmotic strength, which impact water availability within seconds. Yet, established mechanisms of water homeostasis act over much slower timescales2,3; we therefore postulated the existence of a rapid compensatory response. We find that this function is performed by water potential-driven changes in macromolecular assembly, particularly biomolecular condensation of intrinsically disordered proteins. The formation and dissolution of biomolecular condensates liberates and captures free water, respectively, quickly counteracting thermal or osmotic perturbations of water potential, which is consequently robustly buffered in the cytoplasm. Our results indicate that biomolecular condensation constitutes an intrinsic biophysical feedback response that rapidly compensates for intracellular osmotic and thermal fluctuations. We suggest that preserving water availability within the concentrated cytosol is an overlooked evolutionary driver of protein (dis)order and function.


Assuntos
Substâncias Macromoleculares , Proteínas , Solventes , Termodinâmica , Água , Morte Celular , Citosol/química , Citosol/metabolismo , Homeostase , Substâncias Macromoleculares/química , Substâncias Macromoleculares/metabolismo , Concentração Osmolar , Pressão , Proteínas/química , Proteínas/metabolismo , Solventes/química , Solventes/metabolismo , Temperatura , Fatores de Tempo , Água/química , Água/metabolismo
4.
Nature ; 620(7976): 1089-1100, 2023 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-37433327

RESUMO

There has been considerable recent progress in designing new proteins using deep-learning methods1-9. Despite this progress, a general deep-learning framework for protein design that enables solution of a wide range of design challenges, including de novo binder design and design of higher-order symmetric architectures, has yet to be described. Diffusion models10,11 have had considerable success in image and language generative modelling but limited success when applied to protein modelling, probably due to the complexity of protein backbone geometry and sequence-structure relationships. Here we show that by fine-tuning the RoseTTAFold structure prediction network on protein structure denoising tasks, we obtain a generative model of protein backbones that achieves outstanding performance on unconditional and topology-constrained protein monomer design, protein binder design, symmetric oligomer design, enzyme active site scaffolding and symmetric motif scaffolding for therapeutic and metal-binding protein design. We demonstrate the power and generality of the method, called RoseTTAFold diffusion (RFdiffusion), by experimentally characterizing the structures and functions of hundreds of designed symmetric assemblies, metal-binding proteins and protein binders. The accuracy of RFdiffusion is confirmed by the cryogenic electron microscopy structure of a designed binder in complex with influenza haemagglutinin that is nearly identical to the design model. In a manner analogous to networks that produce images from user-specified inputs, RFdiffusion enables the design of diverse functional proteins from simple molecular specifications.


Assuntos
Aprendizado Profundo , Proteínas , Domínio Catalítico , Microscopia Crioeletrônica , Glicoproteínas de Hemaglutininação de Vírus da Influenza/química , Glicoproteínas de Hemaglutininação de Vírus da Influenza/metabolismo , Glicoproteínas de Hemaglutininação de Vírus da Influenza/ultraestrutura , Ligação Proteica , Proteínas/química , Proteínas/metabolismo , Proteínas/ultraestrutura
5.
EMBO J ; 42(23): e114473, 2023 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-37872872

RESUMO

The microtubule motor dynein mediates polarised trafficking of a wide variety of organelles, vesicles and macromolecules. These functions are dependent on the dynactin complex, which helps recruit cargoes to dynein's tail and activates motor movement. How the dynein-dynactin complex orchestrates trafficking of diverse cargoes is unclear. Here, we identify HEATR5B, an interactor of the adaptor protein-1 (AP1) clathrin adaptor complex, as a novel player in dynein-dynactin function. HEATR5B was recovered in a biochemical screen for proteins whose association with the dynein tail is augmented by dynactin. We show that HEATR5B binds directly to the dynein tail and dynactin and stimulates motility of AP1-associated endosomal membranes in human cells. We also demonstrate that the Drosophila HEATR5B homologue is an essential gene that selectively promotes dynein-based transport of AP1-bound membranes to the Golgi apparatus. As HEATR5B lacks the coiled-coil architecture typical of dynein adaptors, our data point to a non-canonical process orchestrating motor function on a specific cargo. We additionally show that HEATR5B promotes association of AP1 with endosomal membranes independently of dynein. Thus, HEATR5B co-ordinates multiple events in AP1-based trafficking.


Assuntos
Dineínas , Proteínas Associadas aos Microtúbulos , Humanos , Dineínas/metabolismo , Complexo Dinactina/metabolismo , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Transporte Biológico/fisiologia , Microtúbulos/metabolismo , Endossomos/metabolismo
6.
Nature ; 589(7842): 468-473, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33408408

RESUMO

Ordered two-dimensional arrays such as S-layers1,2 and designed analogues3-5 have intrigued bioengineers6,7, but with the exception of a single lattice formed with flexible linkers8, they are constituted from just one protein component. Materials composed of two components have considerable potential advantages for modulating assembly dynamics and incorporating more complex functionality9-12. Here we describe a computational method to generate co-assembling binary layers by designing rigid interfaces between pairs of dihedral protein building blocks, and use it to design a p6m lattice. The designed array components are soluble at millimolar concentrations, but when combined at nanomolar concentrations, they rapidly assemble into nearly crystalline micrometre-scale arrays nearly identical to the computational design model in vitro and in cells without the need for a two-dimensional support. Because the material is designed from the ground up, the components can be readily functionalized and their symmetry reconfigured, enabling formation of ligand arrays with distinguishable surfaces, which we demonstrate can drive extensive receptor clustering, downstream protein recruitment and signalling. Using atomic force microscopy on supported bilayers and quantitative microscopy on living cells, we show that arrays assembled on membranes have component stoichiometry and structure similar to arrays formed in vitro, and that our material can therefore impose order onto fundamentally disordered substrates such as cell membranes. In contrast to previously characterized cell surface receptor binding assemblies such as antibodies and nanocages, which are rapidly endocytosed, we find that large arrays assembled at the cell surface suppress endocytosis in a tunable manner, with potential therapeutic relevance for extending receptor engagement and immune evasion. Our work provides a foundation for a synthetic cell biology in which multi-protein macroscale materials are designed to modulate cell responses and reshape synthetic and living systems.


Assuntos
Desenho de Fármacos , Engenharia de Proteínas , Proteínas/síntese química , Proteínas/metabolismo , Células 3T3 , Animais , Biologia Celular , Sobrevivência Celular , Biologia Computacional , Endocitose , Escherichia coli/genética , Escherichia coli/metabolismo , Técnicas In Vitro , Cinética , Ligantes , Camundongos , Microscopia de Força Atômica , Modelos Moleculares , Biologia Sintética
9.
Artigo em Inglês | MEDLINE | ID: mdl-38438190

RESUMO

Designing proteins with tailored structures and functions is a long-standing goal in bioengineering. Recently, deep learning advances have enabled protein structure prediction at near-experimental accuracy, which has catalyzed progress in protein design as well. We review recent studies that use structure-prediction neural networks to design proteins, via approaches such as activation maximization, inpainting, or denoising diffusion. These methods have led to major improvements over previous methods in wet-lab success rates for designing protein binders, metalloproteins, enzymes, and oligomeric assemblies. These results show that structure-prediction models are a powerful foundation for developing protein-design tools and suggest that continued improvement of their accuracy and generality will be key to unlocking the full potential of protein design.


Assuntos
Proteínas , Proteínas/química , Conformação Proteica , Redes Neurais de Computação , Modelos Moleculares , Engenharia de Proteínas , Dobramento de Proteína
10.
bioRxiv ; 2024 Sep 22.
Artigo em Inglês | MEDLINE | ID: mdl-39345553

RESUMO

The design of inducibly assembling protein nanomaterials is an outstanding challenge. Here, we describe the computational design of a protein filament formed from a monomeric subunit which binds a peptide ligand. The cryoEM structure of the micron scale fibers is very close to the computational design model. The ligand acts as a tunable allosteric modulator: while not part of the fiber subunit-subunit interfaces, the assembly of the filament is dependent on ligand addition, with longer peptides having more extensive interaction surfaces with the monomer promoting more rapid growth. Seeded growth and capping experiments reveal that the filaments grow primarily from one end. Oligomers containing 12 copies of the peptide ligand nucleate fiber assembly from monomeric subunit and peptide mixtures at concentrations where assembly occurs very slowly, likely by generating critical local concentrations of monomer in the assembly competent conformation. Following filament assembly, the peptide ligand can be exchanged with free peptide in solution, and it can be readily fused to any functional protein of interest, opening the door to a wide variety of tunable engineered materials.

11.
bioRxiv ; 2024 Jul 23.
Artigo em Inglês | MEDLINE | ID: mdl-39091726

RESUMO

Francis Crick's global parameterization of coiled coil geometry has been widely useful for guiding design of new protein structures and functions. However, design guided by similar global parameterization of beta barrel structures has been less successful, likely due to the deviations required from ideal beta barrel geometry to maintain extensive inter-strand hydrogen bonding without introducing considerable backbone strain. Instead, beta barrels and other protein folds have been designed guided by 2D structural blueprints; while this approach has successfully generated new fluorescent proteins, transmembrane nanopores, and other structures, it requires considerable expert knowledge and provides only indirect control over the global barrel shape. Here we show that the simplicity and control over shape and structure provided by global parametric representations can be generalized beyond coiled coils by taking advantage of the rich sequence-structure relationships implicit in RoseTTAFold based inpainting and diffusion design methods. Starting from parametrically generated idealized barrel backbones, both RFjoint inpainting and RFdiffusion readily incorporate the backbone irregularities necessary for proper folding with minimal deviation from the idealized barrel geometries. We show that for beta barrels across a broad range of global beta sheet parameterizations, these methods achieve high in silico and experimental success rates, with atomic accuracy confirmed by an X-ray crystal structure of a novel beta barrel topology, and de novo designed 12, 14, and 16 stranded transmembrane nanopores with conductances ranging from 200 to 500 pS. By combining the simplicity and control of parametric generation with the high success rates of deep learning based protein design methods, our approach makes the design of proteins where global shape confers function, such as beta barrel nanopores, more precisely specifiable and accessible.

12.
Nat Nanotechnol ; 19(7): 1016-1021, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38570702

RESUMO

Biological evolution has led to precise and dynamic nanostructures that reconfigure in response to pH and other environmental conditions. However, designing micrometre-scale protein nanostructures that are environmentally responsive remains a challenge. Here we describe the de novo design of pH-responsive protein filaments built from subunits containing six or nine buried histidine residues that assemble into micrometre-scale, well-ordered fibres at neutral pH. The cryogenic electron microscopy structure of an optimized design is nearly identical to the computational design model for both the subunit internal geometry and the subunit packing into the fibre. Electron, fluorescent and atomic force microscopy characterization reveal a sharp and reversible transition from assembled to disassembled fibres over 0.3 pH units, and rapid fibre disassembly in less than 1 s following a drop in pH. The midpoint of the transition can be tuned by modulating buried histidine-containing hydrogen bond networks. Computational protein design thus provides a route to creating unbound nanomaterials that rapidly respond to small pH changes.


Assuntos
Histidina , Concentração de Íons de Hidrogênio , Histidina/química , Proteínas/química , Nanoestruturas/química , Modelos Moleculares , Ligação de Hidrogênio , Microscopia Crioeletrônica
13.
Brain Pathol ; 34(6): e13263, 2024 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-38659387

RESUMO

Multiple sclerosis (MS) is unsurpassed for its clinical and pathological hetherogeneity, but the biological determinants of this variability are unknown. HLA-DRB1*15, the main genetic risk factor for MS, influences the severity and distribution of MS pathology. This study set out to unravel the molecular determinants of the heterogeneity of MS pathology in relation to HLA-DRB1*15 status. Shotgun proteomics from a discovery cohort of MS spinal cord samples segregated by HLA-DRB*15 status revealed overexpression of the extracellular matrix (ECM) proteins, biglycan, decorin, and prolargin in HLA-DRB*15-positive cases, adding to established literature on a role of ECM proteins in MS pathology that has heretofore lacked systematic pathological validation. These findings informed a neuropathological characterisation of these proteins in a large autopsy cohort of 41 MS cases (18 HLA-DRB1*15-positive and 23 HLA-DRB1*15-negative), and seven non-neurological controls on motor cortical, cervical and lumbar spinal cord tissue. Biglycan and decorin demonstrate a striking perivascular expression pattern in controls that is reduced in MS (-36.5%, p = 0.036 and - 24.7%, p = 0.039; respectively) in lesional and non-lesional areas. A concomitant increase in diffuse parenchymal accumulation of biglycan and decorin is seen in MS (p = 0.015 and p = 0.001, respectively), particularly in HLA-DRB1*15-positive cases (p = 0.007 and p = 0.046, respectively). Prolargin shows a faint parenchymal pattern in controls that is markedly increased in MS cases where a perivascular deposition pattern is observed (motor cortex +97.5%, p = 0.001; cervical cord +49.1%, p = 0.016). Our findings point to ECM proteins and the vascular interface playing a central role in MS pathology within and outside the plaque area. As ECM proteins are known potent pro-inflammatory molecules, their parenchymal accumulation may contribute to disease severity. This study brings to light novel factors that may contribute to the heterogeneity of the topographical variation of MS pathology.


Assuntos
Biglicano , Decorina , Proteínas da Matriz Extracelular , Esclerose Múltipla , Medula Espinal , Humanos , Esclerose Múltipla/patologia , Esclerose Múltipla/metabolismo , Proteínas da Matriz Extracelular/metabolismo , Masculino , Feminino , Pessoa de Meia-Idade , Adulto , Decorina/metabolismo , Decorina/genética , Medula Espinal/patologia , Medula Espinal/metabolismo , Idoso , Biglicano/metabolismo , Biglicano/genética , Cadeias HLA-DRB1/genética , Estudos de Coortes , Proteômica
14.
bioRxiv ; 2024 Jul 16.
Artigo em Inglês | MEDLINE | ID: mdl-39071267

RESUMO

Proteins which bind intrinsically disordered proteins (IDPs) and intrinsically disordered regions (IDRs) with high affinity and specificity could have considerable utility for therapeutic and diagnostic applications. However, a general methodology for targeting IDPs/IDRs has yet to be developed. Here, we show that starting only from the target sequence of the input, and freely sampling both target and binding protein conformation, RFdiffusion can generate binders to IDPs and IDRs in a wide range of conformations. We use this approach to generate binders to the IDPs Amylin, C-peptide and VP48 in a range of conformations with Kds in the 3 -100nM range. The Amylin binder inhibits amyloid fibril formation and dissociates existing fibers, and enables enrichment of amylin for mass spectrometry-based detection. For the IDRs G3bp1, common gamma chain (IL2RG) and prion, we diffused binders to beta strand conformations of the targets, obtaining 10 to 100 nM affinity. The IL2RG binder colocalizes with the receptor in cells, enabling new approaches to modulating IL2 signaling. Our approach should be widely useful for creating binders to flexible IDPs/IDRs spanning a wide range of intrinsic conformational preferences.

15.
bioRxiv ; 2024 Mar 18.
Artigo em Inglês | MEDLINE | ID: mdl-38562682

RESUMO

Despite the central role that antibodies play in modern medicine, there is currently no way to rationally design novel antibodies to bind a specific epitope on a target. Instead, antibody discovery currently involves time-consuming immunization of an animal or library screening approaches. Here we demonstrate that a fine-tuned RFdiffusion network is capable of designing de novo antibody variable heavy chains (VHH's) that bind user-specified epitopes. We experimentally confirm binders to four disease-relevant epitopes, and the cryo-EM structure of a designed VHH bound to influenza hemagglutinin is nearly identical to the design model both in the configuration of the CDR loops and the overall binding pose.

16.
Protein Sci ; 32(11): e4780, 2023 11.
Artigo em Inglês | MEDLINE | ID: mdl-37695922

RESUMO

Predicting the effects of mutations on protein function and stability is an outstanding challenge. Here, we assess the performance of a variant of RoseTTAFold jointly trained for sequence and structure recovery, RFjoint , for mutation effect prediction. Without any further training, we achieve comparable accuracy in predicting mutation effects for a diverse set of protein families using RFjoint to both another zero-shot model (MSA Transformer) and a model that requires specific training on a particular protein family for mutation effect prediction (DeepSequence). Thus, although the architecture of RFjoint was developed to address the protein design problem of scaffolding functional motifs, RFjoint acquired an understanding of the mutational landscapes of proteins during model training that is equivalent to that of recently developed large protein language models. The ability to simultaneously reason over protein structure and sequence could enable even more precise mutation effect predictions following supervised training on the task. These results suggest that RFjoint has a quite broad understanding of protein sequence-structure landscapes, and can be viewed as a joint model for protein sequence and structure which could be broadly useful for protein modeling.


Assuntos
Proteínas , Proteínas/genética , Proteínas/química , Mutação , Sequência de Aminoácidos , Estabilidade Proteica
17.
bioRxiv ; 2023 Mar 24.
Artigo em Inglês | MEDLINE | ID: mdl-36993242

RESUMO

Phase transitions of cellular proteins and lipids play a key role in governing the organisation and coordination of intracellular biology. The frequent juxtaposition of proteinaceous biomolecular condensates to cellular membranes raises the intriguing prospect that phase transitions in proteins and lipids could be co-regulated. Here we investigate this possibility in the ribonucleoprotein (RNP) granule-ANXA11-lysosome ensemble, where ANXA11 tethers RNP granule condensates to lysosomal membranes to enable their co-trafficking. We show that changes to the protein phase state within this system, driven by the low complexity ANXA11 N-terminus, induce a coupled phase state change in the lipids of the underlying membrane. We identify the ANXA11 interacting proteins ALG2 and CALC as potent regulators of ANXA11-based phase coupling and demonstrate their influence on the nanomechanical properties of the ANXA11-lysosome ensemble and its capacity to engage RNP granules. The phenomenon of protein-lipid phase coupling we observe within this system offers an important template to understand the numerous other examples across the cell whereby biomolecular condensates closely juxtapose cell membranes.

18.
Science ; 377(6604): 387-394, 2022 07 22.
Artigo em Inglês | MEDLINE | ID: mdl-35862514

RESUMO

The binding and catalytic functions of proteins are generally mediated by a small number of functional residues held in place by the overall protein structure. Here, we describe deep learning approaches for scaffolding such functional sites without needing to prespecify the fold or secondary structure of the scaffold. The first approach, "constrained hallucination," optimizes sequences such that their predicted structures contain the desired functional site. The second approach, "inpainting," starts from the functional site and fills in additional sequence and structure to create a viable protein scaffold in a single forward pass through a specifically trained RoseTTAFold network. We use these two methods to design candidate immunogens, receptor traps, metalloproteins, enzymes, and protein-binding proteins and validate the designs using a combination of in silico and experimental tests.


Assuntos
Aprendizado Profundo , Engenharia de Proteínas , Proteínas , Sítios de Ligação , Catálise , Ligação Proteica , Engenharia de Proteínas/métodos , Dobramento de Proteína , Estrutura Secundária de Proteína , Proteínas/química
19.
J Cell Biol ; 220(2)2021 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-33416860

RESUMO

Protein micropatterning allows proteins to be precisely deposited onto a substrate of choice and is now routinely used in cell biology and in vitro reconstitution. However, drawbacks of current technology are that micropatterning efficiency can be variable between proteins and that proteins may lose activity on the micropatterns. Here, we describe a general method to enable micropatterning of virtually any protein at high specificity and homogeneity while maintaining its activity. Our method is based on an anchor that micropatterns well, fibrinogen, which we functionalized to bind to common purification tags. This enhances micropatterning on various substrates, facilitates multiplexed micropatterning, and dramatically improves the on-pattern activity of fragile proteins like molecular motors. Furthermore, it enhances the micropatterning of hard-to-micropattern cells. Last, this method enables subcellular micropatterning, whereby complex micropatterns simultaneously control cell shape and the distribution of transmembrane receptors within that cell. Altogether, these results open new avenues for cell biology.


Assuntos
Fibrinogênio/metabolismo , Microtecnologia/métodos , Animais , Linhagem Celular , Proteínas de Fluorescência Verde/metabolismo , Humanos , Ligantes , Polietilenoglicóis/química , Frações Subcelulares/metabolismo
20.
Nat Commun ; 12(1): 6035, 2021 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-34654800

RESUMO

Between 6-20% of the cellular proteome is under circadian control and tunes mammalian cell function with daily environmental cycles. For cell viability, and to maintain volume within narrow limits, the daily variation in osmotic potential exerted by changes in the soluble proteome must be counterbalanced. The mechanisms and consequences of this osmotic compensation have not been investigated before. In cultured cells and in tissue we find that compensation involves electroneutral active transport of Na+, K+, and Cl- through differential activity of SLC12A family cotransporters. In cardiomyocytes ex vivo and in vivo, compensatory ion fluxes confer daily variation in electrical activity. Perturbation of soluble protein abundance has commensurate effects on ion composition and cellular function across the circadian cycle. Thus, circadian regulation of the proteome impacts ion homeostasis with substantial consequences for the physiology of electrically active cells such as cardiomyocytes.


Assuntos
Fenômenos Fisiológicos Celulares , Ritmo Circadiano/fisiologia , Transporte de Íons/fisiologia , Osmose , Animais , Sistema Cardiovascular/patologia , Células Cultivadas , Cloretos/metabolismo , Fibroblastos , Homeostase , Pulmão , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Potássio/metabolismo , Proteoma , Sódio/metabolismo , Membro 2 da Família 12 de Carreador de Soluto/genética
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