scholarly journals Mieap forms membraneless organelles to compartmentalize and facilitate cardiolipin metabolism

2021 ◽  
Author(s):  
Naoki Ikari ◽  
Katsuko Honjo ◽  
Yoko Sagami ◽  
Yasuyuki Nakamura ◽  
Hirofumi Arakawa
Keyword(s):  
Quality Control ◽  
Enzymatic Reactions ◽  
Liquid Droplets ◽  
Inner Mitochondrial Membrane ◽  
Mitochondrial Quality Control ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Inducible Protein ◽  
Deficient Mice ◽  
Liquid Liquid Phase Separation

Abstract Liquid droplets function as membraneless organelles that compartmentalize and facilitate efficient biological reactions. They are formed by proteins with intrinsically disordered regions (IDRs) via liquid–liquid phase separation. Mieap/SPATA18, a p53-inducible protein, participates in suppression of colorectal tumors by promoting mitochondrial quality control. However, the regulatory mechanism involved remains unclear. Here, we report that Mieap is an IDR-containing protein that drives formation of liquid droplets in mitochondria. Mieap liquid droplets specifically phase separate the mitochondrial phospholipid, cardiolipin. Lipidomic analysis of cardiolipin suggests that Mieap promotes enzymatic reactions involved in cardiolipin metabolism, including biosynthesis and remodeling. Accordingly, four cardiolipin biosynthetic enzymes, TAMM41, PGS1, PTPMT1, and CRLS1, and two remodeling enzymes, PLA2G6 and TAZ, are phase-separated by Mieap liquid droplets. Mieap-deficient mice exhibit altered crista structure in mitochondria of various tissues, including brown fat, and tend to become obese. These results suggest that Mieap drives formation of membraneless organelles to compartmentalize and promote cardiolipin metabolism at the inner mitochondrial membrane, thus potentially contributing to mitochondrial quality control.

scholarly journals Mieap forms membraneless organelles to compartmentalize and facilitate cardiolipin metabolism

2020 ◽  
Author(s):  
Naoki Ikari ◽  
Katsuko Honjo ◽  
Yoko Sagami ◽  
Yasuyuki Nakamura ◽  
Hirofumi Arakawa
Keyword(s):  
Quality Control ◽  
Critical Role ◽  
Enzymatic Reactions ◽  
Liquid Droplets ◽  
Inner Mitochondrial Membrane ◽  
Mitochondrial Quality Control ◽  
Kidney Mitochondria ◽  
Intrinsically Disordered ◽  
Liver And Kidney ◽  
Inducible Protein

AbstractLiquid droplets function as membraneless organelles that compartmentalize and facilitate efficient biological reactions in cells. They are formed by proteins with an intrinsically disordered region(s) (IDR) via liquid–liquid phase separation. Mieap/SPATA18, a p53-inducible protein, plays a critical role in the suppression of human and murine colorectal tumors via mitochondrial quality control. However, the regulatory mechanism underlying this process remains unclear. Here, we report that Mieap is an IDR-containing protein that drives the formation of liquid droplets in the mitochondria. Mieap liquid droplets (MLDs) specifically phase separate the mitochondrial phospholipid cardiolipin. Lipidomic analysis of cardiolipin suggested that Mieap promotes enzymatic reactions involved in cardiolipin metabolism, including biosynthesis and remodeling. Accordingly, four cardiolipin biosynthesis enzymes, TAMM41, PGS1, PTPMT1, and CRLS1, and two remodeling enzymes, PLA2G6 and TAZ, are phase separated by MLDs. Mieap-deficient mice exhibited altered cristae structure in the liver and kidney mitochondria and a trend of obesity. These results suggest that Mieap drives the formation of membraneless organelles to compartmentalize and promotes cardiolipin metabolism at the inner mitochondrial membrane, thus playing a possible role in mitochondrial quality control.

scholarly journals Previously uncharacterized interactions between the folded and intrinsically disordered domains impart asymmetric effects on UBQLN2 phase separation

2021 ◽  
Author(s):  
Tongyin Zheng ◽  
Carlos A. Castañeda
Keyword(s):  
Quality Control ◽  
Phase Separation ◽  
Liquid Phase ◽  
Protein Quality ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Uba Domain ◽  
Phase Separation Behavior ◽  
Separation Behavior ◽  
Liquid Liquid Phase Separation

AbstractShuttle protein UBQLN2 functions in protein quality control (PQC) by binding to proteasomal receptors and ubiquitinated substrates via its N-terminal ubiquitin-like (UBL) and C-terminal ubiquitin-associated (UBA) domains, respectively. Between these two folded domains are intrinsically disordered STI1-I and STI1-II regions, connected by disordered linkers. The STI1 regions bind other components, such as HSP70, that are important to the PQC functions of UBQLN2. We recently determined that the STI1-II region enables UBQLN2 to undergo liquid-liquid phase separation (LLPS) to form liquid dropletsin vitroand biomolecular condensates in cells. However, how the interplay between the folded (UBL/UBA) domains and the intrinsically-disordered regions mediates phase separation is largely unknown. Using engineered domain deletion constructs, we found that removing the UBA domain inhibits UBQLN2 LLPS while removing the UBL domain enhances LLPS, suggesting that UBA and UBL domains contribute asymmetrically in modulating UBQLN2 LLPS. To explain these differential effects, we interrogated the interactions that involve the UBA and UBL domains across the entire UBQLN2 molecule using NMR spectroscopy. To our surprise, aside from well-studied canonical UBL:UBA interactions, there also exist moderate and weak interactions between the UBL and STI1-I/STI1-II domains, and between the UBA domain and the linker connecting the two STI1 regions, respectively. Our findings are essential for the understanding of both the molecular driving forces of UBQLN2 LLPS and the effects of ligand binding to UBL, UBA, or STI1 domains on the phase behavior and physiological functions of UBQLN2.Impact of Work StatementZheng and Castañeda show that interplay between the folded domains and intrinsically disordered regions regulates liquid-liquid phase separation behavior of UBQLN2, a protein quality control (PQC) shuttle protein. Despite their similar size, the folded UBL and UBA domains inhibit and promote phase separation, respectively, due to their previously uncharacterized, asymmetric interactions with the middle intrinsically-disordered region. These results strongly suggest that PQC components, including proteasomal receptors, are likely to modulate UBQLN2 phase separation behavior in cells.

scholarly journals The arrested state of processing bodies supports mRNA regulation in early development

2021 ◽  
Author(s):  
M. Sankaranarayanan ◽  
Ryan J. Emenecker ◽  
Marcus Jahnel ◽  
Irmela R. E. A. Trussina ◽  
Matt Wayland ◽  
...  
Keyword(s):  
Physical Properties ◽  
Electrostatic Interactions ◽  
Processing Bodies ◽  
P Bodies ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Dead Box ◽  
Premature Release ◽  
Liquid Liquid Phase Separation

ABSTRACTBiomolecular condensates that form via liquid-liquid phase separation can exhibit diverse physical states. Despite considerable progress, the relevance of condensate physical states forin vivobiological function remains limited. Here, we investigated the physical properties ofin vivoprocessing bodies (P bodies) and their impact on mRNA storage in matureDrosophilaoocytes. We show that the conserved DEAD-box RNA helicase Me31B forms P body condensates which adopt a less dynamic, arrested physical state. We demonstrate that structurally distinct proteins and hydrophobic and electrostatic interactions, together with RNA and intrinsically disordered regions, regulate the physical properties of P bodies. Finally, using live imaging, we show that the arrested state of P bodies is required to prevent the premature release ofbicoid(bcd) mRNA, a body axis determinant, and that P body dissolution leads tobcdrelease. Together, this work establishes a role for arrested states of biomolecular condensates in regulating cellular function in a developing organism.

scholarly journals A new class of disordered elements controls DNA replication through initiator self-assembly

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Matthew W Parker ◽  
Maren Bell ◽  
Mustafa Mir ◽  
Jonchee A Kao ◽  
Xavier Darzacq ◽  
...  
Keyword(s):  
Dna Replication ◽  
Self Assembly ◽  
Origin Recognition Complex ◽  
Replication Initiation ◽  
Linear Arrangement ◽  
New Class ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Liquid Liquid Phase Separation

The initiation of DNA replication in metazoans occurs at thousands of chromosomal sites known as origins. At each origin, the Origin Recognition Complex (ORC), Cdc6, and Cdt1 co-assemble to load the Mcm2-7 replicative helicase onto chromatin. Current replication models envisage a linear arrangement of isolated origins functioning autonomously; the extent of inter-origin organization and communication is unknown. Here, we report that the replication initiation machinery of D. melanogaster unexpectedly undergoes liquid-liquid phase separation (LLPS) upon binding DNA in vitro. We find that ORC, Cdc6, and Cdt1 contain intrinsically disordered regions (IDRs) that drive LLPS and constitute a new class of phase separating elements. Initiator IDRs are shown to regulate multiple functions, including chromosome recruitment, initiator-specific co-assembly, and Mcm2-7 loading. These data help explain how CDK activity controls replication initiation and suggest that replication programs are subject to higher-order levels of inter-origin organization.

scholarly journals Transcription Regulators and Membraneless Organelles Challenges to Investigate Them

2021 ◽  
Vol 22 (23) ◽  
pp. 12758
Author(s):  
Katarzyna Sołtys ◽  
Andrzej Ożyhar
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Liquid Phase Separation ◽  
Cellular Processes ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Experimental Systems ◽  
Transcription Regulators ◽  
Crucial Information ◽  
Liquid Liquid Phase Separation

Eukaryotic cells are composed of different bio-macromolecules that are divided into compartments called organelles providing optimal microenvironments for many cellular processes. A specific type of organelles is membraneless organelles. They are formed via a process called liquid–liquid phase separation that is driven by weak multivalent interactions between particular bio-macromolecules. In this review, we gather crucial information regarding different classes of transcription regulators with the propensity to undergo liquid–liquid phase separation and stress the role of intrinsically disordered regions in this phenomenon. We also discuss recently developed experimental systems for studying formation and properties of membraneless organelles.

scholarly journals CRIB effector disorder: exquisite function from chaos

2018 ◽  
Vol 46 (5) ◽  
pp. 1289-1302 ◽  
Author(s):  
Darerca Owen ◽  
Helen R. Mott
Keyword(s):  
Protein Binding ◽  
G Protein ◽  
Multiple Roles ◽  
Cellular Processes ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Binding Regions ◽  
Protein Binding Region ◽  
Basic Region ◽  
Liquid Liquid Phase Separation

The CRIB (Cdc42/Rac interactive binding) family of small G-protein effectors contain significant regions with intrinsic disorder. The G-protein-binding regions are contained within these intrinsically disordered regions. Most CRIB proteins also contain stretches of basic residues associated with their G-protein-binding regions. The basic region (BR) and G-protein-binding region together allow the CRIB effectors to bind to their cognate G-protein via a dock- and coalesce-binding mechanism. The BRs of these proteins take on multiple roles: steering G-protein binding, interacting with elements of the membrane and regulating intramolecular regulatory interactions. The ability of these regions of the CRIBs to undergo multivalent interactions and mediate charge neutralizations equips them with all the properties required to drive liquid–liquid phase separation and therefore to initiate and drive signalosome formation. It is only recently that the structural plasticity in these proteins is being appreciated as the driving force for these vital cellular processes.

scholarly journals Dynamic Formation of Liquid Droplets Triggered by Sequential Enzymatic Reactions

2020 ◽  
Author(s):  
Tomoto Ura ◽  
Shunsuke Tomita ◽  
Kentaro Shiraki
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Enzyme Reaction ◽  
Model System ◽  
Liquid Phase Separation ◽  
Enzymatic Reactions ◽  
Liquid Droplets ◽  
Dynamic Formation ◽  
Liquid Liquid Phase Separation

<p>A model system was developed that dynamically generates two different liquid droplets via liquid–liquid phase separation coupled with a sequential glycolytic reaction. The sequential two-enzyme reaction triggers the formation/dissolution of the liquid droplets. The droplets, in turn, compartmentalize each enzymatic step and generate feedback to accelerate the overall reaction.</p>

scholarly journals Intrinsic disorder codes for leaps of protein expression

2020 ◽  
Author(s):  
Chi-Ning Chuang ◽  
Tai-Ting Woo ◽  
Shih-Ying Tsai ◽  
Wan-Chen Li ◽  
Chia-Ling Chen ◽  
...  
Keyword(s):  
Quality Control ◽  
Protein Folding ◽  
Protein Expression ◽  
Posttranslational Modifications ◽  
Protein Quality ◽  
Intrinsic Disorder ◽  
Protein Homeostasis ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Regulate Protein

AbstractIntrinsically disordered regions (IDRs) are protein sequences lacking fixed or ordered three-dimensional structures. Many IDRs are endowed with important molecular functions such as physical interactions, posttranslational modifications or solubility enhancement. We reveal that several biologically important IDRs can act as N-terminal fusion carriers to promote target protein folding or protein quality control, thereby enhancing protein expression. This nanny function has a reasonably strong correlation with high S/T/Q/N amino acid content in IDRs and it is tunable (e.g., via phosphorylation) to regulate protein homeostasis. We propose a hypothesis that “N-terminal intrinsic disorder facilitates abundance” (NIDFA) to explain how some yeast proteins use their N-terminal IDRs (N-IDRs) to generate high levels of protein product. These N-IDRs are versatile toolkits for functional divergence in signaling and evolution.SignificanceDisorder within an otherwise well-structured protein is mostly found in intrinsically disordered regions (IDRs). IDRs can provide many advantages to proteins, including: (1) mediating protein-protein or protein-peptide interactions by adopting different conformations; (2) facilitating protein regulation via diverse posttranslational modifications; and (3) regulating the half-lives of proteins that have been targeted for proteasomal degradation. Here, we report that several biologically important IDRs in S. cerevisiae can act as N-terminal fusion carriers to promote target protein folding or protein quality control, thereby enhancing protein expression. We demonstrate by genetic and bioinformatic analyses that this nanny function is well correlated with high content of serine, threonine, glutamine and asparagine in IDRs and is tunable (e.g., via phosphorylation) to regulate protein homeostasis.

scholarly journals The (un)structural biology of biomolecular liquid-liquid phase separation using NMR spectroscopy

2020 ◽  
Vol 295 (8) ◽  
pp. 2375-2384 ◽  
Author(s):  
Anastasia C. Murthy ◽  
Nicolas L. Fawzi
Keyword(s):  
Phase Separation ◽  
Nmr Spectroscopy ◽  
Liquid Phase ◽  
Structural Biology ◽  
Liquid Phase Separation ◽  
Biological Processes ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Molecular Aspects ◽  
Liquid Liquid Phase Separation

Liquid-liquid phase separation (LLPS) of proteins and nucleic acids is a phenomenon that underlies membraneless compartmentalization of the cell. The underlying molecular interactions that underpin biomolecular LLPS have been of increased interest due to the importance of membraneless organelles in facilitating various biological processes and the disease association of several of the proteins that mediate LLPS. Proteins that are able to undergo LLPS often contain intrinsically disordered regions and remain dynamic in solution. Solution-state NMR spectroscopy has emerged as a leading structural technique to characterize protein LLPS due to the variety and specificity of information that can be obtained about intrinsically disordered sequences. This review discusses practical aspects of studying LLPS by NMR, summarizes recent work on the molecular aspects of LLPS of various protein systems, and discusses future opportunities for characterizing the molecular details of LLPS to modulate phase separation.

scholarly journals Multitiered and Cooperative Surveillance of Mitochondrial Phosphatidylserine Decarboxylase 1

2017 ◽  
Vol 37 (17) ◽  
Author(s):  
Oluwaseun B. Ogunbona ◽  
Ouma Onguka ◽  
Elizabeth Calzada ◽  
Steven M. Claypool
Keyword(s):  
Quality Control ◽  
Heat Exposure ◽  
Mitochondrial Pathway ◽  
Temperature Sensitive ◽  
Intermembrane Space ◽  
Control Mechanisms ◽  
Inner Mitochondrial Membrane ◽  
Mitochondrial Quality Control ◽  
Inner Membrane ◽  
Α Subunit

ABSTRACT Phosphatidylserine decarboxylase 1 (Psd1p), an ancient enzyme that converts phosphatidylserine to phosphatidylethanolamine in the inner mitochondrial membrane, must undergo an autocatalytic self-processing event to gain activity. Autocatalysis severs the protein into a large membrane-anchored β subunit that noncovalently associates with the small α subunit on the intermembrane space side of the inner membrane. Here, we determined that a temperature sensitive (ts) PSD1 allele is autocatalytically impaired and that its fidelity is closely monitored throughout its life cycle by multiple mitochondrial quality control proteases. Interestingly, the proteases involved in resolving misfolded Psd1ts vary depending on its autocatalytic status. Specifically, the degradation of a Psd1ts precursor unable to undergo autocatalysis requires the unprecedented cooperative and sequential actions of two inner membrane proteases, Oma1p and Yme1p. In contrast, upon heat exposure postautocatalysis, Psd1ts β subunits accumulate in protein aggregates that are resolved by Yme1p acting alone, while the released α subunit is degraded in parallel by an unidentified protease. Importantly, the stability of endogenous Psd1p is also influenced by Yme1p. We conclude that Psd1p, the key enzyme required for the mitochondrial pathway of phosphatidylethanolamine production, is closely monitored at several levels and by multiple mitochondrial quality control mechanisms present in the intermembrane space.

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