scholarly journals Granule regulation by phase separation during Drosophila oogenesis

2020 ◽  
Vol 4 (3) ◽  
pp. 355-364
Author(s):  
M. Sankaranarayanan ◽  
Timothy T. Weil
Keyword(s):  
Phase Separation ◽  
Translational Control ◽  
Protein Complexes ◽  
Physiological Role ◽  
Drosophila Oogenesis ◽  
And Function ◽  
Rna Protein Complexes ◽  
Liquid Liquid Phase Separation

Drosophila eggs are highly polarised cells that use RNA–protein complexes to regulate storage and translational control of maternal RNAs. Ribonucleoprotein granules are a class of biological condensates that form predominantly by intracellular phase separation. Despite extensive in vitro studies testing the physical principles regulating condensates, how phase separation translates to biological function remains largely unanswered. In this perspective, we discuss granules in Drosophila oogenesis as a model system for investigating the physiological role of phase separation. We review key maternal granules and their properties while highlighting ribonucleoprotein phase separation behaviours observed during development. Finally, we discuss how concepts and models from liquid–liquid phase separation could be used to test mechanisms underlying granule assembly, regulation and function in Drosophila oogenesis.

scholarly journals Membrane-associated phase separation: organization and function emerge from a two-dimensional milieu

2021 ◽  
Author(s):  
Jonathon A Ditlev
Keyword(s):  
Phase Separation ◽  
Cell Biology ◽  
Cellular Environment ◽  
Condensate Formation ◽  
Associated Proteins ◽  
Available Technology ◽  
And Function ◽  
Surrounding Membrane

Abstract Liquid‒liquid phase separation (LLPS) of biomolecules has emerged as an important mechanism that contributes to cellular organization. Phase separated biomolecular condensates, or membrane-less organelles, are compartments composed of specific biomolecules without a surrounding membrane in the nucleus and cytoplasm. LLPS also occurs at membranes, where both lipids and membrane-associated proteins can de-mix to form phase separated compartments. Investigation of these membrane-associated condensates using in vitro biochemical reconstitution and cell biology has provided key insights into the role of phase separation in membrane domain formation and function. However, these studies have generally been limited by available technology to study LLPS on model membranes and the complex cellular environment that regulates condensate formation, composition, and function. Here, I briefly review our current understanding of membrane-associated condensates, establish why LLPS can be advantageous for certain membrane-associated condensates, and offer a perspective for how these condensates may be studied in the future.

scholarly journals Liquid–Liquid Phase Separation in Crowded Environments

2020 ◽  
Vol 21 (16) ◽  
pp. 5908 ◽  
Author(s):  
Alain A. M. André ◽  
Evan Spruijt
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Macromolecular Crowding ◽  
Liquid Phase Separation ◽  
The Impact ◽  
Crowded Environments ◽  
Liquid Liquid Phase Separation ◽  
In Cells

Biomolecular condensates play a key role in organizing cellular fluids such as the cytoplasm and nucleoplasm. Most of these non-membranous organelles show liquid-like properties both in cells and when studied in vitro through liquid–liquid phase separation (LLPS) of purified proteins. In general, LLPS of proteins is known to be sensitive to variations in pH, temperature and ionic strength, but the role of crowding remains underappreciated. Several decades of research have shown that macromolecular crowding can have profound effects on protein interactions, folding and aggregation, and it must, by extension, also impact LLPS. However, the precise role of crowding in LLPS is far from trivial, as most condensate components have a disordered nature and exhibit multiple weak attractive interactions. Here, we discuss which factors determine the scope of LLPS in crowded environments, and we review the evidence for the impact of macromolecular crowding on phase boundaries, partitioning behavior and condensate properties. Based on a comparison of both in vivo and in vitro LLPS studies, we propose that phase separation in cells does not solely rely on attractive interactions, but shows important similarities to segregative phase separation.

scholarly journals Mice Deficient in the Axonemal Protein Tektin-t Exhibit Male Infertility and Immotile-Cilium Syndrome Due to Impaired Inner Arm Dynein Function

2004 ◽  
Vol 24 (18) ◽  
pp. 7958-7964 ◽  
Author(s):  
Hiromitsu Tanaka ◽  
Naoko Iguchi ◽  
Yoshiro Toyama ◽  
Kouichi Kitamura ◽  
Tohru Takahashi ◽  
...  
Keyword(s):  
Physiological Role ◽  
Electron Microscopic Examination ◽  
Electron Microscopic ◽  
Vitro Fertilization ◽  
Homozygous Mutant ◽  
Causal Genes ◽  
And Function ◽  
Ciliary Dyskinesia

ABSTRACT The haploid germ cell-specific Tektin-t protein is a member of the Tektin family of proteins that form filaments in flagellar, ciliary, and axonemal microtubules. To investigate the physiological role of Tektin-t, we generated mice with a mutation in the tektin-t gene. The homozygous mutant males were infertile, while the females were fully fertile. Sperm morphology and function were abnormal, with frequent bending of the sperm flagella and marked defects in motility. In vitro fertilization assays showed that the defective spermatozoa were able to fertilize eggs. Electron microscopic examination showed that the dynein inner arm structure was disrupted in the sperm flagella of tektin-t-deficient mice. Furthermore, homozygous mutant mice had functionally defective tracheal cilia, as evidenced by altered dynein arm morphology. These results indicate that Tektin-t participates in dynein inner arm formation or attachment and that the loss of Tektin-t results in impaired motility of both flagella and cilia. Therefore, the tektin-t gene is one of the causal genes for immotile-cilium syndrome/primary ciliary dyskinesia.

scholarly journals TDP-43 α-helical structure tunes liquid–liquid phase separation and function

2020 ◽  
Vol 117 (11) ◽  
pp. 5883-5894 ◽  
Author(s):  
Alexander E. Conicella ◽  
Gregory L. Dignon ◽  
Gül H. Zerze ◽  
Hermann Broder Schmidt ◽  
Alexandra M. D’Ordine ◽  
...  
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Structural Changes ◽  
Rna Binding ◽  
Helical Structure ◽  
Liquid Phase Separation ◽  
Control Assembly ◽  
And Function ◽  
Liquid Liquid Phase Separation

Liquid–liquid phase separation (LLPS) is involved in the formation of membraneless organelles (MLOs) associated with RNA processing. The RNA-binding protein TDP-43 is present in several MLOs, undergoes LLPS, and has been linked to the pathogenesis of amyotrophic lateral sclerosis (ALS). While some ALS-associated mutations in TDP-43 disrupt self-interaction and function, here we show that designed single mutations can enhance TDP-43 assembly and function via modulating helical structure. Using molecular simulation and NMR spectroscopy, we observe large structural changes upon dimerization of TDP-43. Two conserved glycine residues (G335 and G338) are potent inhibitors of helical extension and helix–helix interaction, which are removed in part by variants at these positions, including the ALS-associated G335D. Substitution to helix-enhancing alanine at either of these positions dramatically enhances phase separation in vitro and decreases fluidity of phase-separated TDP-43 reporter compartments in cells. Furthermore, G335A increases TDP-43 splicing function in a minigene assay. Therefore, the TDP-43 helical region serves as a short but uniquely tunable module where application of biophysical principles can precisely control assembly and function in cellular and synthetic biology applications of LLPS.

scholarly journals HANP1/H1T2, a Novel Histone H1-Like Protein Involved in Nuclear Formation and Sperm Fertility

2005 ◽  
Vol 25 (16) ◽  
pp. 7107-7119 ◽  
Author(s):  
Hiromitsu Tanaka ◽  
Naoko Iguchi ◽  
Ayako Isotani ◽  
Kouichi Kitamura ◽  
Yoshiro Toyama ◽  
...  
Keyword(s):  
Biochemical Analysis ◽  
Nuclear Protein ◽  
Physiological Role ◽  
Histone H1 ◽  
Vitro Fertilization ◽  
Rich Domain ◽  
And Function ◽  
Specific Nuclear Protein

ABSTRACT We cloned a testis-specific cDNA from mice that encodes a histone H1-like, haploid germ cell-specific nuclear protein designated HANP1/H1T2. The HANP1/H1T2 protein was specifically localized to the nuclei of murine spermatids during differentiation steps 5 to 13 but not to the nuclei of mature sperm. HANP1/H1T2 contains an arginine-serine-rich domain and an ATP/GTP binding site, and it binds to DNA, ATP, and protamine. To investigate the physiological role of HANP1/H1T2, we generated Hanp1/H1T2-disrupted mutant mice. Homozygous Hanp1/H1T2 mutant males were infertile, but females were fertile. Although a substantial number of sperm were recovered from the epididymides, their shape and function were abnormal. During sperm morphogenesis, the formation of nuclei was disturbed and protamine-1 and -2 were only weakly detectable in the nuclei. The chromatin packaging was aberrant, as demonstrated by electron microscopy and biochemical analysis. The mutant sperm exhibited deficient motility and were not competent to fertilize eggs under in vitro fertilization conditions; however, they were capable of fertilizing eggs via intracytoplasmic sperm injection that resulted in the birth of healthy progeny. Thus, we found that HANP1/H1T2 is essential for nuclear formation in functional spermatozoa and is specifically involved in the replacement of histones with protamines during spermiogenesis. At the time of submission of the manuscript, we found an independent publication by Martianov et al. (I. Martianov, S. Brancorsini, R. Catena, A. Gansmuller, N. Kotaja, M. Parvinen, P. Sassone-Corsi, and I. Davidson, Proc. Natl. Acad. Sci. USA 102:2808-2813, 2005) that reported similar results.

scholarly journals Translational inhibition and phase separation primes the epigenetic silencing of transposons

2020 ◽  
Author(s):  
Eun Yu Kim ◽  
Ling Wang ◽  
Zhen Lei ◽  
Hui Li ◽  
Wenwen Fan ◽  
...  
Keyword(s):  
Phase Separation ◽  
Epigenetic Silencing ◽  
Regulatory Mechanisms ◽  
Genome Integrity ◽  
Liquid Droplets ◽  
Ribosome Stalling ◽  
Insight Into ◽  
Liquid Liquid Phase Separation

AbstractTransposons are mobile DNAs that can cause fatal mutations. To counteract these genome invaders, the host genomes deploy small interfering (si) RNAs to initiate and establish the epigenetic silencing. However, the regulatory mechanisms for the selective recognition of transposons by the host genomes remain still elusive. Here we show that plant transposon RNAs undergo frequent ribosome stalling caused by their inherently unfavourable codon sequence usage. The ribosome stalling then causes the RNA truncation and the localization to siRNA bodies, which are both critical prerequisites for the siRNA processing. In addition, SGS3, the key protein in the siRNA biogenesis pathway, forms liquid droplets in vitro through its prion-like domains implicating the role of liquid-liquid phase separation in the formation of the siRNA bodies. Our study provides a novel insight into the regulatory mechanisms for the recognition of invasive genetic elements which is essential for the maintenance of genome integrity.

scholarly journals LncRNAs: Architectural Scaffolds or More Potential Roles in Phase Separation

2021 ◽  
Vol 12 ◽  
Author(s):  
Jie Luo ◽  
Lei Qu ◽  
Feiran Gao ◽  
Jun Lin ◽  
Jian Liu ◽  
...  
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Biological Activities ◽  
Disease Diagnosis ◽  
Liquid Phase Separation ◽  
Diagnosis And Treatment ◽  
Biological Processes ◽  
And Function ◽  
Liquid Liquid Phase Separation

Biomolecules specifically aggregate in the cytoplasm and nucleus, driving liquid-liquid phase separation (LLPS) formation and diverse biological processes. Extensive studies have focused on revealing multiple functional membraneless organelles in both the nucleus and cytoplasm. Condensation compositions of LLPS, such as proteins and RNAs affecting the formation of phase separation, have been gradually unveiled. LncRNAs possessing abundant second structures usually promote phase separation formation by providing architectural scaffolds for diverse RNAs and proteins interaction in both the nucleus and cytoplasm. Beyond scaffolds, lncRNAs may possess more diverse functions, such as functioning as enhancer RNAs or buffers. In this review, we summarized current studies on the function of phase separation and its related lncRNAs, mainly in the nucleus. This review will facilitate our understanding of the formation and function of phase separation and the role of lncRNAs in these processes and related biological activities. A deeper understanding of the formation and maintaining of phase separation will be beneficial for disease diagnosis and treatment.

scholarly journals Role of protein conformation and weak interactions on γ-gliadin liquid-liquid phase separation

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Line Sahli ◽  
Denis Renard ◽  
Véronique Solé-Jamault ◽  
Alexandre Giuliani ◽  
Adeline Boire
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Electrostatic Interactions ◽  
Storage Proteins ◽  
Liquid Phase Separation ◽  
Water Mixture ◽  
Terminal Domain ◽  
Liquid Liquid Phase Separation

Abstract Wheat storage proteins, gliadins, were found to form in vitro condensates in 55% ethanol/water mixture by decreasing temperature. The possible role of this liquid-liquid phase separation (LLPS) process on the in vivo gliadins storage is elusive and remains to be explored. Here we use γ-gliadin as a model of wheat proteins to probe gliadins behavior in conditions near physiological conditions. Bioinformatic analyses suggest that γ-gliadin is a hybrid protein with N-terminal domain predicted to be disordered and C-terminal domain predicted to be ordered. Spectroscopic data highlight the disordered nature of γ-gliadin. We developed an in vitro approach consisting to first solubilize γ-gliadin in 55% ethanol (v/v) and to progressively decrease ethanol ratio in favor of increased aqueous solution. Our results show the ability of γ-gliadin to self-assemble into dynamic droplets through LLPS, with saturation concentrations ranging from 25.9 µM ± 0.85 µM (35% ethanol (v/v)) to 3.8 µM ± 0.1 µM (0% ethanol (v/v)). We demonstrate the importance of the predicted ordered C-terminal domain of γ-gliadin in the LLPS by highlighting the protein condensates transition from a liquid to a solid state under reducing conditions. We demonstrate by increasing ionic strength the role displayed by electrostatic interactions in the phase separation. We also show the importance of hydrogen bonds in this process. Finally, we discuss the importance of gliadins condensates in their accumulation and storage in the wheat seed.

scholarly journals TDP-43 α-helical structure tunes liquid-liquid phase separation and function

2019 ◽  
Author(s):  
Alexander E. Conicella ◽  
Gregory L. Dignon ◽  
Gül H. Zerze ◽  
Hermann Broder Schmidt ◽  
Alexandra M. D’Ordine ◽  
...  
Keyword(s):  
Phase Separation ◽  
Liquid Phase ◽  
Structural Changes ◽  
Helical Structure ◽  
Liquid Phase Separation ◽  
Gene Assay ◽  
And Function ◽  
Lateral Sclerosis ◽  
Liquid Liquid Phase Separation

AbstractLiquid-liquid phase separation (LLPS) is involved in the formation of membraneless organelles (MLOs) associated with RNA processing. Present in several MLOs, TDP-43 undergoes LLPS and is linked to the pathogenesis of amyotrophic lateral sclerosis (ALS). While some disease variants of TDP-43 disrupt self-interaction and function, here we show that designed single mutations can enhance TDP-43 assembly and function via modulating helical structure. Using molecular simulation and NMR spectroscopy, we observe large structural changes in a dimeric TDP-43. Two conserved glycine residues (G335 and G338) are potent inhibitors of helical extension and helix-helix interaction, which are removed in part by variants including the ALS-associated G335D. Substitution to helix-enhancing alanine at either of these positions dramatically enhances phase separation in vitro and decreases fluidity of phase separated TDP-43 reporter compartments in cells. Furthermore, G335A increases TDP-43 splicing function in a mini-gene assay. Therefore, TDP-43 helical region serves as a short but uniquely tunable module that shows promise as for controlling assembly and function in cellular and synthetic biology applications of LLPS.

Role of the 807 C/T Polymorphism of the α2 Gene in Platelet GP Ia Collagen Receptor Expression and Function

1999 ◽  
Vol 81 (06) ◽  
pp. 951-956 ◽  
Author(s):  
J. Corral ◽  
R. González-Conejero ◽  
J. Rivera ◽  
F. Ortuño ◽  
P. Aparicio ◽  
...  
Keyword(s):  
Venous Thrombosis ◽  
Cell Types ◽  
Receptor Expression ◽  
Case Control Studies ◽  
Platelet Surface ◽  
Vitro Analysis ◽  
And Function ◽  
In Vitro Analysis

SummaryThe variability of the platelet GP Ia/IIa density has been associated with the 807 C/T polymorphism (Phe 224) of the GP Ia gene in American Caucasian population. We have investigated the genotype and allelic frequencies of this polymorphism in Spanish Caucasians. The T allele was found in 35% of the 284 blood donors analyzed. We confirmed in 159 healthy subjects a significant association between the 807 C/T polymorphism and the platelet GP Ia density. The T allele correlated with high number of GP Ia molecules on platelet surface. In addition, we observed a similar association of this polymorphism with the expression of this protein in other blood cell types. The platelet responsiveness to collagen was determined by “in vitro” analysis of the platelet activation and aggregation response. We found no significant differences in these functional platelet parameters according to the 807 C/T genotype. Finally, results from 3 case/control studies involving 302 consecutive patients (101 with coronary heart disease, 104 with cerebrovascular disease and 97 with deep venous thrombosis) determined that the 807 C/T polymorphism of the GP Ia gene does not represent a risk factor for arterial or venous thrombosis.

Sign in / Sign up

Export Citation Format

Share Document