Establishment of heterochromatin in domain-size-dependent bursts

Methylation of histone H3K9 is a hallmark of epigenetic silencing in eukaryotes. Nucleosome modifications often rely on positive feedback where enzymes are recruited by modified nucleosomes. A combination of local and global feedbacks has been proposed to account for some dynamic properties of heterochromatin, but the range at which the global feedbacks operate and the exact mode of heterochromatin propagation are not known. We investigated these questions in fission yeast. Guided by mathematical modeling, we incrementally increased the size of the mating-type region and profiled heterochromatin establishment over time. We observed exponential decays in the proportion of cells with active reporters, with rates that decreased with domain size. Establishment periods varied from a few generations in wild type to >200 generations in the longest region examined, and highly correlated silencing of two reporters located outside the nucleation center was observed. On a chromatin level, this indicates that individual regions are silenced in sudden bursts. Mathematical modeling accounts for these bursts if heterochromatic nucleosomes facilitate a deacetylation or methylation reaction at long range, in a distance-independent manner. A likely effector of three-dimensional interactions is the evolutionarily conserved Swi6HP1 H3K9me reader, indicating the bursting behavior might be a general mode of heterochromatin propagation.

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Synthetic chromosome fusion: effects on genome structure and function

10.1101/381137 ◽

2018 ◽

Author(s):

Jingchuan Luo ◽

Guillaume Mercy ◽

Luis A. Vale-Silva ◽

Xiaoji Sun ◽

Neta Agmon ◽

...

Keyword(s):

Genome Structure ◽

Three Dimensional ◽

Wild Type ◽

3D Interaction ◽

Chromosome Fusion ◽

Size Dependent ◽

Varying Length ◽

And Function ◽

Chromosome I ◽

Wild Type Yeast

SUMMARYAs part of the Synthetic Yeast 2.0 (Sc2.0) project, we designed and synthesized synthetic chromosome I. The total length of synI is ~21.4% shorter than wild-type chromosome I, the smallest chromosome in Saccharomyces cerevisiae. SynI was designed for attachment to another synthetic chromosome due to concerns of potential instability and karyotype imbalance. We used a variation of a previously developed, robust CRISPR-Cas9 method to fuse chromosome I to other chromosome arms of varying length: chrIXR (84 kb), chrIIIR (202 kb) and chrIVR (1 Mb). All fusion chromosome strains grew like wild-type so we decided to attach synI to synIII. Through the investigation of three-dimensional structures of fusion chromosome strains, unexpected loops and twisted structures were formed in chrIII-I and chrIX-III-I fusion chromosomes, which depend on silencing protein Sir3. These results suggest a previously unappreciated 3D interaction between HMR and the adjacent telomere. We used these fusion chromosomes to show that axial element Red1 binding in meiosis is not strictly chromosome size dependent even though Red1 binding is enriched on the three smallest chromosomes in wild-type yeast, and we discovered an unexpected role for centromeres in Red1 binding patterns.

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p53 CRISPR Deletion Affects DNA Structure and Nuclear Architecture

Journal of Clinical Medicine ◽

10.3390/jcm9020598 ◽

2020 ◽

Vol 9 (2) ◽

pp. 598 ◽

Cited By ~ 2

Author(s):

Aline Rangel-Pozzo ◽

Samuel Booth ◽

Pak Lok Ivan Yu ◽

Madhurendra Singh ◽

Galina Selivanova ◽

...

Keyword(s):

Tumor Suppressor ◽

Three Dimensional ◽

Nuclear Architecture ◽

Super Resolution ◽

Dna Structure ◽

Tp53 Gene ◽

Binding Pocket ◽

Main Function ◽

Wild Type ◽

Independent Manner

The TP53 gene is a key tumor suppressor. Although the tumor suppressor p53 was one of the first to be characterized as a transcription factor, with its main function potentiated by its interaction with DNA, there are still many unresolved questions about its mechanism of action. Here, we demonstrate a novel role for p53 in the maintenance of nuclear architecture of cells. Using three-dimensional (3D) imaging and spectral karyotyping, as well as super resolution microscopy of DNA structure, we observe significant differences in 3D telomere signatures, DNA structure and DNA-poor spaces as well gains or losses of chromosomes, between normal and tumor cells with CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)-deleted or wild-type TP53. Additionally, treatment with Nutlin-3 results in differences in nuclear architecture of telomeres in wild-type but not in p53 knockout MCF-7 (Michigan Cancer Foundation-7) cells. Nutlin-3 binds to the p53-binding pocket of mouse double minute 2 (MDM2) and blocks the p53-MDM2 interaction. Moreover, we demonstrate that another p53 stabilizing small molecule, RITA (reactivation of p53 and induction of tumor cell apoptosis), also induces changes in 3D DNA structure, apparently in a p53 independent manner. These results implicate p53 activity in regulating nuclear organization and, additionally, highlight the divergent effects of the p53 targeting compounds Nutlin-3 and RITA.

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Fast Overlap Detection between Hard-Core Colloidal Cuboids and Spheres. The OCSI Algorithm

Algorithms ◽

10.3390/a14030072 ◽

2021 ◽

Vol 14 (3) ◽

pp. 72

Author(s):

Luca Tonti ◽

Alessandro Patti

Keyword(s):

Colloidal Particles ◽

Dynamic Properties ◽

Wide Spectrum ◽

Three Dimensional ◽

Hard Core ◽

Detection Algorithm ◽

Three Dimensional Objects ◽

Aggregation Behaviour ◽

Sphere Radius ◽

User Friendly

Collision between rigid three-dimensional objects is a very common modelling problem in a wide spectrum of scientific disciplines, including Computer Science and Physics. It spans from realistic animation of polyhedral shapes for computer vision to the description of thermodynamic and dynamic properties in simple and complex fluids. For instance, colloidal particles of especially exotic shapes are commonly modelled as hard-core objects, whose collision test is key to correctly determine their phase and aggregation behaviour. In this work, we propose the Oriented Cuboid Sphere Intersection (OCSI) algorithm to detect collisions between prolate or oblate cuboids and spheres. We investigate OCSI’s performance by bench-marking it against a number of algorithms commonly employed in computer graphics and colloidal science: Quick Rejection First (QRI), Quick Rejection Intertwined (QRF) and a vectorized version of the OBB-sphere collision detection algorithm that explicitly uses SIMD Streaming Extension (SSE) intrinsics, here referred to as SSE-intr. We observed that QRI and QRF significantly depend on the specific cuboid anisotropy and sphere radius, while SSE-intr and OCSI maintain their speed independently of the objects’ geometry. While OCSI and SSE-intr, both based on SIMD parallelization, show excellent and very similar performance, the former provides a more accessible coding and user-friendly implementation as it exploits OpenMP directives for automatic vectorization.

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Design, Fabrication, and Testing of a Novel 3D 3-Fingered Electrothermal Microgripper with Multiple Degrees of Freedom

Micromachines ◽

10.3390/mi12040444 ◽

2021 ◽

Vol 12 (4) ◽

pp. 444

Author(s):

Guoning Si ◽

Liangying Sun ◽

Zhuo Zhang ◽

Xuping Zhang

Keyword(s):

Degrees Of Freedom ◽

Dynamic Properties ◽

Three Dimensional ◽

Polyimide Film ◽

Zebrafish Embryos ◽

Multiple Degrees Of Freedom ◽

Electrothermal Actuator ◽

3D Space ◽

Pick And Place

This paper presents the design, fabrication, and testing of a novel three-dimensional (3D) three-fingered electrothermal microgripper with multiple degrees of freedom (multi DOFs). Each finger of the microgripper is composed of a V-shaped electrothermal actuator providing one DOF, and a 3D U-shaped electrothermal actuator offering two DOFs in the plane perpendicular to the movement of the V-shaped actuator. As a result, each finger possesses 3D mobilities with three DOFs. Each beam of the actuators is heated externally with the polyimide film. The durability of the polyimide film is tested under different voltages. The static and dynamic properties of the finger are also tested. Experiments show that not only can the microgripper pick and place microobjects, such as micro balls and even highly deformable zebrafish embryos, but can also rotate them in 3D space.

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Deciphering the Role of Residues Involved in Disorder-To-Order Transition Regions in Archaeal tRNA Methyltransferase 5

Genes ◽

10.3390/genes12030399 ◽

2021 ◽

Vol 12 (3) ◽

pp. 399

Author(s):

Ambuj Srivastava ◽

Dhanusha Yesudhas ◽

Shandar Ahmad ◽

M. Michael Gromiha

Keyword(s):

Three Dimensional ◽

Md Simulations ◽

Order Transition ◽

Free Form ◽

Wild Type ◽

Intrinsically Disordered ◽

Intrinsically Disordered Regions ◽

Type Complex ◽

In The Wild ◽

Trna Methyltransferase

tRNA methyltransferase 5 (Trm5) enzyme is an S-adenosyl methionine (AdoMet)-dependent methyltransferase which methylates the G37 nucleotide at the N1 atom of the tRNA. The free form of Trm5 enzyme has three intrinsically disordered regions, which are highly flexible and lack stable three-dimensional structures. These regions gain ordered structures upon the complex formation with tRNA, also called disorder-to-order transition (DOT) regions. In this study, we performed molecular dynamics (MD) simulations of archaeal Trm5 in free and complex forms and observed that the DOT residues are highly flexible in free proteins and become stable in complex structures. The energetic contributions show that DOT residues are important for stabilising the complex. The DOT1 and DOT2 are mainly observed to be important for stabilising the complex, while DOT3 is present near the active site to coordinate the interactions between methyl-donating ligands and G37 nucleotides. In addition, mutational studies on the Trm5 complex showed that the wild type is more stable than the G37A tRNA mutant complex. The loss of productive interactions upon G37A mutation drives the AdoMet ligand away from the 37th nucleotide, and Arg145 in DOT3 plays a crucial role in stabilising the ligand, as well as the G37 nucleotide, in the wild-type complex. Further, the overall energetic contribution calculated using MMPBSA corroborates that the wild-type complex has a better affinity between Trm5 and tRNA. Overall, our study reveals that targeting DOT regions for binding could improve the inhibition of Trm5.

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Functional Assessment of 12 Rare Allelic CYP2C9 Variants Identified in a Population of 4773 Japanese Individuals

Journal of Personalized Medicine ◽

10.3390/jpm11020094 ◽

2021 ◽

Vol 11 (2) ◽

pp. 94

Author(s):

Masaki Kumondai ◽

Akio Ito ◽

Evelyn Marie Gutiérrez Rico ◽

Eiji Hishinuma ◽

Akiko Ueda ◽

...

Keyword(s):

Enzymatic Activity ◽

Three Dimensional ◽

Warfarin Dose ◽

Catalytic Efficiency ◽

Therapeutic Window ◽

Wild Type ◽

Drug Metabolizing Enzyme ◽

Novel Variants ◽

Metabolizing Enzyme ◽

Almost All

Cytochrome P450 2C9 (CYP2C9) is an important drug-metabolizing enzyme that contributes to the metabolism of approximately 15% of clinically used drugs, including warfarin, which is known for its narrow therapeutic window. Interindividual differences in CYP2C9 enzymatic activity caused by CYP2C9 genetic polymorphisms lead to inconsistent treatment responses in patients. Thus, in this study, we characterized the functional differences in CYP2C9 wild-type (CYP2C9.1), CYP2C9.2, CYP2C9.3, and 12 rare novel variants identified in 4773 Japanese individuals. These CYP2C9 variants were heterologously expressed in 293FT cells, and the kinetic parameters (Km, kcat, Vmax, catalytic efficiency, and CLint) of (S)-warfarin 7-hydroxylation and tolbutamide 4-hydroxylation were estimated. From this analysis, almost all novel CYP2C9 variants showed significantly reduced or null enzymatic activity compared with that of the CYP2C9 wild-type. A strong correlation was found in catalytic efficiencies between (S)-warfarin 7-hydroxylation and tolbutamide 4-hydroxylation among all studied CYP2C9 variants. The causes of the observed perturbation in enzyme activity were evaluated by three-dimensional structural modeling. Our findings could clarify a part of discrepancies among genotype–phenotype associations based on the novel CYP2C9 rare allelic variants and could, therefore, improve personalized medicine, including the selection of the appropriate warfarin dose.

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Structural analysis of mutations in the Drosophila beta 2-tubulin isoform reveals regions in the beta-tubulin molecular required for general and for tissue-specific microtubule functions.

Genetics ◽

10.1093/genetics/139.1.267 ◽

1995 ◽

Vol 139 (1) ◽

pp. 267-286 ◽

Cited By ~ 2

Author(s):

J D Fackenthal ◽

J A Hutchens ◽

F R Turner ◽

E C Raff

Keyword(s):

Amino Acid ◽

Three Dimensional ◽

Variable Region ◽

Internal Variable ◽

Microtubule Assembly ◽

Dimensional Structure ◽

Wild Type ◽

Beta Tubulin ◽

Assembly Kinetics ◽

Beta 2

Abstract We have determined the lesions in a number of mutant alleles of beta Tub85D, the gene that encodes the testis-specific beta 2-tubulin isoform in Drosophila melanogaster. Mutations responsible for different classes of functional phenotypes are distributed throughout the beta 2-tubulin molecule. There is a telling correlation between the degree of phylogenetic conservation of the altered residues and the number of different microtubule categories disrupted by the lesions. The majority of lesions occur at positions that are evolutionarily highly conserved in all beta-tubulins; these lesions disrupt general functions common to multiple classes of microtubules. However, a single allele B2t6 contains an amino acid substitution within an internal cluster of variable amino acids that has been identified as an isotype-defining domain in vertebrate beta-tubulins. Correspondingly, B2t6 disrupts only a subset of microtubule functions, resulting in misspecification of the morphology of the doublet microtubules of the sperm tail axoneme. We previously demonstrated that beta 3, a developmentally regulated Drosophila beta-tubulin isoform, confers the same restricted morphological phenotype in a dominant way when it is coexpressed in the testis with wild-type beta 2-tubulin. We show here by complementation analysis that beta 3 and the B2t6 product disrupt a common aspect of microtubule assembly. We therefore conclude that the amino acid sequence of the beta 2-tubulin internal variable region is required for generation of correct axoneme morphology but not for general microtubule functions. As we have previously reported, the beta 2-tubulin carboxy terminal isotype-defining domain is required for suprastructural organization of the axoneme. We demonstrate here that the beta 2 variant lacking the carboxy terminus and the B2t6 variant complement each other for mild-to-moderate meiotic defects but do not complement for proper axonemal morphology. Our results are consistent with the hypothesis drawn from comparisons of vertebrate beta-tubulins that the two isotype-defining domains interact in a three-dimensional structure in wild-type beta-tubulins. We propose that the integrity of this structure in the Drosophila testis beta 2-tubulin isoform is required for proper axoneme assembly but not necessarily for general microtubule functions. On the basis of our observations we present a model for regulation of axoneme microtubule morphology as a function of tubulin assembly kinetics.

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Three-dimensional mathematical modeling of local scour

Journal of Applied Mechanics ◽

10.2208/journalam.8.803 ◽

2005 ◽

Vol 8 ◽

pp. 803-812 ◽

Cited By ~ 6

Author(s):

Hao ZHANG ◽

Hajime NAKAGAWA ◽

Taisuke ISHIGAKI ◽

Yasunori MUTO ◽

Yasuyuki BABA

Keyword(s):

Mathematical Modeling ◽

Three Dimensional ◽

Local Scour

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Substitution of murine ferrochelatase glutamate-287 with glutamine or alanine leads to porphyrin substrate-bound variants

Biochemical Journal ◽

10.1042/bj3560217 ◽

2001 ◽

Vol 356 (1) ◽

pp. 217-222 ◽

Cited By ~ 5

Author(s):

Ricardo FRANCO ◽

Alice S. PEREIRA ◽

Pedro TAVARES ◽

Arianna MANGRAVITA ◽

Michael J. BARBER ◽

...

Keyword(s):

Absorption Spectra ◽

Catalytic Mechanism ◽

Protoporphyrin Ix ◽

Three Dimensional ◽

Iron Chelation ◽

Enzymic Activity ◽

Dimensional Structure ◽

Wild Type ◽

Wild Type Enzyme ◽

Flow Experiments

Ferrochelatase (EC 4.99.1.1) is the terminal enzyme of the haem biosynthetic pathway and catalyses iron chelation into the protoporphyrin IX ring. Glutamate-287 (E287) of murine mature ferrochelatase is a conserved residue in all known sequences of ferrochelatase, is present at the active site of the enzyme, as inferred from the Bacillus subtilis ferrochelatase three-dimensional structure, and is critical for enzyme activity. Substitution of E287 with either glutamine (Q) or alanine (A) yielded variants with lower enzymic activity than that of the wild-type ferrochelatase and with different absorption spectra from the wild-type enzyme. In contrast to the wild-type enzyme, the absorption spectra of the variants indicate that these enzymes, as purified, contain protoporphyrin IX. Identification and quantification of the porphyrin bound to the E287-directed variants indicate that approx. 80% of the total porphyrin corresponds to protoporphyrin IX. Significantly, rapid stopped-flow experiments of the E287A and E287Q variants demonstrate that reaction with Zn2+ results in the formation of bound Zn-protoporphyrin IX, indicating that the endogenously bound protoporphyrin IX can be used as a substrate. Taken together, these findings suggest that the structural strain imposed by ferrochelatase on the porphyrin substrate as a critical step in the enzyme catalytic mechanism is also accomplished by the E287A and E287Q variants, but without the release of the product. Thus E287 in murine ferrochelatase appears to be critical for the catalytic process by controlling the release of the product.

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Multi-Objective Optimization of a Microturbine Compact Recuperator

Journal of Engineering for Gas Turbines and Power ◽

10.1115/1.2836479 ◽

2008 ◽

Vol 130 (3) ◽

Cited By ~ 9

Author(s):

Diego Micheli ◽

Valentino Pediroda ◽

Stefano Pieri

Keyword(s):

Heat Transfer ◽

Three Dimensional ◽

Numerical Procedure ◽

Design Procedure ◽

Domain Size ◽

Multidisciplinary Optimization ◽

Computational Domain ◽

Surface Geometry ◽

Multi Objective ◽

Flux Boundary Conditions

An automatic approach for the multi-objective shape optimization of microgas turbine heat exchangers is presented. According to the concept of multidisciplinary optimization, the methodology integrates a CAD parametric model of the heat transfer surfaces, a three-dimensional meshing tool, and a CFD solver, all managed by a design optimization platform. The repetitive pattern of the surface geometry has been exploited to reduce the computational domain size, and the constant flux boundary conditions have been imposed to better suit the real operative conditions. A new approach that couples cold and warm fluids in a periodic unitary cell is introduced. The effectiveness of the numerical procedure was verified comparing the numerical results with available literature data. The optimization objectives are maximizing the heat transfer rate and minimizing both friction factor and heat transfer surface. The paper presents the results of the optimization of a 50kWMGT recuperator. The design procedure can be effectively extended and applied to any industrial heat exchanger application.

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