The Biochemistry and Evolution of the Dinoflagellate Nucleus.

Dinoflagellates are known to possess a highly aberrant nucleus-the so-called dinokaryon-that exhibits a multitude of exceptional biological features. These include: (1) Permanently condensed chromosomes; (2) DNA in a cholesteric liquid crystalline state, (3) extremely large DNA content (up to 200 pg); and, perhaps most strikingly, (4) a deficit of histones-the canonical building blocks of all eukaryotic chromatin. Dinoflagellates belong to the Alveolata clade (dinoflagellates, apicomplexans, and ciliates) and, therefore, the biological oddities observed in dinoflagellate nuclei are derived character states. Understanding the sequence of changes that led to the dinokaryon has been difficult in the past with poor resolution of dinoflagellate phylogeny. Moreover, lack of knowledge of their molecular composition has constrained our understanding of the molecular properties of these derived nuclei. However, recent advances in the resolution of the phylogeny of dinoflagellates, particularly of the early branching taxa; the realization that divergent histone genes are present; and the discovery of dinoflagellate-specific nuclear proteins that were acquired early in dinoflagellate evolution have all thrown new light nature and evolution of the dinokaryon

High resolution imaging reveals heterogeneity in chromatin states between cells that is not inherited through cell division

BACKGROUND: Genomes of eukaryotes exist as chromatin, and it is known that different chromatin states can influence gene regulation. Chromatin is not a static structure, but is known to be dynamic and vary between cells. In order to monitor the organisation of chromatin in live cells we have engineered fluorescent fusion proteins which recognize specific operator sequences to tag pairs of syntenic gene loci. The separation of these loci was then tracked in three dimensions over time using fluorescence microscopy. RESULTS: We established a work flow for measuring the distance between two fluorescently tagged, syntenic gene loci with a mean measurement error of 63 nm. In general, physical separation was observed to increase with increasing genomic separations. However, the extent to which chromatin is compressed varies for different genomic regions. No correlation was observed between compaction and the distribution of chromatin markers from genomic datasets or with contacts identified using capture based approaches. Variation in spatial separation was also observed within cells over time and between cells. Differences in the conformation of individual loci can persist for minutes in individual cells. Separation of reporter loci was found to be similar in related and unrelated daughter cell pairs. CONCLUSIONS: The directly observed physical separation of reporter loci in live cells is highly dynamic both over time and from cell to cell. However, consistent differences in separation are observed over some chromosomal regions that do not correlate with factors known to influence chromatin states. We conclude that as yet unidentified parameters influence chromatin configuration. We also find that while heterogeneity in chromatin states can be maintained for minutes between cells, it is not inherited through cell division. This may contribute to cell-to-cell transcriptional heterogeneity. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12860-016-0111-y) contains supplementary material, which is available to authorized users

Exploring the syntax-phonology interface in Arabic

Despite an abundance of research on Arabic syntax and phonology as separate domains, there is as yet relatively little research at the syntax-phonology interface in Arabic. This paper begins by providing an overview of what we know so far, in an effort to identify reasons for the lack of work at the interface to date. The paper then presents a review of prior work on the syntax-phonology mapping in Egyptian Arabic (EA) - set in the context of developments in the wider syntax-phonology literature - in order to show that interface work requires expertise in both phonetics/phonology and syntax. Some early results are then presented from a pilot study which compares for the first time the basic syntax-phonology mapping patterns in two dialects of Arabic - EA, and Jordanian Arabic (JA) - and explores whether dialect-internal, inter-speaker variation, previously observed in EA, is also found in JA

Publisher Correction: Genetic tool development in marine protists: emerging model organisms for experimental cell biology.

An amendment to this paper has been published and can be accessed via a link at the top of the paper

Genetic tool development in marine protists: emerging model organisms for experimental cell biology

Abstract: Diverse microbial ecosystems underpin life in the sea. Among these microbes are many unicellular eukaryotes that span the diversity of the eukaryotic tree of life. However, genetic tractability has been limited to a few species, which do not represent eukaryotic diversity or environmentally relevant taxa. Here, we report on the development of genetic tools in a range of protists primarily from marine environments. We present evidence for foreign DNA delivery and expression in 13 species never before transformed and for advancement of tools for eight other species, as well as potential reasons for why transformation of yet another 17 species tested was not achieved. Our resource in genetic manipulation will provide insights into the ancestral eukaryotic lifeforms, general eukaryote cell biology, protein diversification and the evolution of cellular pathways

Rôle de l'organisation du chromosome III dans la directionalité du mating-type switch chez S. cerevisiae

La recombinaison homologue est un processus essentiel dans l'évolution des espèces, pourtant les mécanismes qui régulent cet événement sont encore mal connus. Une idée émergente est que l'architecture nucléaire et la conformation des chromosomes influents sur la recombinaison en favorisant la rencontre entre séquences homologues. Le changement de type sexuel chez la levure S.cerevisiae constitue un excellent modèle pour explorer ce mécanisme. En effet, le changement du type sexuel chez la levure est directionnel c'est-à-dire que suite à une induction d'une cassure double brin (DSB) au niveau du locus MAT par l'endonuclease HO, la cellule répare cette cassure par conversion génique en utilisant un des deux loci HMLa ou HMRa qui se retrouve à l'extrémité de chaque bras du chromosome III. Le locus HMLa sert de donneur d'information si la cellule est de type a et le locus HMRa est utilise lorsque la cellule est a. Nous avons étudiés le rôle de l'organisation nucléaire de ces trois loci ainsi que leur dynamique dans le choix du donneur par microscopie à fluorescence sur des cellules vivantes en utilisant les deux systèmes LacO et TetO ainsi qu'un troisième operator le λO que j'ai adapté pour l'usage chez la levure. Nous avons montré que le chromosome III adopte une organisation mating-type dépendante. Cette organisation spécifique se résume à la formation d'une boucle par l'interaction du RE avec la région centromérique. Cette organisation n'implique pas des facteurs de la compaction de la chromatine comme Asf1 ni des facteurs de mise en silence et d'ancrage des télomères comme Sir4 mais implique Fkh1. De plus, nous pensons que cette organisation est responsable à la relocalisation de HML dans les cellules a dans un endroit qui rend cette séquence recombinogène. En perspectives, la disponibilité de trois FROS ainsi qu'un programme capable d'analyser la position en 3D de trois loci de différentes couleurs, va permettre leur application dans l'étude de l'organisation nucléaire dans d'autres mécanismes cellulaires.Epigenetic regulation of gene expression involves chromatin compaction and organization of genomic loci in the nuclear volume. However, little is known about the influence of chromatin structure and dynamics in recombination. Haploid S.cerevisiae can change mating-type at every cell cycle. During this process, two transcriptionally silent haploid mating-type loci, HMLa and HMRa, located at the opposite ends of chromosome III, are required. Conversion of the mating-type begins with a site-specific cut by the HO endonuclease at the transcribed MAT locus located near the center of the chromosome. The resulting DSB is then repaired by homologous recombination using either HMLa or HMRa as a template. MAT a-cells use HMLa as a template in 85% of the cells whereas a-cells prefer to use HMRa as a template in 90% of the cells. We developed a new fluorescent repressor operator system (FROS) which relies on the transgenic expression of the bacterial λ repressor fused to a fluorescent protein which can bind to its respective λ operator DNA sequence integrated as a short multicopy tandem array in the yeast genome. Combined with the existing FROS based on LacO and TetO it now allows to visualize three specific genomic sites simultaneously. This triple labelling strategy was applied to study the choreography of the three mating-type loci in Saccharomyces cerevisiae. During this study, we uncovered a mating-type specific organization of the chromosome III in S.cerevisiae. This chromosome organization does not seem to be affected after the abolishment of the chromatin fiber compaction (asf1 mutant) or the silencing ans anchoring of telomeres (sir4 mutant) but is affected following FKH1 deletion. We propose a model in which mating-type specific conformation of chromosome III correlates with the efficiency and the outcome of the gene conversion event between the HM and MAT loci. In fact, the left arm seems to form a loop due to transient interaction of the Recombination Enhancer with centromere proximal sequences. This loop positions the HML region at a strategic region to render it highly recombinogenic. In alpha-cells this specific organization is absent leading to a sequestration of HMLa away from MAT. In addition, we analyzed the behavior of the donor sequences HMLa and HMRa relative to MAT following a DSB at MAT. HMLa seems to sweep away from MAT during the minutes of repair in alpha-cells and come back to its original position following the repair. HMRa gets closer to MAT 30min after HO induction. In conclusion, we propose that the nuclear positioning of the HMLa along the cell cycle is non random and thus controlling the mating-type switch. In perspective, the adaptation of a new FROS system and the development of an algorithm able to analyze 3 loci simultaneously present a new tool to study the chromosome organization role in different cellular mechanism in yeast as well as in other models

Role of the chromosome three organization in the directionality of the mating type switch in S. cerevisiae

TOULOUSE3-BU Sciences (315552104) / SudocSudocFranceF

The Biochemistry and Evolution of the Dinoflagellate Nucleus

Dinoflagellates are known to possess a highly aberrant nucleus—the so-called dinokaryon—that exhibits a multitude of exceptional biological features. These include: (1) Permanently condensed chromosomes; (2) DNA in a cholesteric liquid crystalline state, (3) extremely large DNA content (up to 200 pg); and, perhaps most strikingly, (4) a deficit of histones—the canonical building blocks of all eukaryotic chromatin. Dinoflagellates belong to the Alveolata clade (dinoflagellates, apicomplexans, and ciliates) and, therefore, the biological oddities observed in dinoflagellate nuclei are derived character states. Understanding the sequence of changes that led to the dinokaryon has been difficult in the past with poor resolution of dinoflagellate phylogeny. Moreover, lack of knowledge of their molecular composition has constrained our understanding of the molecular properties of these derived nuclei. However, recent advances in the resolution of the phylogeny of dinoflagellates, particularly of the early branching taxa; the realization that divergent histone genes are present; and the discovery of dinoflagellate-specific nuclear proteins that were acquired early in dinoflagellate evolution have all thrown new light nature and evolution of the dinokaryon