Effective Potentials and Orbits in Weyl Conformastatic Slender Disk

Centromeres, the chromosomal loci where spindle fibers attach during cell division to segregate chromosomes, are typically found within satellite arrays in plants and animals. Satellite arrays have been difficult to analyze because they comprise megabases of tandem head-to-tail highly repeated DNA sequences. Much evidence suggests that centromeres are epigenetically defined by the location of nucleosomes containing the centromere-specific histone H3 variant cenH3, independently of the DNA sequences where they are located; however, the reason that cenH3 nucleosomes are generally found on rapidly evolving satellite arrays has remained unclear. Recently, long read sequencing technology has clarified the structures of satellite arrays and sparked rethinking of how they evolve, while new experiments and analyses have helped bring both understanding and further speculation about the role these highly repeated sequences play in centromere identification.

Creation of a synthesis-friendly inflammation-inducible promoter suitable for cell therapy

The development of ‘smart’ cell-based therapeutics requires cells that first recognize conditions consistent with disease (e.g. inflammation) and then subsequently release therapeutic proteins, thereby reducing potential toxicity from otherwise continuous expression. Promoters containing NF-κB response elements are often used as reporters of inflammation; however, endogenous promoters have crosstalk with other pathways, and current synthetic promoters have many exact sequence repeats of NF-κB response elements which make them both difficult to synthesize and inherently genetically unstable. Herein, a synthesis-friendly inflammation-inducible promoter (named SFNp) was created by the packing of 14 NF-κB response elements, which have no repeats >9 bp, followed by a minimal cytomegalovirus promoter. In stably expressing human embryonic kidney 293 cells, we assessed the ability of SFNp to inducibly transcribe genes for reporting expression, changing cell morphology, and performing cell fusion. These experiments represent simple milestones for potentially using SFNp in the development of cell-based therapeutics. As strongly repeated DNA can compromise the long-term stability of genetic circuits, new designs used in ‘smart’ cell therapy will become more reliant on synthesis-friendly components like SFNp.

Multi - level dynamics of heterochromatin and repair sites undergoing homologous recombination

Studies across different organisms show that nuclear architecture and dynamics play central roles in different aspects of homologous recombination (HR) repair. Here we review the most recent discoveries in this field, ranging from directed motions mediating relocalization pathways, to global chromatin mobilization, local DNA looping, and changes in repair focus properties associated with clustering and phase separation. We will highlight how these dynamics work in different contexts, including the molecular mechanisms and regulatory pathways involved. We will also discuss how they function in pericentromeric heterochromatin, which presents a unique environment for HR repair given the abundance of repeated DNA sequences prone to aberrant recombination, the 'silent' chromatin state, and the phase separation characterizing this domain.

Satellitome Analysis of Rhodnius prolixus, One of the Main Chagas Disease Vector Species

The triatomine Rhodnius prolixus is the main vector of Chagas disease in countries such as Colombia and Venezuela, and the first kissing bug whose genome has been sequenced and assembled. In the repetitive genome fraction (repeatome) of this species, the transposable elements represented 19% of R. prolixus genome, being mostly DNA transposon (Class II elements). However, scarce information has been published regarding another important repeated DNA fraction, the satellite DNA (satDNA), or satellitome. Here, we offer, for the first time, extended data about satellite DNA families in the R. prolixus genome using bioinformatics pipeline based on low-coverage sequencing data. The satellitome of R. prolixus represents 8% of the total genome and it is composed by 39 satDNA families, including four satDNA families that are shared with Triatoma infestans, as well as telomeric (TTAGG)n and (GATA)n repeats, also present in the T. infestans genome. Only three of them exceed 1% of the genome. Chromosomal hybridization with these satDNA probes showed dispersed signals over the euchromatin of all chromosomes, both in autosomes and sex chromosomes. Moreover, clustering analysis revealed that most abundant satDNA families configured several superclusters, indicating that R. prolixus satellitome is complex and that the four most abundant satDNA families are composed by different subfamilies. Additionally, transcription of satDNA families was analyzed in different tissues, showing that 33 out of 39 satDNA families are transcribed in four different patterns of expression across samples.

Characterisation of the S1 satellite DNA family in Rana dalmatina

In this study, the DNA of Rana dalmatina was digested with Asp 718I and the two bands of highly repeated DNA produced were cloned and characterised. The largest fragment (494 bp) corresponded to the entire repetitive unit of the major satellite DNA (RdS1a), while the smaller fragment of 385 bp corresponded to the major fragment of RdS1a produced by digestion. A fragment of 332 bpbcorresponding to the repetitive unit of satellite S1b (RdS1b) was instead achieved by digestion with Eco RV. RdS1b is highly homologous to the corresponding portion of the repetition of RdS1a and presents the first 36 bp repeated and inverted. This suggested that RdS1b would have been derived from satellite S1a by two distinct and subsequent events. Further, the high sequence homology and length between RdS1a and the S1a of Rana italica (RiS1a) confirmed the hypothesis that the satellite S1a is antecedent to S1b and inherited from a common ancestor. Southern blots of R. dalmatina genomic DNA digested with Asp 718I produced hybrid bands of fragments of different sizes containing in addition to the satellite S1a, also one or more copies of the S1b satellites. The only sequenced band at the moment corresponded to the repetitive unit of the satellite RdS1a + b (826 bp) deleted of the fragment Asp 718I less than RdS1a (109 bp), while the other double bands should almost certainly correspond to repetitive units of satellites RdS1a + 2b and RdS1a + 3b. Our data suggested different satellite DNA organisation in R. dalmatina, including the tandem structure of the repetitive units of the RdS1a or RdS1b. Our data also suggested the existence in R. dalmatina of at least four different types of hybrid repeating units in all the populations examined.

SetDB1 and Su(var)3-9 play non-overlapping roles in somatic cell chromosomes of Drosophila melanogaster

We explored functional roles of two H3K9-specific histone methyltransferases SetDB1 and Su(var)3-9. Using DamID approach, we generated the binding profile for SetDB1 in Drosophila salivary gland chromosomes, and matched it to the profile of Su(var)3-9. Unlike Su(var)3-9, SetDB1 turned out to be an euchromatic protein that is absent from repeated DNA compartment, and is largely restricted to TSSes and 5'UTRs of ubiquitously expressed genes. Significant SetDB1 association is also observed at insulator protein CP190 binding sites. SetDB1 and H3K9me2/3-enriched sites tend to display poor overlap. At the same time, SetDB1 has clear connection with the distribution of H3K27me3 mark. SetDB1 binds outside the domains possessing this modification, and about half of the borders of H3K27me3 domains are decorated by SetDB1 together with actively transcribed genes. On the basis of poor correlation between the distribution of SetDB1 and H3K9 methylation marks, we speculate that in somatic cells, SetDB1 may contribute to the methylation of a broader set of chromosomal proteins than just H3K9. In addition, SetDB1 can be expected to play a role in the establishment of chromatin functional domains.

The H2A.J histone variant contributes to Interferon-Stimulated Gene expression in senescence by its weak interaction with H1 and the derepression of repeated DNA sequences

The histone variant H2A.J was previously shown to accumulate in senescent human fibroblasts with persistent DNA damage to promote inflammatory gene expression, but its mechanism of action was unknown. We show that H2A.J accumulation contributes to weakening the association of histone H1 to chromatin and increasing its turnover. Decreased H1 in senescence is correlated with increased expression of some repeated DNA sequences, increased expression of STAT/IRF transcription factors, and transcriptional activation of Interferon-Stimulated Genes (ISGs). The H2A.J-specific Val-11 moderates the transcriptional activity of H2A.J, and H2A.J-specific Ser-123 can be phosphorylated in response to DNA damage with potentiation of its transcriptional activity by the phospho-mimetic S123E mutation. Our work demonstrates the functional importance of H2A.J-specific residues and potential mechanisms for its function in promoting inflammatory gene expression in senescence.

Molecular Cytogenetic Analysis of Karyotype and Y Chromosome Conservation in Species of the Genus Talpa (Insectivora)

The Talpidae family has a highly stable karyotype. Most of the chromosome studies in this mammal group, however, employed classical cytogenetic techniques. Molecular cytogenetic analyses are still scarce and, for example, no repeated DNA sequences have been described to date. In this work, we used sequence analysis, chromosomal mapping of a LINE1 retroelement sequence, as well as chromosome painting with a whole Y chromosome probe of T. occidentalis to compare the karyotypes of 3 species of the genus Talpa (T. occidentalis, T. romana, and T. aquitania). Our results demonstrate that in Talpa genomes LINE1 sequences are widely distributed on all chromosomes but are enriched in pericentromeric C-band-positive regions. In addition, these LINE1 accumulate on the Y chromosomes of the 3 Talpa species regardless of their euchromatic or heterochromatic condition. Chromosome painting shows that the Y chromosomes in these 3 species are highly conserved. Interestingly, they share sequences with heterochromatic blocks on chromosome pairs 14 and 16 and, to a lesser degree, with the pericentromeric regions of other autosomes. Together, our analyses demonstrate that the repetitive DNA content of chromosomes from Talpa species is highly conserved.