Lysine 27 dimethylation of Drosophila linker histone dH1 contributes to heterochromatin organization independently of H3K9 methylation

Post-translational modifications (PTMs) of core histones are important epigenetic determinants that correlate with functional chromatin states. However, despite multiple linker histone H1s PTMs have been identified, little is known about their genomic distribution and contribution to the epigenetic regulation of chromatin. Here, we address this question in Drosophila that encodes a single somatic linker histone, dH1. We previously reported that dH1 is dimethylated at K27 (dH1K27me2). Here, we show that dH1K27me2 is a major PTM of Drosophila heterochromatin. At mitosis, dH1K27me2 accumulates at pericentromeric heterochromatin, while, in interphase, it is also detected at intercalary heterochromatin. ChIPseq experiments show that >98% of dH1K27me2 enriched regions map to heterochromatic repetitive DNA elements, including transposable elements, simple DNA repeats and satellite DNAs. Moreover, expression of a mutated dH1K27A form, which impairs dH1K27me2, alters heterochromatin organization, upregulates expression of heterochromatic transposable elements and results in the accumulation of RNA:DNA hybrids (R-loops) in heterochromatin, without affecting H3K9 methylation and HP1a binding. The pattern of dH1K27me2 is H3K9 methylation independent, as it is equally detected in flies carrying a H3K9R mutation, and is not affected by depletion of Su(var)3–9, HP1a or Su(var)4–20. Altogether these results suggest that dH1K27me2 contributes to heterochromatin organization independently of H3K9 methylation.     

Accurate measurement of microsatellite length by disrupting its tandem repeat structure

Tandem repeats of simple sequence motifs, also known as microsatellites, are abundant in the genome. Because their repeat structure makes replication error-prone, variant microsatellite lengths are often generated during germline and other somatic expansions. As such, microsatellite length variations can serve as markers for cancer. However, accurate error-free measurement of microsatellite lengths is difficult with current methods precisely because of this high error rate during amplification. We have solved this problem by using partial mutagenesis to disrupt enough of the repeat structure of initial templates so that their sequence lengths replicate faithfully. In this work, we use bisulfite mutagenesis to convert a C to a U, later read as T. Compared to untreated templates, we achieve three orders of magnitude reduction in the error rate per round of replication. By requiring agreement from two independent first copies of an initial template, we reach error rates below one in a million. We apply this method to a thousand microsatellite loci from the human genome, revealing microsatellite length distributions not observable without mutagenesis.     

The clinical characterization of the adult patient with bipolar disorder aimed at personalization of management

Bipolar disorder is heterogeneous in phenomenology, illness trajectory, and response to treatment. Despite evidence for the efficacy of multimodal­ity interventions, the majority of persons affected by this disorder do not achieve and sustain full syndromal recovery. It is eagerly anticipated that combining datasets across various information sources (e.g., hierarchical “multi‐omic” measures, electronic health records), analyzed using advanced computational methods (e.g., machine learning), will inform future diagnosis and treatment selection. In the interim, identifying clinically meaningful subgroups of persons with the disorder having differential response to specific treatments at point‐of‐care is an empirical priority. This paper endeavours to synthesize salient domains in the clinical characterization of the adult patient with bipolar disorder, with the overarching aim to improve health outcomes by informing patient management and treatment considerations. Extant data indicate that characterizing select domains in bipolar disorder provides actionable information and guides shared decision making. For example, it is robustly established that the presence of mixed features – especially during depressive episodes – and of physical and psychiatric comorbidities informs illness trajectory, response to treatment, and suicide risk. In addition, early environmental exposures (e.g., sexual and physical abuse, emotional neglect) are highly associated with more complicated illness presentations, inviting the need for developmentally‐oriented and integrated treatment approaches. There have been significant advances in validating subtypes of bipolar disorder (e.g., bipolar I vs. II disorder), particularly in regard to pharmacological interventions. As with other severe mental disorders, social functioning, interpersonal/family relationships and internalized stigma are domains highly relevant to relapse risk, health outcomes, and quality of life. The elevated standardized mortality ratio for completed suicide and suicidal behaviour in bipolar disorder invites the need for characterization of this domain in all patients. The framework of this paper is to describe all the above salient domains, providing a synthesis of extant literature and recommendations for decision support tools and clinical metrics that can be implemented at point‐of‐care.     

Nuclear localization of TRK-A in liver cells

The liver represents a site of expression of neurotrophins and their receptors. We have characterized the expression and intracellular localization of the nerve growth factor (NGF) receptor, Trk-A, in liver cells in vivo and in vitro. In both normal and fibrotic liver tissue, Trk-A immunostaining was present in different cell types, including parenchymal cells and cells of the inflammatory infiltrate. In hepatocytes and activated stellate cells (HSC), Trk-A showed a predominant nuclear localization, both in the presence and absence of injury. In cultured HSC, Trk-A was found to be functional, because exposure of the cells to recombinant NGF resulted in stimulation of cell migration and activation of intracellular signaling pathways, including Ras-ERK and PI3K/Akt. Remarkably, in cultured HSC, Trk-A staining was found constitutively in the nucleus. In these cells, Trk-A could be stained only by antibodies directed against the intracellular domain but not by those recognizing the extracellular portion of Trk-A suggesting that the intracellular portion of the receptor is the major determinant of nuclear Trk-A staining. In contrast to HSC, freshly isolated hepatocytes did not show any nuclear localization of the intracellular portion of Trk-A. In pheocromocytoma cells, nuclear staining for Trk-A was not present in conditions of serum deprivation, but could be induced by exposure to NGF or to a mixture of soluble mediators. We conclude that nuclear localization of the intracellular domain of Trk-A is observed constitutively in liver cells such as HSC, while in other cell types it could be induced in response to soluble factors.     

Biogeography of dinoflagellate cysts in northwest Atlantic estuaries

Few biogeographic studies of dinoflagellate cysts include the near‐shore estuarine environment. We determine the effect of estuary type, biogeography, and water quality on the spatial distribution of organic‐walled dinoflagellate cysts from the Northeast USA (Maine to Delaware) and Canada (Prince Edward Island). A total of 69 surface sediment samples were collected from 27 estuaries, from sites with surface salinities >20. Dinoflagellate cysts were examined microscopically and compared to environmental parameters using multivariate ordination techniques. The spatial distribution of cyst taxa reflects biogeographic provinces established by other marine organisms, with Cape Cod separating the northern Acadian Province from the southern Virginian Province. Species such as Lingulodinium machaerophorum and Polysphaeridinium zoharyi were found almost exclusively in the Virginian Province, while others such as Dubridinium spp. and Islandinium? cezare were more abundant in the Acadian Province. Tidal range, sea surface temperature (SST), and sea surface salinity (SSS) are statistically significant parameters influencing cyst assemblages. Samples from the same type of estuary cluster together in canonical correspondence analysis when the estuaries are within the same biogeographic province. The large geographic extent of this study, encompassing four main estuary types (riverine, lagoon, coastal embayment, and fjord), allowed us to determine that the type of estuary has an important influence on cyst assemblages. Due to greater seasonal variations in SSTs and SSSs in estuaries compared to the open ocean, cyst assemblages show distinct latitudinal trends. The estuarine context is important for understanding present‐day species distribution, the factors controlling them, and to better predict how they may change in the future.     

Conspecific plant–soil feedback scales with population size in Lobelia siphilitica (Lobeliaceae)

Plant–soil interactions directly affect plant success in terms of establishment, survival, growth and reproduction. Negative plant–soil feedback on such traits may therefore reduce the density and abundance of plants of a given species at a given site. Furthermore, if conspecific feedback varies among population sites, it could help explain geographic variation in plant population size. We tested for among-site variation in conspecific plant–soil feedback in a greenhouse experiment using seeds and soils from 8 natural populations of Lobelia siphilitica hosting 30–330 plants. The first cohort of seeds was grown on soil collected from each native site, while the second cohort was grown on the soil conditioned by the first. Our goal was to distinguish site-specific effects mediated by biotic and/or abiotic soil properties from those inherent in seed sources. Cohort 1 plants grown from seeds produced in small populations performed better in terms of germination, growth, and survival compared to plants produced in large populations. Plant performance decreased substantially between cohorts, indicating strong negative feedback. Most importantly, the strength of negative feedback scaled linearly (i.e., was less negative) with increasing size of the native plant population, particularly for germination and survival, and was better explained by soil- rather than seed-source effects. Even with a small number of sites, our results suggest that the potential for negative plant–soil feedback varies among populations of L. siphilitica, and that small populations were more susceptible to negative feedback. Conspecific plant–soil feedback may contribute to plant population size variation within a species’ native range.     

A quantitative trait locus involved in maize yield is tightly associated to the r1 gene on the long arm of chromosome 10

We produced and studied for 3?years two synthetic populations of maize differing in their constitution only for the selected alleles present at the red color 1 (r1) locus (R-sc vs. r?Cr). r1 is a regulatory gene conferring anthocyanin pigmentation in different tissues: the R-sc allele confers pigmentation only in the aleurone seed layer, while the r?Cr allele confers pigmentation in several tissues such as root, silk and anther but the seed is colourless. The colourless population (r?Cr/r?Cr) was characterized by improved agronomic features, such as ear weight and plant height, compared with the R-sc/R-sc coloured population. This finding was confirmed by studying single F4 R/r families where the presence of the r?Cr allele conferred positive features, acting as a dominant trait. Quantitative trait locus (QTL) analysis performed using molecular markers on the long arm of chromosome 10 (bin 10.06), where the r1 gene maps, identified a QTL map position for plant height tightly associated to the r1 gene. Thus the r1 gene may represent a major QTL or it could be closely linked to another gene involved in the agronomic performance of the two populations studied.     

Lysosomal fusion and SNARE function are impaired by cholesterol accumulation in lysosomal storage disorders

The function of lysosomes relies on the ability of the lysosomal membrane to fuse with several target membranes in the cell. It is known that in lysosomal storage disorders (LSDs), lysosomal accumulation of several types of substrates is associated with lysosomal dysfunction and impairment of endocytic membrane traffic. By analysing cells from two severe neurodegenerative LSDs, we observed that cholesterol abnormally accumulates in the endolysosomal membrane of LSD cells, thereby reducing the ability of lysosomes to efficiently fuse with endocytic and autophagic vesicles. Furthermore, we discovered that soluble N‐ethylmaleimide‐sensitive factor attachment protein (SNAP) receptors (SNAREs), which are key components of the cellular membrane fusion machinery are aberrantly sequestered in cholesterol‐enriched regions of LSD endolysosomal membranes. This abnormal spatial organization locks SNAREs in complexes and impairs their sorting and recycling. Importantly, reducing membrane cholesterol levels in LSD cells restores normal SNARE function and efficient lysosomal fusion. Our results support a model by which cholesterol abnormalities determine lysosomal dysfunction and endocytic traffic jam in LSDs by impairing the membrane fusion machinery, thus suggesting new therapeutic targets for the treatment of these disorders.     

The conformational quality of insoluble recombinant proteins is enhanced at low growth temperatures

Protein aggregation is a major bottleneck during the bacterial production of recombinant proteins. In general, the induction of gene expression at sub-optimal growth temperatures improves the solubility of aggregation-prone polypeptides and minimizes inclusion body (IB) formation. However, the effect of low temperatures on the quality of the recombinant protein, especially within the insoluble cell fraction, has been hardly ever explored. In this work, we have examined the conformational status of a recombinant GFP protein when produced in Escherichia coli below 37 degrees C. As expected, the fraction of aggregated protein largely decreased at lower temperatures, while the conformational quality of both soluble and aggregated GFP, as reflected by its specific fluorescence emission, progressively improved. This observation indicates that physicochemical conditions governing protein folding affect concurrently the quality of the soluble and the aggregated forms of a misfolding-prone protein, and that protein misfolding and aggregation are clearly not coincident events.