Validation of DNA methylation profiling in formalin-fixed paraffin-embedded samples using the Infinium HumanMethylation450 Microarray

A formalin-fixed paraffin-embedded (FFPE) sample usually yields highly degraded DNA, which limits the use of techniques requiring high-quality DNA, such as Infinium Methylation microarrays. To overcome this restriction, we have applied an FFPE restoration procedure consisting of DNA repair and ligation processes in a set of paired fresh-frozen (FF) and FFPE samples. We validated the FFPE results in comparison with matched FF samples, enabling us to use FFPE samples on the Infinium HumanMethylation450 Methylation array.     

On the evolution of protamines in bony Fish: Alternatives to the “Retroviral horizontal transmission” hypothesis

Fish protamines are highly specialized molecules which are responsible for chromatin condensation during the last stages of spermatogenesis (spermiogenesis). However, not all fish contain protamines in their sperm nuclei; rather, there seems to be a random distribution of protamines within this group. The origin of this sporadic presence of protamines in the sperm and its significance have not yet been precisely determined. In this paper we have conducted an exhaustive survey of the literature available on the different types of nuclear protein composition of the sperm of teleost fish in order to try to correlate these data with what is presently known about the taxonomy of this group. The results of this analysis have allowed us to make the following observations. The divergence between protamines and histones has occurred several times during the evolution of the bony fish. However, the relative frequency of this divergence is almost negligible during the differentiation of genera and species (intrafamily variation) and is very small during the differentiation of families (interfamily variation). Nevertheless, the divergence is very noticeable among the different orders. It is therefore possible to conclude from all this that the sporadic distribution of protamines in bony fish is not a random event as initially believed. Furthermore, such a heterogeneous distribution of protamines cannot be easily accounted for by a mechanism of horizontal retroviral transmission through repeated and independent acquisition of a prot amine gene as has been recently proposed (Jankowski, Stater, Dixon (1986) J Mol Evol 23:1–10). Rather, it could possibly be explained by a repeated and independent loss of the expression of the protamine gene (or loss of the gene itself) which mainly occurred during the diversification of the orders of this group.Correspondence to: J. Ausio     

Protein phosphatases in higher plants: multiplicity of type 2A phosphatases in Arabidopsis thaliana

Two DNA fragments, AP-1 and AP-2, encoding amino acid sequences closely related to Ser/Thr protein phosphatases were amplified from Arabidopsis thaliana genomic DNA. Fragment AP-1 was used to screen. A. thaliana cDNA libraries and several positive clones were isolated. Clones EP8a and EP14a were sequenced and found to encode almost identical proteins (97% identity). Both proteins are 306 amino acids in length and are very similar (79–80% identity) to the mammalian isotypes of the catalytic subunit of protein phosphatase 2A. Therefore, they have been designated PP2A-1 and PP2A-2. A third cDNA clone, EP7, was isolated and sequenced. The polypeptide encoded (308 amino acids, lacking the initial Met codon) is 80% identical with human phosphatases 2A and was named PP2A-3. The PP2A-3 protein is extremely similar (95% identity) to the predicted protein from a cDNA clone previously found in Brassica napus. Southern blot analysis of genomic DNA using AP-1 and AP-2 probes, as well as probes derived from clones EP7, EP8a and EP14a strongly indicates that at least 6 genes closely related to type 2A phosphatases are present in the genome of A. thaliana. Northern blot analysis using the same set of probes demonstrates that, at the seedling stage, the mRNA levels for PP2A-1, PP2A-3 and the gene containing the AP-1 sequence are much higher than those of PP2A-2 and AP-2. These results demonstrate that a multiplicity of type 2A phosphatases might be differentially expressed in higher plants.     

The distribution of coastal fish eDNA sequences in the Anthropocene

Aim

Coastal fishes have a fundamental role in marine ecosystem functioning and contributions to people, but face increasing threats due to climate change, habitat degradation and overexploitation. The extent to which human pressures are impacting coastal fish biodiversity in comparison with geographic and environmental factors at large spatial scale is still under scrutiny. Here, we took advantage of environmental DNA (eDNA) metabarcoding to investigate the relationship between fish biodiversity, including taxonomic and genetic components, and environmental but also socio-economic factors.

Location

Tropical, temperate and polar coastal areas.

Time period

Present day.

Major taxa studied

Marine fishes.

Methods

We analysed fish eDNA in 263 stations (samples) in 68 sites distributed across polar, temperate and tropical regions. We modelled the effect of environmental, geographic and socio-economic factors on α- and β-diversity. We then computed the partial effect of each factor on several fish biodiversity components using taxonomic molecular units (MOTU) and genetic sequences. We also investigated the relationship between fish genetic α- and β-diversity measured from our barcodes, and phylogenetic but also functional diversity.

Results

We show that fish eDNA MOTU and sequence α- and β-diversity have the strongest correlation with environmental factors on coastal ecosystems worldwide. However, our models also reveal a negative correlation between biodiversity and human dependence on marine ecosystems. In areas with high dependence, diversity of all fish, cryptobenthic fish and large fish MOTUs declined steeply. Finally, we show that a sequence diversity index, accounting for genetic distance between pairs of MOTUs, within and between communities, is a reliable proxy of phylogenetic and functional diversity.

Main conclusions

Together, our results demonstrate that short eDNA sequences can be used to assess climate and direct human impacts on marine biodiversity at large scale in the Anthropocene and can further be extended to investigate biodiversity in its phylogenetic and functional dimensions.     

Total Tocopherols,Carotenoids, and Fatty Acids Content Variation of Pistacia atlantica from Different Organs’ Crude Oils and Their Antioxidant Activity during Development Stages

The current study investigated the effect of developmental stages on the chemical composition and the antioxidant activity of fifteen crude oil samples obtained from Pistacia atlantica Desf. leaves, galls, and fruits. Twelve fatty acids were detected by GC/FID, linolenic acid (C18 : 3) was the major fatty acid detected in leaves crude oils that registered [41.73 % (P<0.05)] on the last stage. The best content of tocopherols and carotenoids was recorded at the last stage for leaves and galls oils, respectively, with values of [1.530±0.01, 0.52±0.01 (P<0.05) mg α‐tocopherol equivalent/g DW] and [86.60±0.95, 69.15±0.13 (P<0.05) μg β‐carotene equivalent/g DW]. For fruits oils, the content varied depending on the levels of fruits maturation. The results from DPPH, FRAP, and ABTS assays revealed that the antioxidant activity increased with the increasing content of tocopherols and carotenoids in leaves and galls oils during development stages, and varied for fruits oils depending on the ripening stages. Moreover, according to PCA analysis, the best phytoconstituent content and antioxidant activity were attributed to P. atlantica Desf. fruit's crude oils. Also, a strong relationship was found between the antioxidant activity and bioactive phytochemical components, such as tocopherols, carotenoids, and omega‐three fatty acid, which confirmed that P. atlantica Desf. crude oils present a valuable source of natural antioxidant that could be used for pharmaceutical and food industries purposes.     

Spatial graphs highlight how multi-generational dispersal shapes landscape genetic patterns

Current approaches that compare spatial genetic structure of a given species and the dispersal of its mobile phase can detect a mismatch between both patterns mainly due to processes acting at different temporal scales. Genetic structure result from gene flow and other evolutionary and demographic processes over many generations, while dispersal predicted from the mobile phase often represents solely one generation on a single time-step. In this study, we present a spatial graph approach to landscape genetics that extends connectivity networks with a stepping-stone model to represent dispersal between suitable habitat patches over multiple generations. We illustrate the approach with the case of the striped red mullet Mullus surmuletus in the Mediterranean Sea. The genetic connectivity of M. surmuletus was not correlate with the estimated dispersal probability over one generation, but with the stepping-stone estimate of larval dispersal, revealing the temporal scale of connectivity across the Mediterranean Sea. Our results highlight the importance of considering multiple generations and different time scales when relating demographic and genetic connectivity. The spatial graph of genetic distances further untangles intra-population genetic structure revealing the Siculo-Tunisian Strait as an important corridor rather than a barrier for gene flow between the Western- and Eastern Mediterranean basins, and identifying Mediterranean islands as important stepping-stones for gene flow between continental populations. Our approach can be easily extended to other systems and environments.     

Genetic syndromes caused by mutations in epigenetic genes

The orchestrated organization of epigenetic factors that control chromatin dynamism, including DNA methylation, histone marks, non-coding RNAs (ncRNAs) and chromatin-remodeling proteins, is essential for the proper function of tissue homeostasis, cell identity and development. Indeed, deregulation of epigenetic profiles has been described in several human pathologies, including complex diseases (such as cancer, cardiovascular and neurological diseases), metabolic pathologies (type 2 diabetes and obesity) and imprinting disorders. Over the last decade it has become increasingly clear that mutations of genes involved in epigenetic mechanism, such as DNA methyltransferases, methyl-binding domain proteins, histone deacetylases, histone methylases and members of the SWI/SNF family of chromatin remodelers are linked to human disorders, including Immunodeficiency Centromeric instability Facial syndrome 1, Rett syndrome, Rubinstein–Taybi syndrome, Sotos syndrome or alpha-thalassemia/mental retardation X-linked syndrome, among others. As new members of the epigenetic machinery are described, the number of human syndromes associated with epigenetic alterations increases. As recent examples, mutations of histone demethylases and members of the non-coding RNA machinery have recently been associated with Kabuki syndrome, Claes-Jensen X-linked mental retardation syndrome and Goiter syndrome. In this review, we describe the variety of germline mutations of epigenetic modifiers that are known to be associated with human disorders, and discuss the therapeutic potential of epigenetic drugs as palliative care strategies in the treatment of such disorders.