Neurohumoral basis of circadian rhythmicity in Nephrops norvegicus (L)

A circadian rhythm of activity is demonstrated in single neurons of Nephrops norvegicus (L.). Eyestalk extracts depress neural and locomotor activity. Entrainment of rhythmicity is achieved by the environmental light cycle, apparently acting through the eye.     

Productivity: key factor affecting grazing exclusion effects on vegetation and soil

In this study, we inquire into the effects of short-term goat grazing abandonment on plant species and functional composition, bare ground and net primary productivity (NPP) in two traditionally grazed pastures located in the Canarian Network of Natural Protected Areas and the Natura 2000 Network. In addition, we analyse soil chemical properties, biomass tannin content and energetic value to find out how grazing abandonment affects soil fertility and forage quality of these agroecosystems. Grazing exclusion effects on plant species and functional composition, as well as on soil fertility depended on the productivity of the studied pasture. Erect forbs and shrubs (endemic to Macaronesian region and native) were favoured by grazing removal in the most productive pasture, while soil fertility decreased in the driest and least productive site. An increase in NPP after exclusion was consistent among study sites. Although we consider goat grazing as necessary for maintaining traditional agroecosystems, we also suggest controlling it over time, allowing some periods of rest to give endemic shrub species time to recover from near propagule sources.     

Adenosine A2A receptor (A2AR) is a fine-tune regulator of the collagen1:collagen3 balance

Adenosine is a potent endogenous anti-inflammatory and immunosuppressive metabolite that is a potent modulator of tissue repair. However, the adenosine A_2A receptor (A_2AR)-mediated promotion of collagen synthesis is detrimental in settings such as scarring and scleroderma. The signaling cascade from A_2AR stimulation to increased collagen production is complex and obscure, not least because cAMP and its downstream molecules PKA and Epac1 have been reported to inhibit collagen production. We therefore examined A_2AR-stimulated signaling for collagen production by normal human dermal fibroblasts (NHDF). Collagen1 (Col1) and collagen3 (Col3) content after A_2AR activation by CGS21680 was studied by western blotting. Contribution of PKA and Epac was analyzed by the PKA inhibitor PKI and by knockdowns of the PKA-Cα, -Cβ, -Cγ, Epac1, and Epac2. CGS21680 stimulates Col1 expression at significantly lower concentrations than those required to stimulate Col3 expression. A_2AR stimulates Col1 expression by a PKA-dependent mechanism since PKA inhibition or PKA-Cα and -Cβ knockdown prevents A_2AR-mediated Col1 increase. In contrast, A_2AR represses Col3 via PKA but stimulates both Col1 and Col3 via an Epac2-dependent mechanism. A_2AR stimulation with CGS21680 at 0.1 μM increased Col3 expression only upon PKA blockade. A_2AR activation downstream signaling for Col1 and Col3 expression proceeds via two distinct pathways with varying sensitivity to cAMP activation; more highly cAMP-sensitive PKA activation stimulates Col1 expression, and less cAMP-sensitive Epac activation promotes both Col1 and Col3 expression. These observations may explain the dramatic change in Col1:Col3 ratio in hypertrophic and immature scars, where adenosine is present in higher concentrations than in normal skin.     

BMP9 Is a Proliferative and Survival Factor for Human Hepatocellular Carcinoma Cells

TGF-β family members play a relevant role in tumorigenic processes, including hepatocellular carcinoma (HCC), but a specific implication of the Bone Morphogenetic Protein (BMP) subfamily is still unknown. Although originally isolated from fetal liver, little is known about BMP9, a BMP family member, and its role in liver physiology and pathology. Our results show that BMP9 promotes growth in HCC cells, but not in immortalized human hepatocytes. In the liver cancer cell line HepG2, BMP9 triggers Smad1,5,8 phosphorylation and inhibitor of DNA binding 1 (Id1) expression up- regulation. Importantly, by using chemical inhibitors, ligand trap and gene silencing approaches we demonstrate that HepG2 cells autocrinely produce BMP9 that supports their proliferation and anchorage independent growth. Additionally, our data reveal that in HepG2 cells BMP9 triggers cell cycle progression, and strikingly, completely abolishes the increase in the percentage of apoptotic cells induced by long-term incubation in low serum. Collectively, our data unveil a dual role for BMP9, both promoting a proliferative response and exerting a remarkable anti-apoptotic function in HepG2 cells, which result in a robust BMP9 effect on liver cancer cell growth. Finally, we show that BMP9 expression is increased in 40% of human HCC tissues compared with normal human liver as revealed by immunohistochemistry analysis, suggesting that BMP9 signaling may be relevant during hepatocarcinogenesis in vivo. Our findings provide new clues for a better understanding of BMPs contribution, and in particular BMP9, in HCC pathogenesis that may result in the development of effective and targeted therapeutic interventions.     

Palbociclib,a selective CDK4/6 inhibitor,restricts cell survival and epithelial-mesenchymal transition in Panc-1 and MiaPaCa-2 pancreatic cancer cells

The mortality rate of pancreatic cancer has close parallels to its incidence rate because of limited therapeutics and lack of effective prognosis. Despite various novel chemotherapeutics combinations, the 5-year survival rate is still under 5%. In the current study, we aimed to modulate the aberrantly activated PI3K/AKT pathway and epithelial-mesenchymal transition (EMT) signaling with the treatment of CDK4/6 inhibitor PD-0332991 (palbociclib) in Panc-1 and MiaPaCa-2 pancreatic cancer cells. It was found that PD-0332991 effectively reduced cell viability and proliferation dose-dependently within 24 hours. In addition, PD-0332991 induced cell cycle arrest at the G1 phase by downregulation of aberrant expression of CDK4/6 through the dephosphorylation of Rb in each cell lines. Although PD-0332991 treatment increased epithelial markers and decreased mesenchymal markers, the nuclear translocation of β-catenin was not prevented by PD-0332991 treatment, especially in MiaPaCa-2 cells. Effects of PD-0332991 on the regulation of PI3K/AKT signaling and its downstream targets such as GSK-3 were cell type-dependent. Although the activity of AKT was inhibited in both cell lines, the phosphorylation of GSK-3β at Ser9 increased only in Panc-1. In conclusion, PD-0332991 induced cell cycle arrest and reduced the cell viability of Panc-1 and MiaPaCa-2 cells. However, PD-0332991 differentially affects the regulation of the PI3K/AKT pathway and EMT process in cells due to its distinct influence on Rb and GSK-3/β-catenin signaling. Understanding the effect of PD-0332991 on the aberrantly activated signaling axis may put forward a new therapeutic strategy to reduce the cell viability and metastatic process of pancreatic cancer.     

An Extracellular Ion Pathway Plays a Central Role in the Cooperative Gating of a K2P K+ Channel by Extracellular pH

Proton-gated TASK-3 K⁺ channel belongs to the K_2P family of proteins that underlie the K⁺ leak setting the membrane potential in all cells. TASK-3 is under cooperative gating control by extracellular [H⁺]. Use of recently solved K_2P structures allows us to explore the molecular mechanism of TASK-3 cooperative pH gating. Tunnel-like side portals define an extracellular ion pathway to the selectivity filter. We use a combination of molecular modeling and functional assays to show that pH-sensing histidine residues and K⁺ ions mutually interact electrostatically in the confines of the extracellular ion pathway. K⁺ ions modulate the pK_a of sensing histidine side chains whose charge states in turn determine the open/closed transition of the channel pore. Cooperativity, and therefore steep dependence of TASK-3 K⁺ channel activity on extracellular pH, is dependent on an effect of the permeant ion on the channel pH_o sensors.     

Nucleotide diversity of vernalization and flowering‐time‐related genes in a germplasm collection of meadow fescue (Festuca pratensis Huds. syn. Lolium pratense (Huds.) Darbysh.)

In plant species, control of flowering time is an important factor for adaptation to local natural environments. The Vrn1, CO, FT1 and CK2α genes are key components in the flowering‐specific signaling pathway of grass species. Meadow fescue is an agronomically important forage grass species, which is naturally distributed across Europe and Western Asia. In this study, meadow fescue flowering‐time‐related genes were resequenced to assess nucleotide diversity in European and Western Asian subpopulations. Identified sequence polymorphisms were then converted into PCR‐based molecular genetic markers, and a meadow fescue germplasm collection was genotyped to investigate global allelic variation. Lower nucleotide diversities were observed for the Vrn1 and CO orthologs, while relatively higher values were observed for the FT1 and casein kinase II α‐subunit (CK2α) orthologs. The nucleotide diversity for FT1 orthologs in the Western Asian subpopulation was significantly higher than those of the European subpopulation. Similarly, significant differences in nucleotide diversity for the remaining genes were observed between several combinations of subpopulation. The global allele distribution pattern was consistent with observed level of nucleotide diversity. These results suggested that the degree of purifying selection acting on the genes differs according to geographical location. As previously shown for model plant species, functional specificities of flowering‐time‐related genes may also vary according to environmental conditions.     

Identifying SNP markers tightly associated with six major genes in peach [<Emphasis Type="Italic">Prunus persica</Emphasis> (L.) Batsch] using a high-density SNP array with an objective of marker-assisted selection (MAS)

One of the applications of genomics is to identify genetic markers linked to loci responsible for variation in phenotypic traits, which could be used in breeding programs to select individuals with favorable alleles, particularly at the seedling stage. With this aim, in the framework of the European project FruitBreedomics, we selected five main peach fruit characters and a resistance trait, controlled by major genes with Mendelian inheritance: fruit flesh color Y, fruit skin pubescence G, fruit shape S, sub-acid fruit D, stone adhesion-flesh texture F-M, and resistance to green peach aphid Rm2. They were all previously mapped in Prunus. We then selected three F₁ and three F₂ progenies segregating for these characters and developed genetic maps of the linkage groups including the major genes, using the single nucleotide polymorphism (SNP) genome-wide scans obtained with the International Peach SNP Consortium (IPSC) 9K SNP array v1. We identified SNPs co-segregating with the characters in all cases. Their positions were in agreement with the known positions of the major genes. The number of SNPs linked to each of these, as well as the size of the physical regions encompassing them, varied depending on the maps. As a result, the number of useful SNPs for marker-assisted selection varied accordingly. As a whole, this study establishes a sound basis for further development of MAS on these characters. Additionally, we also discussed some limitations that were observed regarding the SNP array efficiency.     

Marker-assisted introgression (MAI) of almond genes into the peach background: a fast method to mine and integrate novel variation from exotic sources in long intergeneration species

This paper proposes a new breeding strategy, marker-assisted introgression (MAI), to obtain lines of perennial species with a single introgressed fragment from a compatible species two generations after the interspecific hybrid. MAI allows enrichment of the genome of a species with genes from a wild or exotic relative in a short timeframe and with an intermediate step that allows a first exploration of genes/QTLs that the donor species can provide to the target crop. This method has three phases: (1) creating a large backcross one (BC1) population to select, with markers, a reduced number of individuals (15–30, called the prIL set) with a low number of introgressions; (2) phenotyping the prIL set for the traits of interest and inferring the inheritance and map position of segregating major genes/QTLs based on the known genotypes of the prILs; and (3) advancing selected lines carrying the traits of interest to a next generation of backcross or selfing to obtain individuals with a single introgression in the background of the elite commercial germplasm. The proof of concept of this strategy was implemented by using peach as the recurrent species and almond as the donor. The whole process can be done in 9–10 years as the identification of the first line with one introgression was after 5 years (2006–2011), and 4–5 additional years are needed for phenotypic evaluation of selected lines. The expansion of this method to other perennial clonally propagated crops and to other species of Prunus compatible with peach is discussed.