Progesterone Receptor Membrane Component 1 Is a Functional Part of the Glucagon-like Peptide-1 (GLP-1) Receptor Complex in Pancreatic β Cells

Glucagon-like peptide-1 (GLP-1) is an incretin hormone that regulates glucose homeostasis. Because of their direct stimulation of insulin secretion from pancreatic β cells, GLP-1 receptor (GLP-1R) agonists are now important therapeutic options for the treatment of type 2 diabetes. To better understand the mechanisms that control the insulinotropic actions of GLP-1, affinity purification and mass spectrometry (AP-MS) were employed to uncover potential proteins that functionally interact with the GLP-1R. AP-MS performed on Chinese hamster ovary cells or MIN6 β cells, both expressing the human GLP-1R, revealed 99 proteins potentially associated with the GLP-1R. Three novel GLP-1R interactors (PGRMC1, Rab5b, and Rab5c) were further validated through co-immunoprecipitation/immunoblotting, fluorescence resonance energy transfer, and immunofluorescence. Functional studies revealed that overexpression of PGRMC1, a novel cell surface receptor that associated with liganded GLP-1R, enhanced GLP-1-induced insulin secretion (GIIS) with the most robust effect. Knockdown of PGRMC1 in β cells decreased GIIS, indicative of positive interaction with GLP-1R. To gain insight mechanistically, we demonstrated that the cell surface PGRMC1 ligand P4-BSA increased GIIS, whereas its antagonist AG-205 decreased GIIS. It was then found that PGRMC1 increased GLP-1-induced cAMP accumulation. PGRMC1 activation and GIIS induced by P4-BSA could be blocked by inhibition of adenylyl cyclase/EPAC signaling or the EGF receptor–PI3K signal transduction pathway. These data reveal a dual mechanism for PGRMC1-increased GIIS mediated through cAMP and EGF receptor signaling. In conclusion, we identified several novel GLP-1R interacting proteins. PGRMC1 expressed on the cell surface of β cells was shown to interact with the activated GLP-1R to enhance the insulinotropic actions of GLP-1.Glucagon-like peptide-1 (GLP-1)¹ is a gastrointestinal hormone secreted by intestinal L cells upon food intake that is best known for its role in controlling glucose homeostasis. Acting through its cognate glucagon-like peptide-1 receptor (GLP-1R), GLP-1 has several important physiological and pharmacological functions. GLP-1 is best known for enhancing glucose-stimulated insulin secretion (GSIS) from the pancreatic β cells. Importantly, the insulinotropic properties of GLP-1 are maintained in patients with type 2 diabetes (1), which is characterized by insufficient insulin secretion from pancreatic β cells and an inability to maintain glucose homeostasis. Therefore, therapeutic strategies targeting GLP-1R have been developed to treat type 2 diabetes (2, 3). In addition to augmenting insulin secretion, GLP-1 has been known to improve glucose sensing, proinsulin biosynthesis, survival, and proliferation of β cells (3, 4) in a variety of experimental models. GLP-1 also has several extrapancreatic effects, including actions on the central nervous system to inhibit food intake (5), the stomach to decrease gastric emptying and gastric acid secretion (6), and the lungs to stimulate secretion of macromolecules from airways (7). Additionally, GLP-1 has an effect on the heart and possibly the kidney to modulate blood pressure and heart rate (8, 9).The GLP-1R is a member of the B1 family of G protein–coupled receptors (secretin receptor family). In mammals, GLP-1R is expressed in multiple tissues, including pancreatic β cells and δ cells (10), hypothalamus, lung, stomach, heart, kidney (11), and thyroid (12), which in part explains its diverse actions. Upon ligand binding, the GLP-1R is capable of coupling to diverse cell signal transduction pathways, but it is best known for its actions on G protein Gs α and adenylate cyclase activity to increase intracellular cAMP. It is known that other proteins can affect GLP-1R activity in addition to G proteins, including β-arrestin and caveolin, which affect receptor internalization and trafficking. β-Arrestin 1 is also required for proper GLP-1-stimulated cAMP production (13–15). More recently, it was shown that another B1 family member, gastric inhibitory polypeptide receptor heterodimerizes with GLP-1R, decreasing GLP-1-induced β-arrestin recruitment and mobilization (16). Very recently, our group identified several novel potential GLP-1R interactors using a membrane-based split-ubiquitin yeast two-hybrid (MYTH) assay (17). Three β cell–expressing membrane-bound interactors, solute carrier family 15 member 4 (SLC15A4), amyloid β A4 precursor-like protein 1 (APLP1), and adaptor-related protein complex 2 subunit mu (AP2M1), were further selected for individual knockdown in mouse insulinoma (MIN6) β cells using small interfering RNAs (siRNAs). GLP-1-induced insulin secretion was significantly enhanced when these genes were silenced, suggesting that these interactor proteins attenuate GLP-1R activity. These findings demonstrated that GLP-1R protein interactions are complex and the interactors can have measurable effects on receptor trafficking and downstream signaling. Such interactions may in part explain the diverse tissue-specific effects of GLP-1 and offer avenues for controlling GLP-1 actions in a tissue-selective manner.Although the MYTH system is well established (18) and has been applied to study G protein–coupled receptor interactomes (17), it is limited on two fronts. Firstly, it must be performed in yeast which is not an ideal representation of the mammalian system. Secondly, it is technically difficult to activate the receptor in MYTH, thus, effects of ligand stimulation on the receptor interactome cannot be assessed. Recently, affinity purification–mass spectrometry (AP-MS) has become a powerful tool for discovering and examining novel protein–protein interactions, including those between membrane-bound proteins in mammalian cells (19–21). In the current study, we applied AP-MS to discover novel GLP-1R interactors and employed a human GLP-1R harboring a FLAG^® epitope. GLP-1R-Flag was expressed in either Chinese hamster ovary (CHO) cells or MIN6 β cells, and interactors were studied in the presence or absence of GLP-1.     

Stand density and species richness affect carbon storage and net primary productivity in early and late successional temperate forests differently

How stand density and species richness affect carbon (C) storage and net primary productivity (NPP) changes with forest succession is poorly understood. We quantified the C storage of trees and the aboveground NPP in an early successional secondary birch forest (birch forest) and a late successional mixed broadleaf-Korean pine (Pinus koraiensis) forest (mixed forest) in northeastern China. We found that: 1) tree C storage in the mixed forest (120.3 Mg C ha^?1) was significantly higher than that in the birch forest (78.5 Mg C ha^?1), whereas the aboveground NPP was not different between the two forest types; and 2) only stand density had a positive linear relationship with tree C storage and aboveground NPP in the birch forest. In the mixed forest, both tree C storage and aboveground NPP were significantly affected by the combination of the stand density and species richness. The tree C storage to stand density and species richness relationships were hump-shaped. The aboveground NPP increased with increasing stand density, but its relationship to species richness was hump-shaped. We conclude that the effect of stand density and species richness on tree C storage and aboveground NPP was influenced by forest stand succession, and such effects should be considered in studying stand density- and species richness- ecosystem function (e.g., C storage and NPP) relationships in temperate forest ecosystems.     

Isolation and Identification of Pathogens Causing Marigold (Tagetes erecta L.) Black Spot in Beijing,China

Black spot leads to great marigold losses worldwide. The disease is characterized by black spots on leaves and stems in its early stages, and the whole plant has black rot at the advanced stage. In this report, 6 of 217 Alternaria strains isolated from lesions of marigold plants in Beijing were randomly selected. The morphological characteristics and a pathogenic tree based on two protein‐coding genes (gpd and alt a 1) indicated that Alternaria tagetica is the causal agent of marigold black spot in Beijing. All six Alternaria strains could successfully re‐infect marigold, but they could not infect carrot or zinnia by either spore spray in a greenhouse or planting experiments in the epidemic area. This is the first report of the A. tagetica pathogen being isolated from marigold in Beijing.     

Ontogenetic changes in anti-herbivore defensive traits in leaves of four Mediterranean co-occurring <Emphasis Type="Italic">Quercus</Emphasis> species

Investment in anti-herbivore defence in tree species has been one of the priority research topics in plant terrestrial ecology during the last decades. However, despite considerable experimental effort, interspecific differences in the ontogenetic trends in the investment in defence are still a matter of debate, as to date experimental evidence is contradictory. In the present work, insect herbivory levels were measured in seedlings and mature trees of four co-occurring Mediterranean Quercus species with differing leaf life spans, as well as several leaf characteristics that can determine herbivore preference. The measured leaf traits included nitrogen (N), fibre (cellulose, hemicellulose and lignin), total phenolic contents, leaf mass per unit area (LMA) and leaf thickness. The leaves of seedlings had a lower LMA and leaf thickness and lower concentrations of N and cellulose, but higher concentrations of lignin and phenols than those of mature trees. However, the loss of leaf area tended to be more severe for seedlings than for mature trees, although the differences were only significant for deciduous species. This constitutes a confirmation of the strong effects of physical traits on herbivore preferences. The greater resource limitations for defensive mechanisms in seedlings with respect to mature trees would explain that at intraspecific level we do observe a compromise between chemical and physical defences. As a result, seedlings rely on chemical rather than on physical defences.     

西洋参胚性细胞和胚状体细胞中的核内含体与细胞质内含体

１９２１年,Ｍｏｌｉｓｈ首先在光镜下发现植物叶片细胞核中有一种晶体状的内含物（见Ｗｅｉｎｔｒａｕｂ等［１］）。后来人们陆续在某些动、植物的细胞核中观察到了这种结构［２—４］,并称之为核内含体（ｉｎｔｒａｎｕｃｌｅａｒｉｎｃｌｕｓｉｏｎｓ）。Ｂｉｇａｚｚｉ［３］曾在４５种桔梗科植物的细胞中看到了核内含体。但在离体培养的植物细胞中发现内含体的报道很少,至今仅在榛子组织培养分生细胞中看到了类似的结构［５］。我们在对西洋参体细胞胚胎发生进行超微结构研究的过程中发现,在胚性愈伤组织和胚状体细胞的核和细胞质…     

<Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> inhibits <Emphasis Type="Italic">in-vitro Candida</Emphasis> biofilm development

Background  

Elucidation of the communal behavior of microbes in mixed species biofilms may have a major impact on understanding infectious diseases and for the therapeutics. Although, the structure and the properties of monospecies biofilms and their role in disease have been extensively studied during the last decade, the interactions within mixed biofilms consisting of bacteria and fungi such as Candida spp. have not been illustrated in depth. Hence, the aim of this study was to evaluate the interspecies interactions of Pseudomonas aeruginosa and six different species of Candida comprising C. albicans, C. glabrata, C. krusei, C. tropicalis, C. parapsilosis, and C. dubliniensis in dual species biofilm development.     

Isoprene emission protects photosynthesis in sunfleck exposed Grey poplar

In the present study, we combined transient temperature and light stress (sunfleck) and comparably analyzed photosynthetic gas exchange in Grey poplar which has been genetically modified in isoprene emission capacity. Overall, we demonstrate that for poplar leaves the ability to emit isoprene is crucial to maintain photosynthesis when exposed to sunflecks. Net CO₂ assimilation and electron transport rates were strongly impaired in sunfleck-treated non-isoprene emitting poplars. Similar impairment was not detected when the leaves were exposed to high light (lightflecks) only. Within 10 h non-isoprene emitting poplars recovered from sunfleck-related impairment as indicated by chlorophyll fluorescence and microarray analysis. Unstressed leaves of non-isoprene emitting poplars had higher ascorbate contents, but also higher contents of malondialdehyde than wild-type. Microarray analyses revealed lipid and chlorophyll degradation processes in the non-isoprene emitting poplars. Thus, there is evidence for an adjustment of the antioxidative system in the non-isoprene emitting poplars even under normal growth conditions.     

Host Age Preference Behavior in <Emphasis Type="Italic">Aphidius ervi</Emphasis> Haliday (Hymenoptera: Aphidiidae)

Many parasitoid species have preference for certain stages of hosts to parasitize but the underlying behavioral mechanisms of such preference are still poorly understood, making it difficult to evaluate host-parasitoid interactions and their effects on the success of biological control programs. Here, we report our work on a parasitoid Aphidius ervi Haliday on the pea aphid Acyrthosiphon pisum (Harris). We show that with the increase of host age, female parasitoids are more likely to encounter and to attack their hosts but the hosts develop increasingly greater defensive capabilities. Encounter almost always triggers attack attempt; however, increasing attack attempts do not proportionally lead to ovipositor probings and increasing ovipositor probings do not proportionally translate into ovipositions. These asymmetric responses may be interpreted as that A. ervi females prefer to parasitize older aphids for higher fitness return but those aphids can better defend themselves, and as a consequence, A. ervi females may achieve the highest gain by attacking aphids of intermediate ages. We suggest that A. ervi females forage in a manner consistent with the optimal foraging theory, trading off host handling time with fitness returns.     

The invasive Yellow Crazy Ant and the decline of forest ant diversity in Indonesian cacao agroforests

Throughout the tropics, agroforests are often the only remaining habitat with a considerable tree cover. Agroforestry systems can support high numbers of species and are therefore frequently heralded as the future for tropical biodiversity conservation. However, anthropogenic habitat modification can facilitate species invasions that may suppress native fauna. We compared the ant fauna of lower canopy trees in natural rainforest sites with that of cacao trees in agroforests in Central Sulawesi, Indonesia in order to assess the effects of agroforestry on occurrence of the Yellow Crazy Ant Anoplolepis gracilipes, a common invasive species in the area, and its effects on overall ant richness. The agroforests differed in the type of shade-tree composition, tree density, canopy cover, and distance to the village. On average, 43% of the species in agroforests also occurred in the lower canopy of nearby primary forest and the number of forest ant species that occurred on cacao trees was not related to agroforestry characteristics. However, A. gracilipes was the most common non-forest ant species, and forest ant richness decreased significantly with the presence of this species. Our results indicate that agroforestry may have promoted the occurrence of A. gracilipes, possibly because tree management in agroforests negatively affects ant species that depend on trees for nesting and foraging, whereas A. gracilipes is a generalist when it comes to nesting sites and food preference. Thus, agroforestry management that includes the thinning of tree stands can facilitate ant invasions, thereby threatening the potential of cultivated land for the conservation of tropical ant diversity.