Ac2PIM-responsive miR-150 and miR-143 Target Receptor-interacting Protein Kinase 2 and Transforming Growth Factor Beta-activated Kinase 1 to Suppress NOD2-induced Immunomodulators

Specific and coordinated regulation of innate immune receptor-driven signaling networks often determines the net outcome of the immune responses. Here, we investigated the cross-regulation of toll-like receptor (TLR)2 and nucleotide-binding oligomerization domain (NOD)2 pathways mediated by Ac2PIM, a tetra-acylated form of mycobacterial cell wall component and muramyl dipeptide (MDP), a peptidoglycan derivative respectively. While Ac2PIM treatment of macrophages compromised their ability to induce NOD2-dependent immunomodulators like cyclooxygenase (COX)-2, suppressor of cytokine signaling (SOCS)-3, and matrix metalloproteinase (MMP)-9, no change in the NOD2-responsive NO, TNF-alpha, VEGF-A, and IL-12 levels was observed. Further, genome-wide microRNA expression profiling identified Ac2PIM-responsive miR-150 and miR-143 to target NOD2 signaling adaptors, RIP2 and TAK1, respectively. Interestingly, Ac2PIM was found to activate the SRC-FAK-PYK2-CREB cascade via TLR2 to recruit CBP/P300 at the promoters of miR-150 and miR-143 and epigenetically induce their expression. Loss-of-function studies utilizing specific miRNA inhibitors establish that Ac2PIM, via the miRNAs, abrogate NOD2-induced PI3K-PKC delta-MAPK pathway to suppress beta-catenin-mediated expression of COX-2, SOCS-3, and MMP-9. Our investigation has thus underscored the negative regulatory role of Ac2PIM-TLR2 signaling on NOD2 pathway which could broaden our understanding on vaccine potential or adjuvant utilities of Ac2PIM and/or MDP.

Variability in the air-sea interaction patterns and timescales within the south-eastern Bay of Biscay, as observed by HF radar data

Two high-frequency (HF) radar stations were installed on the coast of the south-eastern Bay of Biscay in 2009, providing high spatial and temporal resolution and large spatial coverage of currents in the area for the first time. This has made it possible to quantitatively assess the air-sea interaction patterns and timescales for the period 2009-2010. The analysis was conducted using the Barnett-Preisendorfer approach to canonical correlation analysis (CCA) of reanalysis surface winds and HF radar-derived surface currents. The CCA yields two canonical patterns: the first wind-current interaction pattern corresponds to the classical Ekman drift at the sea surface, whilst the second describes an anticyclonic/cyclonic surface circulation. The results obtained demonstrate that local winds play an important role in driving the upper water circulation. The wind-current interaction timescales are mainly related to diurnal breezes and synoptic variability. In particular, the breezes force diurnal currents in waters of the continental shelf and slope of the south-eastern Bay. It is concluded that the breezes may force diurnal currents over considerably wider areas than that covered by the HF radar, considering that the northern and southern continental shelves of the Bay exhibit stronger diurnal than annual wind amplitudes.

Feed conversion, protein efficiency, digestibility and growth performance of Oreochromis niloticus fed Delonix regia seed meal

Oreochromis niloticus fingerlings (mean weight 5.27~c0.29g) were fed raw and boiled Delonix regia seed meals following standard procedures. The weight gain, specific growth rate (SGR), feed conversion ratio (FCR), protein efficiency ratio (PER), net protein utilization (NPU) were determined as growth indices. Diet formulated with seed boiled for 80 minutes showed significantly (P<0.05) high values for the growth indices. Carcass nutrients composition were significantly (P<0.05) higher than in the control (raw) diet. Delonix regia seed meal when boiled has high potential of being utilized efficiently by O.niloticus. The implications of the respective index in fish metabolism are discussed

Chemical-scale studies of G protein-coupled receptors and ligand-gated ion channels

This dissertation describes studies of G protein-coupled receptors (GPCRs) and ligand-gated ion channels (LGICs) using unnatural amino acid mutagenesis to gain high precision insights into the function of these important membrane proteins.

Chapter 2 considers the functional role of highly conserved proline residues within the transmembrane helices of the D2 dopamine GPCR. Through mutagenesis employing unnatural α-hydroxy acids, proline analogs, and N-methyl amino acids, we find that lack of backbone hydrogen bond donor ability is important to proline function. At one proline site we additionally find that a substituent on the proline backbone N is important to receptor function.

In Chapter 3, side chain conformation is probed by mutagenesis of GPCRs and the muscle-type nAChR. Specific side chain rearrangements of highly conserved residues have been proposed to accompany activation of these receptors. These rearrangements were probed using conformationally-biased β-substituted analogs of Trp and Phe and unnatural stereoisomers of Thr and Ile. We also modeled the conformational bias of the unnatural Trp and Phe analogs employed.

Chapters 4 and 5 examine details of ligand binding to nAChRs. Chapter 4 describes a study investigating the importance of hydrogen bonds between ligands and the complementary face of muscle-type and α4β4 nAChRs. A hydrogen bond involving the agonist appears to be important for ligand binding in the muscle-type receptor but not the α4β4 receptor.

Chapter 5 describes a study characterizing the binding of varenicline, an actively prescribed smoking cessation therapeutic, to the α7 nAChR. Additionally, binding interactions to the complementary face of the α7 binding site were examined for a small panel of agonists. We identified side chains important for binding large agonists such as varenicline, but dispensable for binding the small agonist ACh.

Chapter 6 describes efforts to image nAChRs site-specifically modified with a fluorophore by unnatural amino acid mutagenesis. While progress was hampered by high levels of fluorescent background, improvements to sample preparation and alternative strategies for fluorophore incorporation are described.

Chapter 7 describes efforts toward a fluorescence assay for G protein association with a GPCR, with the ultimate goal of probing key protein-protein interactions along the G protein/receptor interface. A wide range of fluorescent protein fusions were generated, expressed in Xenopus oocytes, and evaluated for their ability to associate with each other.

Membrane activities of Dynamin-related protein 1 (DRP1) and BCL2-related Ovarian Killer (BOK)

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Squilla protein: chemical composition and nutritive value

Protein extract prepared from squilla (Grato squilla nepa), a commercially unexploited crustacean, was analysed for crude protein and essential amino acids. All the essential amino acids except tryptophan and threonine were present in nutritionally adequate amounts. The protein was evaluated for its nutritional quality in respect of growth rate, protein efficiency ratio (PER) and liver nitrogen content by feeding on rats. Growth rates and protein efficiency ratios were similar in rats fed on casein, squilla protein and a combination of squilla protein and casein (1:1) diet. The weight of liver and kidneys were normal.

The study of different protein in diet and water salinity on growth and survival rate in Pacific white shrimp (Litopenaeus vannamei)

In this research reared white western shrimp (Litopenaeus vannamei ,Boone, 1931) with five diet with five different protein level contain 20%, 25%, 30%, 35% and 40% and three salinity level contain 15-17 ppt, 27-30 ppt,and 40-45 ppt researched protein percent effect and water salinity on growth, survival, feed conversion ratio, hemolymph osmolatity, hemolymph protein and corpse protein contain. In this research was 15 sorrow with 3 repeat and used from 45 tanks with 300 liters capacity. Shrimps first weight average was about 2 grams and after 60 days culture cropped down results: Shrimps biomass growth in 15-17 ppt salinity was higher than anther salinities who had meaning different with growth in 40-45 ppt salinity ( p< 0.05). But hadn’t meaning different with growth 27-30 ppt salinity. survival rate in 15-17 ppt salinity was 97.03 who was lower than another salinities. survival percent in 24-30 ppt salinity and 40-45 ppt salinity was 99.33% Highest biomass growth in different diets was in diet number 5 with 40 percent protein that it had meaning different with another diets (p<0.05) . although with informed to product expense in different diets. One kilogram shrimp product expense in different diets hadn’t meaning different (P<0.05) Survival rate in different diets hadn’t meaning different lowest feed conversion ratio was 1.67 in 15-14 salinity that hadn’t meaning different with another salinities also corpse protein quantity in different salinities and different diets hadn’t meaning different. Hemolymph Osmolality in 15-17 ppt salinity was 573.88 mOsm/kg had meaning different with hemolymph osmolality in 27-30 ppt salinity that was 650. 380 mOsm/kg and in 40-45 ppt salinity was 630.38 mOsm/kg. Hemolymph protein in 15-17 ppt salinity was 124.72 mg/ml had meaning different with hemlymph protein in 27-30 ppt salinity that was 136.52 mg/ml but hadn’t meaning different with hemolymph protein in 40-45 ppt salinity that was 128.84 mg/ml. Hemolymph protein in different diets hadn’t meaning different (p<0.05). Keywords: shrimp, Litopenaeus vannamei, protein , salinity, growth, survival rate, FCR, hemolymph osmolality, hemolymph protein.