Correlations between the thermal stability of chloroplast (thylakoid) membranes and the composition and fluidity of their polar lipids upon acclimation of the higher plant,Nerium oleander,to growth temperature

The thermal stability of excitation transfer from pigment proteins to the Photosystem II reaction center of Nerium oleander adjusts by 10 Celsius degrees when cloned plants grown at 20°C/15°C, day/night growth temperatures are shifted to 45°C/32°C growth temperature or vice versa. Concomitant with this adjustment is a decrease in the fluidity of thylakoid membrane polar lipids as determined by spin labeling. The results are consistent with the hypothesis that there is a limiting maximum fluidity compatible with maintenance of native membrane structure and function. This limiting fluidity was about the same as for a number of other species which exhibit a range of thermal stabilities. Inversely correlated shifts in lipid fluidity and thermal stability occurred during the time course of acclimation of N. oleander to new growth temperatures. Thus, the temperature at which the limiting fluidity was reached changed during acclimation while the limiting fluidity remained constant. Although the relative proportion of the major classes of membrane polar lipids remained constant during adjustments in fluidity, large changes occured in the abundance of specific fatty acids. These changes were different for the phospho- and galacto-lipids suggesting that the fatty acid composition of these two lipid classes is regulated by different mechanisms. Comparisons between membrane lipid fluidity and fatty acid composition indicate that fluidity is not a simple linear function of fatty acid composition.     

cDNA cloning,expression, and enzymatic activity of a novel endogenous cellulase from the beetle Batocera horsfieldi

In this study, we report a novel cellulase [β-1,4-endoglucanase (EGase), EC 3.2.1.4] cDNA (Bh-EGase II) belonging to the glycoside hydrolase family (GHF) 45 from the beetle Batocera horsfieldi. The Bh-EGase II gene spans 720 bp and consists of a single exon coding for 239 amino acid residues. Bh-EGase II showed 93.72% protein sequence identity to Ag-EGase II from the beetle Apriona germari. The GHF 45 catalytic site is conserved in Bh-EGase II. Bh-EGase II has three putative N-glycosylation sites at 56–58 (N–K–S), 99–101 (N–S–T), and 237–239 (N–Y–S), respectively. The cDNA encoding Bh-EGase II was expressed in baculovirus-infected insect BmN cells and Bombyx mori larvae. Recombinant Bh-EGase II from BmN cells and larval hemolymph had an enzymatic activity of approximately 928 U/mg. The enzymatic catalysis of recombinant Bh-EGase II showed the highest activity at 50 °C and pH 6.0.     

Co-administration of live attenuated Salmonella enterica serovar Typhimurium expressing swine interleukin-18 and interferon-α provides enhanced Th1-biased protective immunity against inactivated vaccine of pseudorabies virus

The co-administration of two or more cytokines may generate additive or synergistic effects for controlling infectious diseases. However, the practical use of cytokine combinations for the modulation of immune responses against inactivated vaccine has not been demonstrated in livestock yet, primarily due to protein stability, production, and costs associated with mass administration. In light of the current situation, we evaluated the immunomodulatory functions of the combined administration of swine interleukin-18 (swIL-18) and interferon-α (swIFN-α) against an inactivated PrV vaccine using attenuated Salmonella enterica serovar Typhimurium as a cytokine delivery system. Co-administration of S. enterica serovar Typhimurium expressing swIL-18 and swIFN-α produced enhanced Th1-biased humoral and cellular immune responses against the inactivated PrV vaccine, when compared to single administration of S. enterica serovar Typhimurium expressing either swIL-18 or swIFN-α. Also, enhanced immune responses in co-administered piglets occurred rapidly after virulent PrV challenge, and piglets that received co-administration of S. enterica serovar Typhimurium expressing swIL-18 and swIFN-α displayed a greater alleviation of clinical severity following the virulent PrV challenge, as determined by clinical scores and cumulative daily weight gain. Furthermore, this enhancement was confirmed by reduced nasal shedding of PrV following viral challenge. Therefore, these results suggest that oral co-administration of S. enterica serovar Typhimurium expressing swIL-18 and swIFN-α provide enhanced Th1-biased immunity against inactivated PrV vaccine to alleviate clinical signs caused by PrV challenge.     

The Antibody Light-Chain Linker Regulates Domain Orientation and Amyloidogenicity

The antibody light chain (LC) consists of two domains and is essential for antigen binding in mature immunoglobulins. The two domains are connected by a highly conserved linker that comprises the structurally important Arg108 residue. In antibody light chain (AL) amyloidosis, a severe protein amyloid disease, the LC and its N-terminal variable domain (V_L) convert to fibrils deposited in the tissues causing organ failure. Understanding the factors shaping the architecture of the LC is important for basic science, biotechnology and for deciphering the principles that lead to fibril formation. In this study, we examined the structure and properties of LC variants with a mutated or extended linker. We show that under destabilizing conditions, the linker modulates the amyloidogenicity of the LC. The fibril formation propensity of LC linker variants and their susceptibility to proteolysis directly correlate implying an interplay between the two LC domains. Using NMR and residual dipolar coupling-based simulations, we found that the linker residue Arg108 is a key factor regulating the relative orientation of the V_L and C_L domains, keeping them in a bent and dense, but still flexible conformation. Thus, inter-domain contacts and the relative orientation of V_L and C_L to each other are of major importance for maintaining the structural integrity of the full-length LC.     

Facile removal of high mannose structures prior to extracting complex type N‐glycans from de‐N‐glycosylated peptides retained by C18 solid phase to allow more efficient glycomic mapping

The relative amount of high mannose structures within an N‐glycomic pool differs from one source to another, but quite often it predominates over the larger size complex type structures carrying biologically important glyco‐epitopes. An efficient method to separate these two classes of N‐glycans would significantly aid in detecting the lower abundant components by MS. Capitalizing on an initial observation that only high mannose type structures were recovered in the flow‐through fraction when peptide‐N‐glycosidase F digested peptides were passed through a C18 cartridge in 0.1% formic acid, we demonstrated here that native complex type N‐glycans can be retained by C18 cartridge and to be efficiently separated from both the smaller high mannose type structures, as well as de‐N‐glycosylated peptides by stepwise elution with increasing ACN concentration. The weak retention of the largely hydrophilic N‐glycans on C18 resin is dependent not only on size but also increased by the presence of α6‐fucosylation. This was shown by comparing the resulting N‐glycomic profiles of the washed and low‐ACN eluted fractions derived from both a human cancer cell line and an insect cell line.     

Fibroblast growth factor 16 and 18 are expressed in human cardiovascular tissues and induce on endothelial cells migration but not proliferation

Endothelial cells line the blood vessel and precursor endothelial cells appear to have a pivotal effect on the organ formation of the heart, the embryonic development of the kidney, and the liver. Several growth factors including the fibroblast growth factors (FGF) seem to be involved in these processes. Ligands such as basic FGF produced and secreted by endothelial cells may also coordinate cellular migration, differentiation, and proliferation under pathological conditions including wound healing, tumorgenesis, and fibrogenesis in the adult. Recently we demonstrated the expression of two secreted FGFs, FGF16, and FGF18, in HUVEC and in rat aortic tissue. In the present report, we confirmed by RT-PCR analysis that FGF18 is wildly expressed in the cardiovascular tissue, while FGF16 showed a more restricted expression pattern. HUVEC clearly demonstrated chemotaxis towards FGF16 and FGF18. Both FGFs also enhanced cell migration in response to mechanical damage. However, recombinant FGF16 and FGF18 failed to induce endothelial cell proliferation or sprouting in a three-dimensional in vitro angiogenesis assay. Fgf18 expression was earlier reported in the liver, and we detected FGF18 expression in liver vascular and liver sinusoidal endothelial cells (LSECs), but not in hepatic parenchymal cells. Recombinant FGF18 stimulated DNA synthesis in primary hepatocytes, suggesting, that endothelial FGF18 might have a paracrine function in promoting growth of the parenchymal tissue. Interestingly, FGF2, which is mitogenic on endothelial cells and hepatocytes stimulates a sustained MAPK activation in both cell types, while FGF18 causes a short transient activation of the MAPK pathway in endothelial cells but a sustained activation in hepatocytes. Therefore, the difference in the time course of MAPK activation by the different FGFs appears to be the cause for the different cellular responses.     

Site-specific cleavage of the 40S ribosomal subunit reveals eukaryote-specific ribosomal protein S28 in the subunit head

After resolving the crystal structure of the prokaryotic ribosome, mapping the proteins in the eukaryotic ribosome is a challenging task. We applied RNase H digestion to split the human 40S ribosomal subunit into head and body parts. Mass spectrometry of the proteins in the 40S subunit head revealed the presence of eukaryote-specific ribosomal protein S28e. Recombinant S28e was capable of specific binding to the 3′ major domain of the 18S rRNA (K_a = 8.0 ± 0.5 × 10⁹ M⁻¹). We conclude that S28e has a binding site on the 18S rRNA within the 40S subunit head.

Structured summary

MINT-8044084: S8 (uniprotkb:P62241) and S19 (uniprotkb:P39019) colocalize (MI:0403) by cosedimentation through density gradient (MI:0029)MINT-8044095: S8 (uniprotkb:P62241), S19 (uniprotkb:P39019) and S13 (uniprotkb:P62277) colocalize (MI:0403) by cosedimentation through density gradient (MI:0029)MINT-8044024: S29 (uniprotkb:P62273), S28 (uniprotkb:P62857), S21 (uniprotkb:P63220), S20 (uniprotkb:P60866), S26 (uniprotkb:P62854), S25 (uniprotkb:P62851), S12 (uniprotkb:P25398), S17 (uniprotkb:P08708), S19 (uniprotkb:P39019), S14 (uniprotkb:P62263), S16 (uniprotkb:P62249) and S11 (uniprotkb:P62280) colocalize (MI:0403) by cosedimentation through density gradient (MI:0029)MINT-8044065: S29 (uniprotkb:P62273), S28 (uniprotkb:P62857), S19 (uniprotkb:P39019), S14 (uniprotkb:P62263) and S16 (uniprotkb:P62249) colocalize (MI:0403) by cosedimentation through density gradient (MI:0029)     

Morphological, morphometric, meristic and osteological evidence for two species of hake (Actinopterygii: Gadiformes: Merluccius) in Argentinean waters

Morphologically, both classic and landmark-based morphometry and meristic analyses of 241 specimens of Merluccius, along with the re-examination of six paratype specimens of Merluccius hubbsi, the holotype and three paratypes of Merluccius patagonicus and the syntype of Merluccius australis revealed the presence of only two species of Merluccius in Argentinean waters. Internal structures (hyomandibula, urohyal and sagitta otolith) of M. hubbsi were compared to those reported for M. patagonicus and were shown to have identical morphology. Type specimens of M. patagonicus showed a complete overlap in morphometric and meristic characters with M. hubbsi, whereas M. australis had a greater number of second dorsal and anal-fin rays, and more lateral-line scales. In addition, M. australis had a smaller eye and longer snout than M. hubbsi and M. patagonicus. The results indicate that there is no evidence for a third hake species in Argentinean waters. Merluccius patagonicus is a synonym of M. hubbsi.     

Complete inhibition of in vivo glioma growth by oncostatin M

We describe here the oncostatin M (OSM)-dependent inhibition of in vivo tumour formation after intracerebral inoculation of glioblastoma cells in mice. We generated human glioblastoma cells transfected with the OSM gene under the control of a tetracycline-response promoter. Upon removal of tetracycline from the medium, cells exhibited a differentiated cell morphology, while proliferation was significantly inhibited. After implantation of these cells into nude mice brains, large tumours developed in animals lacking OSM expression, whereas no tumour formation was observed in mice with induced OSM expression. Our results suggest that OSM exerts pronounced antitumorigenic effects on glioblastoma cells in vivo and provide arguments for a therapeutic application of OSM in humans.