Leptodora kindtii survival in the laboratory

Leptodora kindtii, a pelagic predatory cladoceran, suffers high mortality on transfer to laboratory, which makes the experimental work difficult. We investigated the causes of high mortality, using four variables: water volume, animal density, light intensity, and origin of water for culturing, i.e., water from native or a non-native lake. For the experiments we used Leptodora and water from Lake Loosdrecht and Lake Maarsseveen (The Netherlands). Water was found to be the most important factor; the animals did not necessarily do better in lake water from which they were collected. Water volume and animal density were of limited importance, and light intensity did not affect survival.     

1H-1H correlations across N–H•••N hydrogen bonds in nucleic acids

In 2HJ(NN)-COSY experiments, which correlate protons with donor/acceptor nitrogens across Nd...HNa bonds, the receptor nitrogen needs to be assigned in order to unambiguously identify the hydrogen bond. For many situations this is a non-trivial task which is further complicated by poor dispersion of (Na,Nd) resonances. To address these problems, we present pulse sequences to obtain direct, internucleotide correlations between protons in uniformly 13C/15N labeled nucleic acids containing Nd...HNa hydrogen bonds. Specifically, the pulse sequence H2(N1N3)H3 correlates H2(A,omega1):H3(U,omega2) protons across Watson-CrickA-U and mismatched G.A base pairs, the sequences H5(N3N1)H1/H6(N3N1)H1 correlate H5(C,omega1)/H6(C,omega1):H1(G,omega2) protons across Watson-Crick G-C base pairs, and the H2(N2N7)H8 sequence correlates NH2(G,A,C;omega1):H8(G,A;omega2) protons across G.G, A.A, sheared G.A and other mismatch pairs. These 1H-1H connectivities circumvent the need for independent assignment of the donor/acceptor nitrogen and related degeneracy issues associated with poorly dispersed nitrogen resonances. The methodology is demonstrated on uniformly 13C/15N labeled samples of (a) an RNA regulatory element involving the HIV-1 TAR RNA fragment, (b) a multi-stranded DNA architecture involving a G.(C-A) triad-containing G-quadruplex and (c) a peptide-RNA complex involving an evolved peptide bound to the HIV-1 Rev response element (RRE) RNA fragment.     

Novel 2D and 3D multiple-quantum bi-directional HCNCH experiments for the correlation of ribose and base protons/carbons in 13C/15N labeled RNA

Multiple-quantum 2D and 3D bi-directional HCNCH experiments are presented for the correlation of base and ribose protons/carbons in ¹³C/¹⁵N labeled HIV-1 TAR RNA. In both 2D and 3D experiments, the magnetization of H1 is transferred to H6/H8 and H1 through H1-C1-N1/9-C6/8-H6/8 and H1-C1-N1/9-C1-H1 pathways, and the magnetization of H6/8 is transferred to H1 and H6/8 through H6/8-C6/8-N1/9-C1-H1 and H6/8-C6/8-N1/9-C6/8-H6/8 pathways. Chemical shifts of four different nuclei (H1, C1, C6/8 and H6/8) are sampled in the 2D experiment. The correlation of base and ribose protons/carbons is established by the rectangular arrangement of crossover and out-and-back peaks in the proton/carbon correlated spectrum. The rectangular connections can be further resolved using the nitrogen dimension in a ¹H/¹³C/¹⁵N 3D experiment. Furthermore, by taking advantage of the well separated chemical shifts of N1 (pyrimidine) and N9 (purine), the 2D spectrum can be simplified into two sub-spectra based on their base type. Both experiments were tested on a ¹³C/¹⁵N labeled 27-mer HIV-1 TAR RNA containing a UUCG hairpin loop.     

Leptodora kindtii survival in the laboratory

Leptodora kindtii, a pelagic predatory cladoceran, suffers high mortality on transfer to laboratory, which makes the experimental work difficult. We investigated the causes of high mortality, using four variables: water volume, animal density, light intensity, and origin of water for culturing, i.e., water from native or a non-native lake. For the experiments we used Leptodora and water from Lake Loosdrecht and Lake Maarsseveen (The Netherlands). Water was found to be the most important factor; the animals did not necessarily do better in lake water from which they were collected. Water volume and animal density were of limited importance, and light intensity did not affect survival.     

Cyclopiazonic acid production by Aspergillus flavus and its effects on broiler chickens

Cyclopiazonic acid (CPA) was purified from cultures of Aspergillus flavus, and ca. 14 g of the toxin was collected for use in feeding studies. Chicken rations were artificially contaminated with purified CPA at concentrations of 10, 50, and 100 ppm (microgram/g) and fed ad libitum to eight groups of chickens for 7 weeks. Chickens receiving feed with 100 ppm of CPA had high mortality, decreased weight gain, and poor feed conversion when compared with birds receiving other doses. Postmortem examination showed that chickens fed the two greatest doses of CPA had proventricular lesions characterized by mucosal erosion and hyperemia (100 ppm) and by thick mucosa and dilated proventricular lumens (50 ppm). Birds given 100 ppm of CPA in feed also had numerous yellow foci in their livers and spleens. Microscopic examination of tissues of birds that received 100 ppm of CPA revealed ulcerative proventriculitis, mucosal necrosis in the gizzard, and hepatic and splenic necrosis and inflammation. Birds given 50 ppm of CPA had hyperplasia of the proventricular mucosal epithelium. Birds given 10 ppm of CPA and control birds had no significant treatment-related lesions.     

Female tannin avoidance: a possible explanation for habitat and dietary segregation of giraffes (Giraffa camelopardalis peralta) in Niger

Niger is host to the last free‐roaming herd of G.c. peralta (Giraffa camelopardalis peralta). We examined the foraging preferences of these giraffe in their dry‐season habitats, with the goal of preserving the herd in the regions that they currently inhabit. The current dry‐season habitat comprises two distinct vegetation zones. In both of these zones the giraffe must exist alongside the people of this region. The giraffes exhibit a sexual segregation in their dry‐season habitat selection and forage choices. The females show a strong preference for the intermediate zone (IM) when lactating. The males and pregnant females show a preference for the Dallol Bosso (DB). Nursing cows exhibit an avoidance of tannins. Bulls and non‐nursing cows prefer high protein and high fat forage, while subadults show a strong preference for high protein and carbohydrate contents and moderate tannin levels. Combretum glutinosum is the preferred species for adults of both sexes in the IM. Males and females have strong preferences for both Acacia nilotica and Acacia seyal in the DB. Sub‐adults of both sexes strongly prefer Prosopis africana in the IM. Unlike females, males retain their preference for A. nilotica when in the IM.     

Towards structural genomics of RNA: rapid NMR resonance assignment and simultaneous RNA tertiary structure determination using residual dipolar couplings

We report a new residual dipolar couplings (RDCs) based NMR procedure for rapidly determining RNA tertiary structure demonstrated on a uniformly (15)N/(13)C-labeled 27 nt variant of the trans-activation response element (TAR) RNA from HIV-I. In this procedure, the time-consuming nuclear Overhauser enhancement (NOE)-based sequential assignment step is replaced by a fully automated RDC-based assignment strategy. This approach involves examination of all allowed sequence-specific resonance assignment permutations for best-fit agreement between measured RDCs and coordinates for sub-structures in a target RNA. Using idealized A-form geometries to model Watson-Crick helices and coordinates from a previous X-ray structure to model a hairpin loop in TAR, the best-fit RDC assignment solutions are determined very rapidly (相似文献   

Peptide-triggered conformational switch in HIV-1 RRE RNA complexes

We have used NMR spectroscopy to determine the solution structure of a complex between an oligonucleotide derived from stem IIB of the Rev responsive element (RRE-IIB) of HIV-1 mRNA and an in vivo selected, high affinity binding Arg-rich peptide. The peptide binds in a partially alpha-helical conformation into a pocket within the RNA deep groove. Comparison with the structure of a complex between an alpha-helical Rev peptide and RRE-IIB reveals that the sequence of the bound peptide determines the local conformation of the RRE peptide binding site. A conformational switch of an unpaired uridine base was revealed; this points out into the solvent in the Rev peptide complex, but it is stabilized inside the RNA deep groove by stacking with an Arg side chain in the selected peptide complex. The conformational switch has been visualized by NMR chemical shift mapping of the uridine H5/H6 atoms during a competition experiment in which Rev peptide was displaced from RRE-IIB by the higher affinity binding selected peptide.     

Concerted motions in HIV-1 TAR RNA may allow access to bound state conformations: RNA dynamics from NMR residual dipolar couplings.

Ground-state dynamics in RNA is a critical precursor for structural adaptation observed ubiquitously in protein-RNA recognition. A tertiary conformational analysis of the stem-loop structural element in the transactivation response element (TAR) from human immunodeficiency virus type 1 (HIV-I) RNA is presented using recently introduced NMR methods that rely on the measurement of residual dipolar couplings (RDC) in partially oriented systems. Order matrix analysis of RDC data provides evidence for inter-helical motions that are of amplitude 46(+/-4) degrees, of random directional character, and that are executed about an average conformation with an inter-helical angle between 44 degrees and 54 degrees. The generated ensemble of TAR conformations have different organizations of functional groups responsible for interaction with the trans-activator protein Tat, including conformations similar to the previously characterized bound-state conformation. These results demonstrate the utility of RDC-NMR for simultaneously characterizing RNA tertiary dynamics and average conformation, and indicate an avenue for TAR complex formation involving tertiary structure capture.