Structure of the small nuclear RNP particle U1: identification of the two structural protuberances with RNP-antigens A and 70K

We have investigated the structure of the small nuclear RNP (snRNP) U1 by combining EM of complete and partially protein-deficient particles with immunoelectron microscopy employing mAbs against known components of the U1 snRNP. It was found that the two main protuberances of this particle can be identified with the U1-specific proteins A and 70K. The 70K protuberance is the one lying closer to the 5' terminus of the snRNA, as identified by its 5'-terminal m3G cap. The round-shaped main body of U1 snRNP represents its core RNP domain containing the common snRNP proteins. Functional implications of these results are discussed. Our results may also point to the physical basis for the production of autoantibodies directed against specific groups of snRNP proteins. The physical grouping of the common proteins (Sm epitopes) and the specific proteins (RNP epitopes) could result in one or the other being presented to the immune system as is the case in patients suffering from SLE or MCTD, respectively.     

Purification, cloning, and RXR identity of the HeLa cell factor with which RAR or TR heterodimerizes to bind target sequences efficiently.

We have purified and cloned a HeLa cell nuclear protein that strongly stimulates binding of retinoic acid and thyroid hormone receptors (RARs and TRs) to response elements. The purified protein is a human retinoid X receptor beta (hRXR beta). Three murine members of the RXR family (mRXR alpha, beta, and gamma) have also been cloned, and their interactions with RARs and TRs have been investigated. Under conditions where RAR, RXR, and TR bound poorly as homodimers to various response elements, strongly cooperative RAR-RXR and TR-RXR binding was observed. The binding efficiency was dependent on the sequence, relative orientation, and spacing of the repeated motifs of response elements. We show also that unstable RAR-RXR heterodimers were formed in solution, and that C-terminal sequences and the DNA-binding domains of both receptors were required for efficient formation of stable heterodimers on response elements. These findings suggest a convergence of the signaling pathways of some members of the nuclear receptor superfamily.     

Effect of pH on cell viability and product yields in d-xylose fermentations by Candida shehatae

 Candida shehatae were sequentially subjected to aerobic conditions for cellular growth, followed by anaerobic conditions for ethanol production from D-xylose at pH 2.5, 4.5 and 6.0. Ethanol yields increased from 0.25 g/g to 0.37 g/g and xylitol yields decreased from 0.33 g/g to 0.1 g/g as the pH was increased from 2.5 to 6.0. Cell viability, measured by plate counts and methylene blue staining, decreased in all of the fermentations, following the switch from aerobic to anaerobic conditions. However, pH 6.0 was shown to extend cell viability and increase the final ethanol concentration from 45 g/l to 55 g/l, compared to the yield at pH 4.5. Received: 25 April 1995/Received revision: 5 September 1995/Accepted: 20 September 1995     

Molecular analysis of cystinuria in Libyan Jews: exclusion of the SLC3A1 gene and mapping of a new locus on 19q.

Cystinuria is a hereditary disorder of amino acid transport and is manifested by the development of kidney stones. In some patients the disease is caused by mutations in the SLC3A1 gene, which is located on the short arm of chromosome 2 and encodes a renal/intestinal transporter for cystine and the dibasic amino acids. In Israel cystinuria is especially common among Jews of Libyan origin. After excluding SLC3A1 as the disease-causing gene in Libyan Jewish patients, we performed a genomewide search that shows that the Libyan Jewish cystinuria gene maps to the long arm of chromosome 19. Significant linkage was obtained for seven chromosome 19 markers. A maximal LOD score of 9.22 was obtained with the marker D19S882. Multipoint data and recombination analysis placed the gene in an 8-cM interval between the markers D19S409 and D19S208. Significant linkage disequilibrium was observed for alleles of four markers, and a specific haplotype comprising the markers D19S225, D19S208, D19S220, and D19S422 was found in 11 of 17 carrier chromosomes, versus 1 of 58 Libyan Jewish noncarrier chromosomes.     

Accelerated molecular dynamics simulations of the octopamine receptor using GPUs: discovery of an alternate agonist‐binding position

Octopamine receptors (OARs) perform key biological functions in invertebrates, making this class of G‐protein coupled receptors (GPCRs) worth considering for insecticide development. However, no crystal structures and very little research exists for OARs. Furthermore, GPCRs are large proteins, are suspended in a lipid bilayer, and are activated on the millisecond timescale, all of which make conventional molecular dynamics (MD) simulations infeasible, even if run on large supercomputers. However, accelerated Molecular Dynamics (aMD) simulations can reduce this timescale to even hundreds of nanoseconds, while running the simulations on graphics processing units (GPUs) would enable even small clusters of GPUs to have processing power equivalent to hundreds of CPUs. Our results show that aMD simulations run on GPUs can successfully obtain the active and inactive state conformations of a GPCR on this reduced timescale. Furthermore, we discovered a potential alternate active‐state agonist‐binding position in the octopamine receptor which has yet to be observed and may be a novel GPCR agonist‐binding position. These results demonstrate that a complex biological system with an activation process on the millisecond timescale can be successfully simulated on the nanosecond timescale using a simple computing system consisting of a small number of GPUs. Proteins 2016; 84:1480–1489. © 2016 Wiley Periodicals, Inc.     

Morphogenesis of sclerotome and neural crest in avian embryos

Summary The distribution of sclerotome and neural crest cells of avian embryos was studied by light and electron microscopy. Sclerotome cells radiated from the somites towards the notochord, to occupy the perichordal space. Neural crest cells, at least initially, also entered cell-free spaces. At the cranial somitic levels they moved chiefly dorsal to the somites, favouring the rostral part of each somite. These cells did not approach the perichordal space. More caudally (i.e. trunk levels), neural crest cells initially moved ventrally between the somites and neural tube. Adjacent to the caudal half of each somite, these cells penetrated no further than the myosclerotomal border, but opposite the rostral somite half, they were found next to the sclerotome almost as far ventrally as the notochord. However, they did not appear to enter the perichordal space, in contrast to sclerotome cells.When tested in vitro, sclerotome cells migrated towards notochords co-cultured on fibronectin-rich extracellular material, and on collagen gels. In contrast, neural crest cells avoided co-cultured notochords. This avoidance was abolished by inclusion of testicular hyaluronidase and chondroitinase ABC in the culture medium, but not by hyaluronidase from Streptomyces hyalurolyticus. The results suggest that sclerotome and neural crest mesenchyme cells have a different distribution with respect to the notochord, and that differential responses to notochordal extracellular material, possibly chondroitin sulphate proteoglycan, may be responsible for this.