Generalized lattice graphs for 2D-visualization of biological information

Several graph representations have been introduced for different data in theoretical biology. For instance, complex networks based on Graph theory are used to represent the structure and/or dynamics of different large biological systems such as protein-protein interaction networks. In addition, Randic, Liao, Nandy, Basak, and many others developed some special types of graph-based representations. This special type of graph includes geometrical constrains to node positioning in space and adopts final geometrical shapes that resemble lattice-like patterns. Lattice networks have been used to visually depict DNA and protein sequences but they are very flexible. However, despite the proved efficacy of new lattice-like graph/networks to represent diverse systems, most works focus on only one specific type of biological data. This work proposes a generalized type of lattice and illustrates how to use it in order to represent and compare biological data from different sources. We exemplify the following cases: protein sequence; mass spectra (MS) of protein peptide mass fingerprints (PMF); molecular dynamic trajectory (MDTs) from structural studies; mRNA microarray data; single nucleotide polymorphisms (SNPs); 1D or 2D-Electrophoresis study of protein polymorphisms and protein-research patent and/or copyright information. We used data available from public sources for some examples but for other, we used experimental results reported herein for the first time. This work may break new ground for the application of Graph theory in theoretical biology and other areas of biomedical sciences.     

Effect of estradiol on Ca<Superscript>2+</Superscript> release from intracellular stores in porcine oocytes stimulated by prolactin,theophylline, or guanosine triphosphate

The interaction between prolactin and theophylline as well as between prolactin and guanosine triphosphate during Ca²⁺ release from intracellular stores of estradiol-treated porcine oocytes isolated from the ovary at the stage of follicular growth were studied using fluorescent Ca²⁺-sensitive probe chlortetracycline. In the absence of estradiol, prolactin or theophylline induced Ca²⁺ release from intracellular stores; however, no increase in Ca²⁺ release was observed after their combined action. Conversely, Ca²⁺ release from intracellular stores increased only after the combined exposure to prolactin and theophylline in the presence of estradiol. In the absence of estradiol, guanosine triphosphate induced calcium release alone and together with prolactin. Protein kinase C regulated Ca²⁺ release from intracellular stores after the combined exposure to prolactin and theophylline only in the presence of estradiol; while the activation of protein kinase C required no estradiol during the combined exposure to prolactin and guanosine triphosphate. The data obtained indicate the effect of estradiol on Ca²⁺ release from intracellular stores after the combined exposure to prolactin and theophylline, while no such effect was observed after the combined exposure to prolactin and guanosine triphosphate.     

Role of individual R domain phosphorylation sites in CFTR regulation by protein kinase A

The cystic fibrosis transmembrane conductance regulator (CFTR) plays a critical role in transcellular ion transport and when defective, results in the genetic disease cystic fibrosis. CFTR is novel in the ATP-binding cassette superfamily as an ion channel that is enabled by a unique unstructured regulatory domain. This R domain contains multiple protein kinase A sites, which when phosphorylated allow channel gating. Most of the sites have been indicated to stimulate channel activity, while two of them have been suggested to be inhibitory. It is unknown whether individual sites act coordinately or distinctly. To address this issue, we raised monoclonal antibodies recognizing the unphosphorylated, but not the phosphorylated states of four functionally relevant sites (700, 737, 768, and 813). This enabled simultaneous monitoring of their phosphorylation and dephosphorylation and revealed that both processes occurred rapidly at the first three sites, but more slowly at the fourth. The parallel phosphorylation rates of the stimulatory 700 and the putative inhibitory 737 and 768 sites prompted us to reexamine the role of the latter two. With serines 737 and 768 reintroduced individually into a PKA insensitive variant, in which serines at 15 sites had been replaced by alanines, a level of channel activation by PKA was restored, showing that these sites can mediate stimulation. Thus, we have provided new tools to study the CFTR regulation by phosphorylation and found that sites proposed to inhibit channel activity can also participate in stimulation.     

Mechanisms of COPI vesicle formation

Coat Protein I (COPI) is one of the most intensely investigated coat complexes. Numerous studies have contributed to a general understanding of how coat proteins act to initiate intracellular vesicular transport. This review highlights key recent findings that have shaped our current understanding of how COPI vesicles are formed.     

A high-throughput assay for rapid and simultaneous analysis of perfect markers for important quality and agronomic traits in rice using multiplexed MALDI-TOF mass spectrometry

The application of single nucleotide polymorphisms (SNPs) in plant breeding involves the analysis of a large number of samples, and therefore requires rapid, inexpensive and highly automated multiplex methods to genotype the sequence variants. We have optimized a high-throughput multiplexed SNP assay for eight polymorphisms which explain two agronomic and three grain quality traits in rice. Gene fragments coding for the agronomic traits plant height (semi-dwarf, sd-1 ) and blast disease resistance ( Pi-ta ) and the quality traits amylose content ( waxy ), gelatinization temperature ( alk ) and fragrance ( fgr ) were amplified in a multiplex polymerase chain reaction. A single base extension reaction carried out at the polymorphism responsible for each of these phenotypes within these genes generated extension products which were quantified by a matrix-assisted laser desorption ionization-time of flight system. The assay detects both SNPs and indels and is co-dominant, simultaneously detecting both homozygous and heterozygous samples in a multiplex system. This assay analyses eight functional polymorphisms in one 5 µL reaction, demonstrating the high-throughput and cost-effective capability of this system. At this conservative level of multiplexing, 3072 assays can be performed in a single 384-well microtitre plate, allowing the rapid production of valuable information for selection in rice breeding.     

Characterization of eight single nucleotide polymorphism markers in Aedes aegypti

Sequencing of part of seven genes from Aedes aegypti collected in 16 Brazilian cities revealed the existence of 53 single nucleotide polymorphisms (SNPs), representing one SNP every 52 base pairs. From these 53 SNPs, we selected eight that are independent and highly polymorphic. We describe the use of these markers for differentiation of Brazilian populations of A. aegypti. These are the first SNPs developed for delineating population structure in A. aegypti, and will be a useful complement to epidemiological studies.     

Assessing statistical power of SNPs for population structure and conservation studies

Single nucleotide polymorphisms (SNPs) have been proposed by some as the new frontier for population studies, and several papers have presented theoretical and empirical evidence reporting the advantages and limitations of SNPs. As a practical matter, however, it remains unclear how many SNP markers will be required or what the optimal characteristics of those markers should be in order to obtain sufficient statistical power to detect different levels of population differentiation. We use a hypothetical case to illustrate the process of designing a population genetics project, and present results from simulations that address several issues for maximizing statistical power to detect differentiation while minimizing the amount of effort in developing SNPs. Results indicate that (i) while ~30 SNPs should be sufficient to detect moderate (F_ST = 0.01) levels of differentiation, studies aimed at detecting demographic independence (e.g. F_ST < 0.005) may require 80 or more SNPs and large sample sizes; (ii) different SNP allele frequencies have little affect on power, and thus, selection of SNPs can be relatively unbiased; (iii) increasing the sample size has a strong effect on power, so that the number of loci can be minimized when sample number is known, and increasing sample size is almost always beneficial; and (iv) power is increased by including multiple SNPs within loci and inferring haplotypes, rather than trying to use only unlinked SNPs. This also has the practical benefit of reducing the SNP ascertainment effort, and may influence the decision of whether to seek SNPs in coding or noncoding regions.     

Backbone assignment of HINT1 protein, a mouse histidine triad nucleotide binding protein

HINT1 is a mouse histidine triad nucleotide binding protein. Here we report the assignments for the backbone nitrogen, carbon and proton NMR signals.     

Non-nucleoside inhibitors binding to hepatitis C virus NS5B polymerase reveal a novel mechanism of inhibition

The RNA-dependent RNA polymerase (NS5B) from hepatitis C virus (HCV) is a key enzyme in HCV replication. NS5B is a major target for the development of antiviral compounds directed against HCV. Here we present the structures of three thiophene-based non-nucleoside inhibitors (NNIs) bound non-covalently to NS5B. Each of the inhibitors binds to NS5B non-competitively to a common binding site in the "thumb" domain that is approximately 35 Angstroms from the polymerase active site located in the "palm" domain. The three compounds exhibit IC(50) values in the range of 270 nM to 307 nM and have common binding features that result in relatively large conformational changes of residues that interact directly with the inhibitors as well as for other residues adjacent to the binding site. Detailed comparisons of the unbound NS5B structure with those having the bound inhibitors present show that residues Pro495 to Arg505 (the N terminus of the "T" helix) exhibit some of the largest changes. It has been reported that Pro495, Pro496, Val499 and Arg503 are part of the guanosine triphosphate (GTP) specific allosteric binding site located in close proximity to our binding site. It has also been reported that the introduction of mutations to key residues in this region (i.e. Val499Gly) ablate in vivo sub-genomic HCV RNA replication. The details of NS5B polymerase/inhibitor binding interactions coupled with the observed induced conformational changes provide new insights into the design of novel NNIs of HCV.