Comparative studies of skeletal soluble matrices from some Scleractinian corals and Molluscs

The soluble organic matrices extracted from aragonitic skeletons of five Scleractinia and two Molluscs were analyzed using FTIR spectrometry, HPLC and 2-D gel electrophoresis. All these methods show that scleractinian and molluscan matrices are different. Both matrices are acidic with poorly separated molecular weights. The scleractinian matrices are highly glycosylated, whereas the molluscan matrices are not as shown by stains in 2D-electrophoresis. These results were in accordance with earlier data: high S contents in Scleractinia skeletons and low S contents in Mollusca shells. These results confirm the control of the morphology and the chemical composition of aragonite biocrystals in various taxa.     

Complete mitochondrial genomes of two Japanese precious corals, Paracorallium japonicum and Corallium konojoi (Cnidaria, Octocorallia, Coralliidae): notable differences in gene arrangement

Precious coral are taxonomically a group of corals that belong to the family Coralliidae within the order Alcyonacea, subclass Octocorallia, and class Anthozoa, whose skeletal axes are used for jewelry. They are distributed in the Mediterranean Sea and in waters adjacent to Japan, Taiwan, Midway Island and the Hawaiian Islands. The genus Corallium of the family Coralliidae was recently divided into two genera, Corallium and Paracorallium, based on morphological observations, but insufficient molecular evidence to support this classification has been presented to date. We determined for the first time the complete mitochondrial genome sequence of two precious corals P. japonicum and C. konojoi, in order to clarify their systematic positions. The circular mitochondrial genomes of P. japonicum and C. konojoi are 18,913bp and 18,969bp in length, respectively, and encode 13 typical energy pathway protein coding genes (nad1-6, nad4L, cox1-3, cob, atp6 and atp8), two ribosomal RNA genes (rns and rnl), a transfer RNA (trnM) and a mismatch repair gene homologue msh1. The two genomes have an overall nucleotide sequence identity of 97.5%, which is comparable to that between Acanella eburnea and Keratoisidinae sp. belonging to Octocorallia. Surprisingly, however, their gene arrangements were not identical. Phylogenetic analyses using seven complete mitochondrial genome sequences belonging to species in the subclass Octocorallia indicated that within the subclass, at least three gene order rearrangement events occurred during evolution. Our results support the validity of the morphological classification that separated the family Coralliidae into two genera, Corallium and Paracorallium.     

Comparing G matrices: are common principal components informative?

Common principal components (CPC) analysis is a technique for assessing whether variance-covariance matrices from different populations have similar structure. One potential application is to compare additive genetic variance-covariance matrices, G. In this article, the conditions under which G matrices are expected to have common PCs are derived for a two-locus, two-allele model and the model of constrained pleiotropy. The theory demonstrates that whether G matrices are expected to have common PCs is largely determined by whether pleiotropic effects have a modular organization. If two (or more) populations have modules and these modules have the same direction, the G matrices have a common PC, regardless of allele frequencies. In the absence of modules, common PCs exist only for very restricted combinations of allele frequencies. Together, these two results imply that, when populations are evolving, common PCs are expected only when the populations have modules in common. These results have two implications: (1) In general, G matrices will not have common PCs, and (2) when they do, these PCs indicate common modular organization. The interpretation of common PCs identified for estimates of G matrices is discussed in light of these results.     

Complete mitochondrial genomes of the Japanese pink coral (Corallium elatius) and the Mediterranean red coral (Corallium rubrum): a reevaluation of the phylogeny of the family Coralliidae based on molecular data

Precious corals are soft corals belonging to the family Coralliidae (Anthozoa: Octocorallia: Alcyonacea) and class Anthozoa, whose skeletal axes are used for jewelry.The family Coralliidae includes ca. 40 species and was originally thought to comprise of the single genus Corallium. In 2003, Corallium was split into two genera, Corallium and Paracorallium, and seven species were moved to this newly identified genus on the bases of morphological features. Previously, we determined the complete mitochondrial genome sequence of two precious corals Paracorallium japonicum and Corallium konojoi, in order to clarify their systematic positions. The two genomes showed high nucleotide sequence identity, but their gene order arrangements were not identical. Here, we determined three complete mitochondrial genome sequences from the one specimen of Mediterranean Corallium rubrum and two specimens of Corallium elatius coming from Kagoshima (South Japan). The circular mitochondrial genomes of C. rubrum and C. elatius are 18,915 bp and 18,969–18,970 bp in length, respectively, and encode 14 typical octocorallian protein-coding genes (nad1–6, nad4L, cox1–3, cob, atp6, atp8, and mtMutS, which is an octocoral-specific mismatch repair gene homologue), two ribosomal RNA genes (rns and rnl), and one transfer RNA (trnM). The overall nucleotide differences between C. konojoi and each C. elatius haplotype (T2007 and I2011) are only 10 and 11 nucleotides, respectively; this degree of similarity indicates that C. elatius and C. konojoi are very closely related species. Notably, the C. rubrum mitochondrial genome shows more nucleotide sequence identity to P. japonicum (99.5%) than to its congeneric species C. konojoi (95.3%) and C. elatius (95.3%). Moreover, the gene order arrangement of C. rubrum was the same as that of P. japonicum, while that of C. elatius was the same as C. konojoi. Phylogenetic analysis based on three mitochondrial genes from 24 scleraxonian species shows that the family Coralliidae is separated into two distinct groups, recovering Corallium as a paraphyletic genus. Our results indicate that the currently accepted generic classification of Coralliidae should be reconsidered.     

Bifunctional monolithic affinity hydrogels for dual-protein delivery

Multiple-protein delivery has been proven to be a critical consideration for promoting tissue regeneration. Many polymeric composite biomaterials have been designed and used for modulating dual-protein delivery to enhance tissue regeneration in vitro or in vivo. However, the fabrication conditions and low water contents within the portions of these composite matrices that determine protein release rates are not optimal for maintaining the stability of encapsulated macromolecular therapeutics. In this proof-of-concept work, we aim to resolve this deficiency by single-step fabrication of affinity hydrogels capable of independently delivering two or more proteins. Selective protein-binding sites were incorporated into poly(ethylene glycol) hydrogels via copolymerization with glycidyl methacrylate-iminodiacetic acid (GMIDA) ligands to modulate release of two model proteins, lysozyme and hexahistidine tagged green fluorescent protein (hisGFP), via two distinct matrix-binding mechanisms, namely electrostatic interaction and metal-ion chelation. Differing from composite matrices for dual-protein delivery, the results reported herein indicate that injectable monolithic affinity hydrogels are capable of rapidly encapsulating multiple therapeutic agents under mild physiological conditions and independently controlling their localized delivery. Most importantly, these affinity hydrogels retain high water permeabilities throughout the entire device, characteristics that are necessary for maintaining the stability and viability of encapsulated proteins and cells.     

Comparison of the soluble organic matrices of healthy and diseased shells of Pinctada margaritifera (L.) and Pecten maximus L. (Mollusca, Bivalvia)

Pinctada margaritifera and Pecten maximus are among the mollusks of commercial value. Both are known to show abnormal calcification processes that strongly increase the mortality rate. Several parameters of the soluble organic matrices extracted from the shells of P. margaritifera and P. maximus are analyzed: bulk composition, molecular weights, and acidity. The composition of the matrices of healthy and diseased shells were compared, showing that the protein and the sugar contents are variously modified. Differences between healthy and diseased shells are dissimilar in the two species. This is in accordance with the previously described macroscopic and microscopic alterations (red color, numerous brown membranes). This study does not allow identification the origin of the disease, but provides new insights on the role of sugars in biomineralization processes.     

Morphological and molecular characterization of a new genus and new species of parazoanthid (Anthozoa: Hexacorallia: Zoantharia) associated with Japanese Red Coral

The Order Zoantharia has long been taxonomically neglected primarily due to difficulty in examining the internal morphology of sand-encrusted zoanthids. However, recent work using molecular markers has shown an unexpectedly high diversity of previously “hidden” taxa (families and genera) within Zoantharia (=Zoanthidea, Zoanthiniaria). In this study, unidentified sediment-encrusting zoanthid specimens (n = 8) were collected from living Japanese Red Coral Paracorallium japonicum (Family Coralliidae) during precious coral harvesting by Remotely Operated Vehicle (ROV) and manned submersible (February 2004–January 2006) at depths of 194–250 m at six locations between Ishigaki-jima Island and Kikai-jima Island, southern Japan. DNA sequences (mitochondrial 16S ribosomal DNA [mt 16S rDNA], cytochrome oxidase subunit I [COI], nuclear internal transcribed spacer of ribosomal DNA [ITS-rDNA]) unambiguously place these specimens in a previously undescribed, new monophyletic lineage within the family Parazoanthidae. Corallizoanthus tsukaharai, gen. n. et sp. n. is the first reported zoanthid species associated with the family Coralliidae and unlike other described gorgonian-associated zoanthids (Savalia spp.) does not secrete its own hard axis. Morphologically, C. tsukaharai sp. n. is characterized by generally unitary polyps and bright yellow external coloration. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. Communicated by Biology Editor Dr Ruth Gates     

Plasticization, antiplasticization, and molecular packing in amorphous carbohydrate-glycerol matrices

The molecular packing of amorphous maltodextrin-glycerol matrices is systematically explored by combining positron annihilation lifetime spectroscopy (PALS) with thermodynamic measurements and dilatometry. Maltodextrin-glycerol matrices are equilibrated at a range of water activities between 0 and 0.54 at T = 25 °C to analyze the effect of both water and glycerol on the average molecular hole size and the specific volume of the matrices. In the glassy state, glycerol results in a systematic reduction of the average molecular hole size. In contrast, water interacts with the carbohydrate matrix in a complex way. Thermodynamic clustering theory shows that, at very low water contents the water molecules are well dispersed and are closely associated with the carbohydrate chains. In this regime water acts as an antiplasticizer, whereby it reduces the size of the molecular holes. Conversely, at higher water contents, while still in the glassy state, water acts as a plasticizer by increasing the average hole volume of the carbohydrate matrices. This plasticization-dominated mechanism is likely to be due to the interplay between the ability of water to form hydrogen bonds with the hydroxyl residues on the carbohydrate chains and its mobility, which is significantly decoupled from the bulk mobility of the matrix. Our findings are of key importance for the understanding of the effect of glycerol on the biostabilization performance of these carbohydrate matrices, as it provides a first insight on how molecular packing can relate to the dynamics in such matrices.     

Structure and composition of the aragonitic crossed lamellar layers in six species of Bivalvia and Gastropoda

The microstructures, the chemical composition and the soluble organic matrices of the aragonitic crossed lamellar layers of the shells of six species of molluscs have been studied. The microstructures and chemical contents are similar, whereas the quantities of organic matrices are variable. All the soluble matrices are glycoproteins, with low S contents. Their molecular weights, the protein-sugar ratios and acidities are variable. Neither a gastropod nor a bivalve pattern is recognized. The diversity of the organic matrices probably plays a main role in the fossilization processes of mollusc shells.     

A Monte Carlo algorithm for computing the IBD matrices using incomplete marker information

Identity-By-Descent (IBD) is a general measurement of the relationship between two groups of genes. If the two groups consist of two homologous genes, one from each individual, the IBD is called the coancestry between the two individuals. Coancestry is an important concept in both population and quantitative genetics. It is the probability that both genes are copies of the same gene in the genealogy. The average coancestry value at a random locus in a population reflects the level of population diversity, effective population size, the level of inbreeding and other attributes. Coancestry is also the building block for the covariance structure used to estimate the additive genetic variance component for a quantitative trait. There are many other types of IBD matrices, depending on the natures of the genes included in each group, and these IBD matrices vary from locus to locus. Molecular markers distributed along the genome provide information that can be used to infer these locus-specific IBD matrices. As a result, we can estimate and test the variance components of a quantitative trait contributed by these loci using the inferred IBD matrices. In this study, we develop the concept of locus-specific epistatic IBD matrices and a Monte Carlo method to infer these IBD matrices. The method is suitable for large pedigrees with arbitrary complexity and various levels of missing marker information. With these locus-specific IBD matrices, we are ready to search for quantitative trait loci along the genome in complicated pedigrees.     

The Method of Random Skewers

Ecological and evolutionary studies are often concerned with the properties of covariance matrices. The method of random skewers (RS method) has been used compare a matrix to an a priori vector or to compare two matrices. The method involves multiplying a matrix by many random vectors drawn from a uniform distribution over all possible vector directions. The comparisons are usually made using the average angle (or cosine) of the response vectors to an a priori vector or to the response vectors corresponding from another matrix. Angles are usually constrained to the interval 0°–90° because the distribution of response vectors is bipolar bimodal. The size of the average angle or cosine depends strongly on the relative sizes of the eigenvalues (especially the first). The distribution of angles between pairs of response vectors from two covariance matrices is more complicated because it depends on the differences in orientation of the eigenvectors and the relative sizes of the eigenvalues of the both matrices. The average absolute value of the angles between these pairs of response vectors depends on the relative sizes of the eigenvalues of the matrices making it difficult to interpret its meaning without knowledge of the eigenvalues and eigenvectors of the two matrices. Thus, it is simpler to just directly compare matrices in terms of these quantities.     

EFFECTS OF IONOPHORE A23187 ON OOCYTES OF COMANTHUS JAPONICA (ECHINODERMATA: CRINOIDEA)*

Micromolar amounts of divalent cation ionophore A23187 stimulate full grown (but unfertilizable) oocytes of Comanthus japonica to undergo a cortical reaction that is incomplete: first, cortical granule contents ejected at exocytosis do not coalesce but remain as individual blebs just outside the oocyte; and, second, about a fourth of the cortical granule population does not undergo exo-cytosis and remains in the cortical cytoplasm. Of the cortical granules remaining in the oocyte, some have unreacted contents and others have morphologically modified contents. Fine structures are compared among unreacted cortical granules, internally-reacted cortical granules, extracellular blebs of cortical granule material and normal fertilization membranes. The comparison strongly suggests that the outer dense layer and inner fibrous layer of the normal fertilization membrane are derived, respectively, from the dense patches and from the matrices of the cortical granules.     

hnRNA and its attachment to a nuclear protein matrix

In this study, DNA-depleted nuclear protein matrices are isolated from HeLa S3 cells. These nuclear matrices consist of peripheral laminae, residual nucleoli, and internal fibrillar structures. High molecular weight, heterogeneous nuclear RNA (hnRNA) is quantitatively associated with these structures and can be released intact only by affecting the integrity of the matrices. It is, therefore, concluded that hnRNA is part of a highly organized nuclear structure. By irradiation of intact cells or isolated nuclear matrices with ultraviolet light, proteins tightly associated with hnRNA can be induced to cross-link with the RNA. Performing the cross-linking in vivo is an extra guarantee that only hnRNA-protein (hnRNP) complexes existing in the intact cell are covalently linked. Such hnRNP complexes were isolated and purified under conditions that completely dissociate nonspecific RNA-protein complexes. By comparison of the hnRNP found in nuclear matrices and the published data on the composition of hnRNP particles, it was found that the so-called hnRNP "packaging" proteins (32,000-38,000 mol wt) were not efficiently cross-linked to hnRNA by UV irradiation. They were, however, present in the matrix preparations, bound to hnRNA, because they were released from nuclear matrices after ribonuclease treatment of these structures. On the other hand, two major hnRNPs (41,500 and 43,000 mol wt) were efficiently cross-linked to hnRNA. These proteins were not released by ribonuclease treatment, which suggests that they are involved in the binding of hnRNA to the nuclear matrix.     

两种常见海水鱼高温贮存过程中挥发性盐基氮和生物胺含量变化

棘头梅童鱼(Collichthys lucidus)和龙头鱼(Harpodon nehereus)是我国沿海常见的两种小型海水鱼, 常被作为水产动物饵料, 也可被人类食用。研究检测了这两种鱼在30℃下贮存48h 每隔6h 的挥发性盐基氮(T-VBN)和9 种生物胺(尸胺、腐胺、组胺、酪胺、5-羟色胺、亚精胺、精胺、多巴胺、章鱼胺)的含量变化, 并对这两种鱼的T-VBN 和生物胺含量与时间的相关性进行分析, 为水产品类饵料安全投喂和人类食品安全提供基础资料。结果表明: 两种鱼在相同贮存条件中T-VBN 和生物胺含量均存在一定差异。T-VBN 含量随着贮存时间的延长而逐渐增加, 棘头梅童鱼T-VBN 含量从0h 的8.19 mg/100 g 增加到48h 的568.05 mg/100 g,龙头鱼从0h 的13.16 mg/100 g 增加到48h 的361.34 mg/100 g, 棘头梅童鱼增长值显著高于龙头鱼(PPPP>0.05);多巴胺在两种鱼体内均未检测到。这两种鱼体内T-VBN、腐胺、尸胺、组胺、酪胺含量与时间的相关性均极其显著(P<0.01)。       

Phylogenetic analysis of correlation structure in stalk-eyed flies (Diasemopsis,Diopsidae)

Morphological divergence among species may be constrained by the pattern of genetic variances and covariances among traits within species. Assessing the existence of such a relationship in nature requires information on the stability of intraspecific correlation and covariance structure and the correspondence of this structure to the pattern of evolutionary divergence within a lineage. Here, we investigate these issues for nine morphological traits and 15 species of stalk-eyed flies in the genus Diasemopsis. Within-species matrices for these traits were generated from phenotypic data for all the Diasemopsis species and from genetic data for a single Diasemopsis species, D. dubia. The among-species pattern of divergence was assessed by calculating the evolutionary correlations for all pairwise combinations of the morphological traits along the phylogeny of these species. Comparisons of intraspecific matrices reveal significant similarity among all species in the phenotypic correlations matrices but not the covariance matrices. In addition, the differences in correlation structure that do exist among species are not related to their phylogenetic placement or change in the means of the traits. Comparisons of the phenotypic and phylogenetic matrices suggest a strong relationship between the pattern of evolutionary change among species and both the intraspecific correlation structure and the stability of this structure among species. The phenotypic and the phylogenetic matrices are significantly similar, and pairs of traits whose intraspecific correlations are more stable across taxa exhibit stronger coevolution on the phylogeny. These results suggest either the existence of strong constraints on the pattern of evolutionary change or a consistent pattern of correlated selection shaping both the phenotypic and phylogenetic matrices. The genetic correlation structure for D. dubia, however, does not correspond with patterns found in the phenotypic and phylogenetic data. Possible reasons for this disagreement are discussed.     

Discrimination of phytochrome dependent light inducible from non-light inducible plant genes. Prediction of a common light-responsive element (LRE) in phytochrome dependent light inducible plant genes.

We aligned 14 5'-leading sequences of small subunit ribulose-1,5-bisphosphate carboxylase (rbcS) genes. A strong consensus sequence ("CCTTATCAT") was located directly upstream of the TATA-box. The occurrence of this motif in other light dependent phytochrome regulated plant genes led to the calculation of two consensus matrices. With these two matrices we are able to distinguish almost all known light induced plant genes which are phytochrome regulated from non-light induced plant genes indicating, that all these genes share a common light-responsive element (LRE). The results obtained by computer analysis are discussed with regard to experimental data.     

Poly(ethyleneoxide) for high resolution and high-speed separation of DNA by capillary electrophoresis

Capillary electrophoresis (CE) with polyacrylamide gels has already been demonstrated to allow single-base resolution of single-stranded DNA. However, linear polyacrylamide is not an ideal matrix because of a high viscosity and difficulties in preparing the polymer with well defined pore sizes. Alternatively, poly(ethyleneoxide) (PEO) with a large range of molecular masses from 300 000 to 8 000 000 is available commercially. In addition, it is easy to prepare homogeneous solutions to provide highly reproducible separation performance with sufficient resolution. Single-base resolution of double-stranded DNA between 123 and 124 base pairs can be achieved by the use of homogeneous matrices prepared from PEO (2.5% M_r 8 000 000), and even better resolution is achieved by using mixed polymer matrices. With further work, it should be possible to change the fractions and the total amounts of polymers to achieve even higher resolution for different samples with different size ranges of fragments. Another advantage of mixed polymer matrices is that relatively high resolution can be obtained while maintaining a relatively low viscosity compared to linear polyacrylamide with identical contents of formamide and urea, which makes it easier to fill these matrices into small capillaries.     

Changes in the ratio of microsomal electron carriers in reconstituted microsomal membranes

The reconstitution of microsomal membrane monooxygenase system with variable contents of the hydroxylating chain enzymatic components was carried out. It was found that during self-assembly of microsomal membranes solubilized with 4% sodium cholate and gel filtration through Sephadex LH-20 in the presence of isolated microsomal enzymes, two forms of cytochrome P-450, i. e. phenobarbital- and 3-methylcholantrene-induced ones, and NADPH-cytochrome P-450 reductase, the exogenous enzymes are incorporated into the microsomal membrane matrices of control and methyl-cholantrene-treated animals. In the membranes reconstituted from the microsomes of the methylcholantrene-induced animals the catalytic activity of cytochrome P-448 in the metabolism of benz(a)pyrene at varying cytochrome P-448 and NADPH-cytochrome P-450 reductase contents were investigated.     

Correlation of the cell phenotype of cultured cell lines with their adhesion to components of the extracellular matrix

The differential adhesion of cultured mammalian clonal cell lines to components of the extracellular matrix was examined by kinetic adhesion and long-term growth assays. Uniform artificial matrices were prepared by air drying collagen Type I solution (C) onto a microtiter well and then air drying a solution containing a single glycosaminoglycan (GAG): hyaluronic acid (HA), chondroitin sulfate-4 (CHS-4), or chondroitin sulfate-6 (CHS-6). The adhesion of [3H]thymidine-prelabeled cells suspended in fibronectin (FN) depleted medium was measured at 2 and 6 hr. Neuroblastoma (N18, Lan 1) and melanoma (B16, G361, S91) cell lines exhibited a significantly greater percentage of cells adhering to one or more C-GAG matrices compared with C matrices. Maximal adhesion at 2 hr was to C-HA. In contrast at 2 hr, two glial, two epithelial, and one fibroblastic cell line showed unchanged or significantly decreased binding to C-GAG compared with C matrices. Further experiments using a neuroblastoma (N18) and a glioma (C6) cell line indicated that the adhesion patterns were not altered either by the method of dissociation from the tissue culture dish, preincubation with exogenous GAG, or the addition of exogenous fibronectin. Assays of N18 and C6 adhesion to matrices made from a non-GAG polyanionic compound, polygalacturonic acid (PGA), did not yield the same adhesion patterns as C-HA matrices. Long-term growth studies of a neuroblastoma (N18) melanoma (S91), and glioma (C6) cell line on nonuniform matrices deliberately prepared with GAG-rich and GAG-poor regions complemented the observations from the kinetic adhesion assays. N18 and S91 cells did not grow on areas which did not contain GAG by toluidine blue staining. However, the C6 cells did not grow on areas which did strongly stain for GAG. A quantitative analysis of the long term growth of N18 and C6 cells substantiated these observations. All these data indicate that the cellular phenotype may be correlated with matrix adhesion. Neuroblastomas and melanomas have a greater affinity for GAG-containing matrices while glial, epithelial, and fibroblastic cells appear to have a greater or equal affinity for collagen matrices.     

Comparison of the soluble matrices of the calcitic prismatic layer of Pinna nobilis (Mollusca,Bivalvia, Pteriomorpha)

The calcitic prisms of the outer layer of the shell of Pinna nobilis, surrounded by thick organic walls, contain a soluble intracrystalline matrix. The structure and composition of the outer interprismatic walls are not well known. The current viewpoint is they are composed of an insoluble matrix. Another thick organic structure, the interlamellar sheet of the nacreous layer, is composed of insoluble and soluble matrices. The composition of two sets of soluble organic matrices from the calcitic layer of Pinna nobilis, extracted with and without the organic walls are compared. According to the various analyses (SEM and AFM, UV and FTIR spectrometry, HPLC, electrophoreses, XANES), the main characteristics of the two matrices are similar, but not identical. Thus, the organic walls contain soluble components. However, the three-layered structure of the interlamellar sheet of the nacreous layer has not been observed.