Characterization of a V κ family in Mus musculus castaneus: sequence analysis

To examine genetic variation at immunoglobulin (Ig) multigene loci over short spans of evolutionary time, we have compared members of an Ig kappa chain variable (V ) region family from several mouse species. In this study, seven unique Igk-V24 family members have been isolated from Mus m. castaneus and characterized by nucleotide sequence determination for comparison to their counterparts in Mus m domesticus (BALB/c), and Mus pahari, representing 1–2 million years of evolution in the former case and 5–8 million years in the latter. Parsimony, together with evolutionary distances calculated for various paris of Igk-V24 family coding regions, relate all family members to a common progenitor existing roughly 24 million years ago (Mya). A significant portion of the M. m. castaneus family consists of pseudogene segments in various degrees of progressive degeneration. The substitution patterns and divergence rates for all gene segments are characteristic of their respective subsets, especially in the areas flanking the coding regions. Complex and variable patterns of diversity are seen in potentially expressed coding regions, which appear to reflect quite different selective pressures on various subregions within the V protein domain. These results indicate that evolutionary pressures are operating at the level of family subsets, their individual members, and subregions within similar molecules.The nucleotide sequence data presented in this report have been submitted to the GenBank nucleotide sequence database and assigned the accession numbers (Igk-CaV24) M80407; (Igk-CaV24A) M80408; (Igk-CaV24B.1) M80409; (Igk-CaV24B.2) M80410; (Igk-CaV24D.1) M80411; (Igk-CaV24D.2) M80412; and (Igk-CaV24D.3) M20892.     

Experiments with β-chain variants of the hemoglobin of Mus musculus

Starch gel electrophoretic and ultracentrifuge methods failed to demonstrate any differences between the hemoglobins of mice of the Shanghai and HBBP/Cag strains and crosses among these strains. The apparent identity of these hemoglobins is thought to stem from the contribution of Asian mice to the British mouse fancy from which the laboratory strains having Hbb-p in part descerd. Maleate buffer of pH 7 or above can be used to prevent the formation of disulfide-bridged dimers of mouse hemoglobins. However, the minor electrophoretic bands of Hbb-p and Hbb-d react with approximately twice as much maleate as the major bands of each of these hemoglobins, although the minor bands like the major contain only one free cysteine group per chain. This can be explained by the alkylation of the -amino of lysine residue 76, but some evidence for the alkylation of histidine in the minor band of Hbb-p is also presented.     

“Pigtail,” a hereditary tail abnormality in the house mouse,Mus musculus

Journal of Genetics - A skeletal abnormality, termed pigtail, in the house mouse is described which phenotypically resembles flexed tail. In a certain small proportion of litters, one or more young...     

Characterization of enzyme-catalysed endogenous β-hydroxylation of phenylethanoid glycosides in Euphrasia rostkoviana Hayne at the molecular level

Phenylethanoid glycosides, the main constituents of the aerial part of eyebright (Euphrasia rostkoviana Hayne) were treated by the endogenous hydroxylase enzyme and the concomitant biotransformation was characterized by applying high-performance liquid chromatography with UV and MS detections and NMR spectroscopy. In the extracts of the untreated (intact) samples, acteoside and eukovoside were determined as main compounds. The enzymatic treatment resulted in the quantitative transformation of these phenylethanoid glycosides into their corresponding hydroxyl derivatives identified as two epimers of β-hydroxyacteoside and β-hydroxyeukovoside. As to the importance of this hydroxylation β-hydroxyeukovoside was identified as a new compound and β-hydroxyacteoside was described for the first time in eyebright. We proved for first time that a β-hydroxylase enzyme is active in eyebright tissues which can transform phenylethanoid glycosides into their β-hydroxyl derivatives. Our new enzymatic method combined with a preparative HPLC facilitates the isolation of β-hydroxyl phenylethanoid glycosides from the aerial part of eyebright.     

Do precocial mammals develop at a faster rate? A comparison of rates of skull development in Sigmodon fulviventer and Mus musculus domesticus

Variation in neonatal maturity among mammals is often explained by variation in gestation length, but species may also differ in developmental rate, a quantity that is difficult to measure because the conventional formalism makes two important and potentially unrealistic assumptions: (1) ontogeny of form can be described by a single line, and (2) species have the same ontogeny of form. We examine two species, one precocial (Sigmodon fulviventer), the other altricial (Mus musculus domesticus), and find that neither assumption is met. Therefore, we introduce an alternative metric, the rate of shape differentiation away from the average neonate. We find that S. fulviventer has a lower developmental rate than M. m. domesticus; consequently, while more mature at birth, S. fulviventer loses ground to M. m. domesticus over time. Surprisingly, despite differences in gestation length and developmental rate, these species reach developmental and life-history milestones at nearly identical degrees of skull shape maturity.     

Determination of adult size in a folivorous moth: constraints at instar level?

1. Reaction norms for size and age at maturity were studied in Epirrita autumnata (Lepidoptera, Geometridae). Growth rates were manipulated by rearing larvae on different levels of food quality and quantity, and instar-specific final weights and development times were recorded.
2. Food level and initial weight of an instar accounted for most of the variance in final weights. Sexual dimorphism in pupal weights could be entirely ascribed to sex differences in initial weights of the last instar.
3. There were problems with considering the reaction norms optimal within the conventional demographic explanatory framework. Because fecundity increases linearly with body size and no costs of large adult size are known, one should expect female larvae to grow larger to increase their individual fitness. It is therefore likely that constraints play a major role in the determination, and evolution, of size in this species.
4. A focus on individual instars may be the best way to reveal the constraints on, and space for, adaptive evolution of insect growth. Some limit to the initial:final weight ratio of an instar, and the fixed number of instars, may represent important constraints.
5. Reaction norms like the ones described in this study lead to strong environmental determination rather than canalization of body size. Food quality throughout larval development may thus be very important for individual fitness and population dynamics in E. autumnata.     

Characterization of a Brassica napus myrosinase pseudogene: myrosinases are members of the BGA family of β-glycosidases

Myrosinase isoenzymes are known to be encoded by two different families of genes denoted MA and MB. Nucleotide sequence analysis of a Brassica napus genomic clone containing a gene for myrosinase revealed it to be a pseudogene of the MA family. The gene spans more than 5 kb and contains at least 12 exons. The exon sequence of the gene is highly similar to myrosinase cDNA sequences. However, the gene displays three potential or actual pseudogene characters. Southern blot analysis using probes from the 3 portions of the genomic and B. napus MA and MB cDNA clones showed that MA type myrosinases are encoded by approximately 4 genes, while MB type myrosinases are encoded by more than 10 genes in B. napus. Northern blots with mRNA from seeds and young leaves probed with the MA-and MB-specific probes showed that the MA and MB myrosinase gene families are differentially expressed. Myrosinases are highly similar to proteins of a -glycosidase enzyme family comprising both -glycosidases and phospho--glycosidases of as diverged species as archaebacteria, bacteria, mammals and plants. By homology to these -glycosidases, putative active site residues in myrosinase are discussed on the basis of the similarity between -glycosidases and cellulases.     

Evolution of a Vκ gene family

To examine the evolution of multigene families we have selected as an example an immunoglobulin light chain variable region subgroup (V24) which has been extensively characterized in inbred mice (Mus musculus domesticus). Homologous genes have been isolated and sequenced from Mus pahari, a genetically and geographically isolated species believed to be the oldest living representative of the genus. Southern blot analysis using probes corresponding to individual genes in this subgroup reveals changes in the overall size of the family occurring at the level of individual genes but not at the level of the entire family. Nucleotide sequence analysis indicates an absence of regulatory sequences such as the CAT and TATA boxes 5 to the coding region, but a decanucleotide sequence involved in light chain expression is highly conserved. Within coding regions highly complex patterns of variation are seen which appear to reflect quite different selective pressures on various subregions of the coding sequence. Complementarity determining regions (CDR) are conserved to different extents, with the first CDR region in all family members being among the most conserved segments of the molecule. Conservation is similarly variable among framework segments, indicating complex and variable evolutionary pressures not only at the level of individual genes or their products but also at subregions within homologous molecules.     

Nomadic behaviour and colony fission in a cooperative spider: life history evolution at the level of the colony?

The concept of colony-level life history evolution is introduced for the cooperative spiders by describing the life cycle and demography of Atbuhna binotata (Araneae: Dictynidae), a species living in groups containing up to several dozen adult females plus their offspring. In a life cycle remarkably similar to that of army ants, the colonies of A binotata were found to reproduce by fission and to alternate nomadic and sedentary phases in tight association with their internal demography. Colonies of other cooperative spiders, on the other hand, remain stationary as they grow for a number of generations before producing propagules that are relatively small subsets of the maternal colony. It is suggested that A. binotata!% peculiar life cycle may have unfolded as a consequence of the two-dimensional architecture of its nests. Expanding two-dimensional nests may fragment more easily than the three-dimensional nests characteristic of other species. A long distance group migration or nomadic phase, described here for the first time for a spider, may have followed as a mechanism to cope with potential disadvantages of fission while selecting for strict synchronization of individual life cycle stages within the nests. It is shown, however, that, as in other cooperative spiders, A. binotatd% sex ratio is also highly female biased. The theoretical implications of biased sex ratios in a species with fissioning colonies are briefly discussed.     

‘We are against a multi-ethnic society’: policies of exclusion at the urban level in Italy

Local policies for immigrants in recent years have attracted a growing interest among scholars. It is increasingly accepted that they are distinct units of analysis in the governance of migration, with significant degrees of autonomy with regard to national policies. Most of the literature, however, deals with the inclusive role of local policies. The argument of this article, on the contrary, is the development of local policies aimed to exclude migrants from various kinds of benefits and rights. It is based on a pilot research, conducted in Lombardy (northern Italy), on seventy cases, referred to forty-seven different local authorities. Then, the outcome of these policies is analysed: the exclusion of migrants is a tool to seek political consent, but is also a battlefield, where anti-discrimination institutions, advocacy groups and courts react against the measures approved by local authorities.     

Genetic study of pancreatic proteinase and α-amylase in mice (Mus musculus)

Genetic variants were found at two loci for pancreatic proteinase in mice. The Prt-1 locus contains a pair of allelic genes, Prt-1 ^a and Prt-1 ^b , ad the Prt-2 locus contains two codominant allelic genes, Prt-2 ^a and Prt-2 ^b .Expression of the two genetic variants of proteinase allowed mice strains used in this study to be classified into three phenotypic classes. Prt-1 ^b andPrt-2 ^a were found in most of the Japanese inbred strains, Prt-1 ^b andPrt-2 ^a were found in most of the inbred strains imported from the United States, and, furthermore, Prt-1 ^b and Prt-2 ^b were present in Japanese feral-origin mice strains. Prt-1, Prt-2, and Amy-2 loci did not belong to the same linkage group.     

Inhibiting astrocytic activation: a novel analgesic mechanism of ketamine at the spinal level?

Although ketamine is widely used as an analgesic agent and has an anti-allodynic effect on neuropathic pain, the underlying analgesic mechanisms are not fully explained by the modern 'neuronal-based' theories. As emerging studies have focused on the critical role of spinal astrocytes in the pathological pain states, we have hypothesized that there exist some 'astrocytes-related' mechanisms in the analgesic function of ketamine. In the present study, using the spinal nerve ligation (SNL) pain model, we investigated the anti-nociceptive effects of intraperitoneal or intrathecal ketamine on SNL-induced neuropathic pain response, meanwhile, we investigated the astrocytic activation after ketamine administration on SNL rats. Behavioral data showed that either intraperitoneal or intrathecal ketamine inhibited SNL-induced allodynia, however, immunohistochemistry showed that SNL induced astrocytic activation was suppressed by intrathecal but not intraperitoneal ketamine. Using quantitative Western blot analysis, our report showed that intrathecal ketamine down-regulated glial fibrillary acidic protein expression, suggesting inhibition of SNL-induced astrocytic activation, which wasn't influenced by intraperitoneal administration. We conclude that intraperitoneal ketamine could alleviate SNL-induced neuropathic pain via the classical 'neuronal-based' mechanisms, but in addition, 'astrocytes-related' mechanisms were also important underlying the anti-allodynic effect of intrathecal ketamine.     

Characterization of a novel β-L-Arabinofuranosidase in Bifidobacterium longum: functional elucidation of A DUF1680 family member

Pfam DUF1680 (PF07944) is an uncharacterized protein family conserved in many species of bacteria, actinomycetes, fungi, and plants. In a previous article, we cloned and characterized the hypBA2 gene as a β-l-arabinobiosidase in Bifidobacterium longum JCM 1217. In this study, we cloned a DUF1680 family member, the hypBA1 gene, which constitutes a gene cluster with hypBA2. HypBA1 is a novel β-l-arabinofuranosidase that liberates l-arabinose from the l-arabinofuranose (Araf)-β1,2-Araf disaccharide. HypBA1 also transglycosylates 1-alkanols with retention of the anomeric configuration. Mutagenesis and azide rescue experiments indicated that Glu-366 is a critical residue for catalytic activity. This report provides the first characterization of a DUF1680 family member, which defines a new family of glycoside hydrolases, the GH family 127.     

Characterization of the α-mannosidase gene family in filamentous fungi: N-glycan remodelling for the development of eukaryotic expression systems

Although filamentous fungi are used extensively for protein expression, their use for the production of heterologous glycoproteins is constrained by the types of N-glycan structures produced by filamentous fungi as compared to those naturally found on the glycoproteins. Attempts are underway to engineer the N-glycan synthetic pathways in filamentous fungi in order to produce fungal expression strains which can produce heterologous glycoproteins carrying specific N-glycan structures. To fully realize this goal, a detailed understanding of the genetic components of this pathway in filamentous fungi is required. In this review, we discuss the characterization of the α-mannosidase gene family in filamentous fungi and its implications for the elucidation of the N-glycan synthetic pathway.     

Characterization of new members of the pregnancy-specific β1-glycoprotein family

Three cDNAs encoding members of the pregnancy-specific 1-glycoprotein (PSG) family were isolated from human term placental cDNA library. All three cDNAs encode proteins with similar domain structure. There is a leader sequence of 34 amino acids followed by an N-domain of 109 amino acids. Immediately after the N-domain are one or two copies of a repeating A-domain of 93 amino acids, a B-domain of 85 amino acids and a C-domain of variable size. The proteins are highly hydrophilic. However, one of them has an 81-amino acid C-domain which is very hydrophobic and could potentially serve as a membrane attachment site. The putative cell-cell recognition tripeptide, Arg-Gly-Asp, is present in the N-domain of two of the proteins. Partial sequence of one of the cDNAs has been found in HeLa cells while cDNAs highly homologous to two of the cDNAs have been found in the fetal liver. Functional roles of the PSG proteins basing on their structure are proposed.Abbreviations PSG Pregnancy-Specific 1-Glycoprotein, according to nomenclature recommended at the ISOBM XVII Meeting, 1989 [31] - CEA Carcinoembryonic Antigen - bp base-pair - kb kilo-base-pair - nt nucleotide - aa amino acid - UTR Untranslated Region - RGD Arg-Gly-Asp The nucleotide sequence of two of the cDNAs presented in this article have been submitted to GenBank under the accession numbers M37102 (hPS91) and M37103 (hPS133). The nucleotide sequence of hPS176 has also been submitted. No accession number is available yet.     

Characterization of a polymorphism in the 3′ part of the chicken vitellogenin gene

Summary An allele giving rise to a polymorphism within the 3 part of the chicken vitellogenin gene was cloned, sequenced, and compared to the previously cloned allele. The polymorphism is formed by a perfect copy of 343 bp from intron 32 in tandem array with a perfect copy of 244 bp from intron 33; this 587-bp element is inserted in a head-to-tail arrangement in intron 33. We propose a mechanism in which an unequal crossing-over resulted in a vitellogenin gene with two exons 33, one of which was subsequently deleted. Thus, intron 33 was enlarged by the tandem repeats without affecting the protein-encoding sequence of the gene. At the boundaries of the repeated elements, two short direct repeats are found that resemble the recombination signals of immunoglobulin genes. They may have had a key role in the formation of the new allele.     

Reconstruction of the astaxanthin biosynthesis pathway in rice endosperm reveals a metabolic bottleneck at the level of endogenous β-carotene hydroxylase activity

Astaxanthin is a high-value ketocarotenoid rarely found in plants. It is derived from β-carotene by the 3-hydroxylation and 4-ketolation of both ionone end groups, in reactions catalyzed by β-carotene hydroxylase and β-carotene ketolase, respectively. We investigated the feasibility of introducing an extended carotenoid biosynthesis pathway into rice endosperm to achieve the production of astaxanthin. This allowed us to identify potential metabolic bottlenecks that have thus far prevented the accumulation of this valuable compound in storage tissues such as cereal grains. Rice endosperm does not usually accumulate carotenoids because phytoene synthase, the enzyme responsible for the first committed step in the pathway, is not present in this tissue. We therefore expressed maize phytoene synthase 1 (ZmPSY1), Pantoea ananatis phytoene desaturase (PaCRTI) and a synthetic Chlamydomonas reinhardtii β-carotene ketolase (sCrBKT) in transgenic rice plants under the control of endosperm-specific promoters. The resulting grains predominantly accumulated the diketocarotenoids canthaxanthin, adonirubin and astaxanthin as well as low levels of monoketocarotenoids. The predominance of canthaxanthin and adonirubin indicated the presence of a hydroxylation bottleneck in the ketocarotenoid pathway. This final rate-limiting step must therefore be overcome to maximize the accumulation of astaxanthin, the end product of the pathway.