Structural organization of the beta-globin locus of B-haplotype sheep

Goats and some sheep synthesize a juvenile hemoglobin, Hb C (alpha 2 beta C2), at birth and produce this hemoglobin exclusively during severe anemia. Sheep that synthesize this juvenile hemoglobin are of the A haplotype. Other sheep, belonging to a separate group, the B haplotype, do not synthesize hemoglobin C and during anemia continue to produce their adult hemoglobin. To understand the basis for this difference we have determined the structural organization of the beta- globin locus of B-type sheep by constructing and isolating overlapping genomic clones. These clones have allowed us to establish the linkage map 5' epsilon I-epsilon II-psi beta I-beta B-epsilon III-epsilon IV- psi beta II-beta F3' in this haplotype. Thus, B sheep lack four genes, including the BC gene, and have only eight genes, compared with the 12 found in the goat globin locus. The goat beta-globin locus is as follows: 5' epsilon I-epsilon II-psi beta X-beta C-epsilon III-epsilon IV-psi beta Z-beta A-epsilon V-epsilon VI-psi beta Y-beta F3'. Southern blot analysis of A-type sheep reveals that these animals have a beta- globin locus similar to that of goat, i.e., 12 globin genes. Thus, the beta-globin locus of B-haplotype sheep resembles that of cows and may have retained the duplicated locus of the ancestor of cows and sheep. Alternatively, the B-sheep locus arrangement may be the result of a deletion of a four-gene set from the triplicated locus.      

Diaphragmatic activity is a determinant of postnatal lung growth

To test the hypothesis that activity of respiratory muscles determines regional growth of lung parenchyma, we studied the effects of unilateral diaphragmatic paralysis on contralateral/ipsilateral lung growth in cats and piglets. Five 10- to 12-wk-old cats and five 8-wk-old piglets underwent unilateral diaphragmatic paralysis by thoracic and cervical phrenectomy, respectively. Five to seven weeks after surgery, when the cats were killed for studies of lung growth, gain in body weight was the same as in five sham-operated controls. At this time, mean pleural pressure ipsilateral to the paralyzed hemidiaphragm was the same as contralateral mean pleural pressure during tidal breathing, and values did not differ from controls. However overall functional residual capacity was lower in the phrenectomized cats (35 +/- 4 ml) than in the controls (55 +/- 11 ml, P less than 0.01). Growth of contralateral lungs relative to ipsilateral lungs was greater in the phrenectomized cats than in the controls, as shown by ratios of contralateral/ipsilateral wet lung weight (1.44 vs. 1.34, P less than 0.01), maximum inflation volume (1.53 vs. 1.33, P less than 0.05), and total protein content (1.45 vs. 1.26, P less than 0.05). Ratios of total protein to DNA and RNA to DNA were unchanged. One week after surgery in the piglets, the ratio of contralateral/ipsilateral wet lung weight was increased (1.61 vs. 1.29, P less than 0.01) and total weight of both lungs was reduced. We conclude that regional growth of lung parenchyma by cell proliferation depends in part on regional distribution of respiratory muscle activity.     

Rotavirus RNA replication: VP2, but not VP6, is necessary for viral replicase activity.

Temperature-sensitive mutants of simian rotavirus SA11 were previously developed and organized into 10 of a possible 11 recombination groups on the basis of genome reassortment studies. Two of these mutants, tsF and tsG, map to genes encoding VP2 (segment 2) and VP6 (segment 6), respectively. To gain insight into the role of these proteins in genome replication, MA104 cells were infected with tsF or tsG and then maintained at permissive temperature (31 degrees C) until 9 h postinfection, when some cells were shifted to nonpermissive temperature (39 degrees C). Subviral particles (SVPs) were recovered from the infected cells at 10.5 and 12 h postinfection and assayed for associated replicase activity in a cell-free system shown previously to support rotavirus genome replication in vitro. The results showed that the level of replicase activity associated with tsF SVPs from cells shifted to nonpermissive temperature was ca. 20-fold less than that associated with tsF SVPs from cells maintained at permissive temperature. In contrast, the level of replicase activity associated with tsG SVPs from cells maintained at nonpermissive temperature was only slightly less (twofold or less) than that associated with tsG SVPs from cells maintained at permissive temperature. Analysis of the structure of replicase particles from tsG-infected cells shifted to nonpermissive temperature showed that they were similar in size and density to virion-derived core particles and contained the major core protein VP2 but lacked the major inner shell protein VP6. Taken together, these data indicate that VP2, but not VP6, is an essential component of enzymatically active replicase particles.     

Influence of broth dilution on the disruption of Escherichia coli

Summary The effect of cell concentration (5 to 150 g/L wet wt after broth dilution) on homogenizer disruption efficiency and homogenate viscosity is reported for E. coli. Broth dilution increases homogenizer efficiency and decreases feed and homogenate viscosity. However, this increase in disruption efficiency is not sufficient to warrant dilution of the broth prior to homogenization. The optimal feed concentration is the maximum possible that does not lead to practical handling difficulties due to high viscosity.     

Radioimmunoassay for the determination of limonin in Citrus

An ¹²⁵I-radioimmunoassay technique has been developed for the triterpenoid bitter principle, limonin. Synthesis of the iodinated tracer and the limonin—bovine serum albumin conjugate are described. The antibody has a high affinity (K_a 1.1 x 10⁹l/mol) and specificity for limonin and the detection limit of the assay is 0.07 ng or 0.7 ppb. Standard curves are linear over a range of 0.5–100 ng limonin, assays can be performed in crude extracts, and several hundred samples can be processed per day. The distribution of limonin in fruits and vegetative parts of Citrus paradisi has been determined, highest values (0.92%) being found in the seeds, lowest (0.0007%) in the juice vesicles of ripe fruits. The potential of this assay method in citrus research is discussed.     

Phylogeny and biogeography of southern African spoon-winged lacewings (Neuroptera: Nemopteridae: Nemopterinae)

Nemopteridae are a charismatic family of lacewings characterised by uniquely extended hind wings. They are an ancient widespread group in the drier regions of the world. The family comprises two subfamilies, Crocinae (thread-wings) and Nemopterinae (spoon- and ribbon-wings). The present distribution of the family has been largely influenced by the vicariant events of plate tectonics, resulting in relict populations in some parts of the world and extensive evolutionary radiations in others, particularly southern Africa where the vast majority of the species are endemic to the Western and Northern Cape Provinces of South Africa. This study aimed to establish the validity of the 11 currently recognised genera and infer their biogeographic history using molecular sequence data from four gene regions. The hypothesis that the Cape nemopterines co-evolved with certain taxa in the Cape Floristic Region was also tested. Phylogenetic analysis supports seven of the 11 currently recognised genera. The crown age of the Nemopterinae is estimated to be at ca. 145.6 Mya, indicating that the group has been present since the late Jurassic. Most of the genera appear to have diversified during the middle Eocene and into the middle Miocene (ca. 44–11 Mya) with recent rapid radiation of several of the genera occurring during the late Miocene (ca. 6–4.5 Mya). While these data support an initial radiation with the Rushioideae (Aizoaceae) it is recommended that further study including observations and gut content be carried out.     

Lysophosphatidic acid,human osteoblast formation,maturation and the role of 1α,25-Dihydroxyvitamin D3 (calcitriol)

The simplest signalling lipid Lysophosphatidic acid (LPA) elicits pleiotropic actions upon most mammalian cell types. Although LPA has an established role in many biological processes, particularly wound healing and cancer, the function of LPA for human osteoblast (hOB) biology is still unravelling. Early studies, identified in this review, gave a reliable indication that LPA, via binding to one of several transmembrane receptors, stimulated multiple intracellular signalling networks coupled to changes in cell growth, fibronectin binding, maturation and survival. The majority of studies exploring the actions of LPA on hOB responses have done so using the lipid in isolation. Our own research has focussed on the co-operation of LPA with the active vitamin D3 metabolite, 1α25,dihydroxycholecalciferol (calcitriol), in light of a serendipitous discovery that calcitriol, in a serum-free culture setting, was unable to promote hOB maturation. We subsequently learnt that the serum-borne factor co-operating with calcitriol to enhance hOB differentiation was LPA bound to the albumin fraction of whole serum. Recent studies from our laboratory have identified that LPA and calcitriol are a potent pairing for securing hOB formation from their stem cell progeny. Greater understanding of the ability of LPA to influence, for example, hOB growth, maturation and survival could be advantageous in developing novel strategies aimed at improving skeletal tissue repair and regeneration. Herein this review provides an insight into the diversity of studies exploring the actions of a small lipid on a major cell type key to bone tissue health and homeostasis. This article is part of a Special Issue entitled Advances in Lysophospholipid Research.     

Patterns of population genetic variation in sympatric chiltoniid amphipods within a calcrete aquifer reveal a dynamic subterranean environment

Calcrete aquifers from the Yilgarn region of arid central Western Australia contain an assemblage of obligate groundwater invertebrate species that are each endemic to single aquifers. Fine-scale phylogeographic and population genetic analyses of three sympatric and independently derived species of amphipod (Chiltoniidae) were carried out to determine whether there were common patterns of population genetic structure or evidence for past geographic isolation of populations within a single calcrete aquifer. Genetic diversity in amphipod mitochondrial DNA (cytochrome c oxidase subunit I gene) and allozymes were examined across a 3.5 km² region of the Sturt Meadows calcrete, which contains a grid of 115 bore holes (=wells). Stygobiont amphipods were found to have high levels of mitochondrial haplotype diversity coupled with low nucleotide diversity. Mitochondrial phylogeographic structuring was found between haplogroups for one of the chiltoniid species, which also showed population structuring for nuclear markers. Signatures of population expansion in two of the three species, match previous findings for diving beetles at the same site, indicating that the system is dynamic. We propose isolation of populations in refugia within the calcrete, followed by expansion events, as the most likely source of intraspecific genetic diversity, due to changes in water level influencing gene flow across the calcrete.