The murine 3β-hydroxysteroid dehydrogenase multigene family: Structure, function and tissue-specific expression

The classical form of the enzyme 5-ene-3β-hydroxysteroid dehydrogenase/isomerase (3βHSD), expressed in adrenal glands and gonads, catalyzes the conversion of 5-ene-3β-hydroxysteroids to 4-ene-3-ketosteroids, an essential step in the biosynthesis of all active steroid hormones. To date, four distinct mouse 3βHSD cDNAs have been isolated and characterized. These cDNAs are expressed in a tissue-specific manner and encode proteins of two functional classes. Mouse 3βHSD I and III function as 3β-hydroxysteroid dehydrogenases and 5-en→4-en isomerases using NAD⁺ as a cofactor. The enzymatic function of 3βHSD II has not been completely characterized. Mouse 3βHSD IV functions only as a 3-ketosteroid reductase using NADPH as a cofactor. The predicted amino acid sequences of the four isoforms exhibit a high degree of identity. Forms II and III are 85 and 83% homologous to form I. Form IV is most distant from the other three with 77 and 73% sequence identity to I and III, respectively. 3βHSD I is expressed in the gonads and adrenal glands of the adult mouse. 3βHSD II and III are expressed in the kidney and liver with the expression of form II greater in kidney and form III greater in liver. Form IV is expressed exclusively in the kidney. Although the amino acid composition of forms I, III and IV predicts proteins of the same molecular weight, the proteins have different mobilities on SDS-polyacrylamide gel electrophoresis. This characteristic allows for differential identification of the expressed proteins. The four structural genes encoding the different isoforms are closely linked within a segment of mouse chromosome 3 that is conserved on human chromosome 1.     

Large-scale physical mapping within the region 22q12.3-13.1 in meningioma

The lack of physical mapping data strongly restricts the analysis of the meningioma chromosomal region that was assigned to the bands 22q12.3-qter. Recently, we reported a new marker D22S16 for chromosome 22 that was assigned to the region 22q13-qter by in situ hybridization. Utilizing somatic cell hybrids we now sublocalized the marker D22S16 within the band region 22q12-13.1, thus placing it in the vicinity of the gene for the platelet derived growth factor (PDGFB). A physical map was established for the regions surrounding the PDGFB gene and the D22S16 marker. By means of pulsed-field gel electrophoresis (PFGE) D22S16 and PDGFB were found to be physically linked within 900 kb. We also identified two CpG clusters bordering the PDGFB gene. For the enzyme NotI, a variation of the PDGFB restriction pattern was found between different individuals. PFGE analysis of the two loci (PDFGB and D22S16) failed to identify major rearrangements in meningioma.     

Acquired thermotolerance and heat shock in the extremely thermophilic archaebacterium Sulfolobus sp. strain B12.

The extreme thermophile Sulfolobus sp. strain B12 exhibits an acquired thermotolerance response. Thus, survival of cells from a 70 degrees C culture at the lethal temperature of 92 degrees C was enhanced by as much as 6 orders of magnitude over a 2-h period if the culture was preheated to 88 degrees C for 60 min or longer before being exposed to the lethal temperature. In eubacteria and eucaryotes, acquired thermotolerance correlates with the induced synthesis of a dozen or so proteins known as heat shock proteins. In this Sulfolobus species, it correlates with the preferential synthesis of primarily one major protein (55 kilodaltons) and, to a much lesser extent, two minor proteins (28 and 35 kilodaltons). Since the synthesis of all other proteins was radically reduced and these proteins were apparently not degraded or exported, their relative abundance within the cell increased during the time the cells were becoming thermotolerant. They could not yet be related to known heat shock proteins. In immunoassays, they were not cross-reactive with antibodies against heat shock proteins from Escherichia coli (DnaK and GroE), which are highly conserved between eubacteria and eucaryotes. However, it appears that if acquired thermotolerance depends on the synthesis of protective proteins, then in this extremely thermophilic archaebacterium it depends primarily on one protein.     

Alpha-globin gene markers identify genetic differences between Australian aborigines and Melanesians.

Australian aborigines exhibit a number of alpha-globin cluster rearrangements involving both alpha- and zeta-globin genes. alpha+-Thalassemia (-alpha/) in this population is heterogeneous and includes the 3.7 types I, II, and III gene deletions. The alpha alpha alpha/ and zeta zeta zeta/ rearrangements are each found in association with two haplotypes, indicating origins from at least two separate DNA crossover events. Differences in alpha-globin cluster rearrangements and in haplotypes between Australian aborigines, Papua New Guinea highlanders and island Melanesians, are consistent with multiple colonizing events into Australia.     

Conserved and divergent aspects of Robo receptor signaling and regulation between Drosophila Robo1 and C. elegans SAX-3

The evolutionarily conserved Roundabout (Robo) family of axon guidance receptors control midline crossing of axons in response to the midline repellant ligand Slit in bilaterian animals including insects, nematodes, and vertebrates. Despite this strong evolutionary conservation, it is unclear whether the signaling mechanism(s) downstream of Robo receptors are similarly conserved. To directly compare midline repulsive signaling in Robo family members from different species, here we use a transgenic approach to express the Robo family receptor SAX-3 from the nematode Caenorhabditis elegans in neurons of the fruit fly, Drosophila melanogaster. We examine SAX-3’s ability to repel Drosophila axons from the Slit-expressing midline in gain of function assays, and test SAX-3’s ability to substitute for Drosophila Robo1 during fly embryonic development in genetic rescue experiments. We show that C. elegans SAX-3 is properly translated and localized to neuronal axons when expressed in the Drosophila embryonic CNS, and that SAX-3 can signal midline repulsion in Drosophila embryonic neurons, although not as efficiently as Drosophila Robo1. Using a series of Robo1/SAX-3 chimeras, we show that the SAX-3 cytoplasmic domain can signal midline repulsion to the same extent as Robo1 when combined with the Robo1 ectodomain. We show that SAX-3 is not subject to endosomal sorting by the negative regulator Commissureless (Comm) in Drosophila neurons in vivo, and that peri-membrane and ectodomain sequences are both required for Comm sorting of Drosophila Robo1.     

Recording fox baiting effort across the landscape using geographic information systems: Facilitating more effective management

Predation on native vertebrates and livestock by the European Red Fox (Vulpes vulpes) remains a significant problem in many parts of Australia. Coordinated approaches to fox control are most effective in protecting these assets and involve placement of baits across the landscape by private and public land managers. However, participation in such programmes varies seasonally and the spatial coverage of baiting is often difficult to determine. Here, we describe a geographic information systems‐based system that assists land managers to collect and use spatial information, minimizing gaps in bait coverage and maximizing bait encounters by foxes to increase the effectiveness of pest control. The coordination of data collection and reporting between land managers should facilitate more effective adaptive management by allowing better strategic planning and increasing landholder involvement, which should, in turn, improve the programme's efficacy, provided other critical conditions and resources are met.     

Population Genetics of the Eastern Hellbender (Cryptobranchus alleganiensis alleganiensis) across Multiple Spatial Scales

Conservation genetics is a powerful tool to assess the population structure of species and provides a framework for informing management of freshwater ecosystems. As lotic habitats become fragmented, the need to assess gene flow for species of conservation management becomes a priority. The eastern hellbender (Cryptobranchus alleganiensis alleganiensis) is a large, fully aquatic paedamorphic salamander. Many populations are experiencing declines throughout their geographic range, yet the genetic ramifications of these declines are currently unknown. To this end, we examined levels of genetic variation and genetic structure at both range-wide and drainage (hierarchical) scales. We collected 1,203 individuals from 77 rivers throughout nine states from June 2007 to August 2011. Levels of genetic diversity were relatively high among all sampling locations. We detected significant genetic structure across populations (F_st values ranged from 0.001 between rivers within a single watershed to 0.218 between states). We identified two genetically differentiated groups at the range-wide scale: 1) the Ohio River drainage and 2) the Tennessee River drainage. An analysis of molecular variance (AMOVA) based on landscape-scale sampling of basins within the Tennessee River drainage revealed the majority of genetic variation (∼94–98%) occurs within rivers. Eastern hellbenders show a strong pattern of isolation by stream distance (IBSD) at the drainage level. Understanding levels of genetic variation and differentiation at multiple spatial and biological scales will enable natural resource managers to make more informed decisions and plan effective conservation strategies for cryptic, lotic species.