The amphotropic murine leukemia virus receptor gene encodes a 71-kilodalton protein that is induced by phosphate depletion.

The amphotropic murine leukemia virus (MuLV) can infect cells from a number of mammals, including humans, via its specific receptor. Basic knowledge of amphotropic MuLV receptor expression is likely to be useful in the development and improvement of gene therapy protocols based on amphotropic-pseudotyped vectors. To investigate the expression of the human receptor for the amphotropic MuLV (GLVR-2, newly termed Pit2), we determined its mRNA levels in several cell lines and found them to vary significantly. Induction of increased levels of mRNA after removal of phosphate from the media was observed in two osteosarcoma cell lines. The increase in GLVR-2 mRNA resulted in a concomitant rise in the levels of a 71-kDa protein specifically recognized by affinity-purified antibodies against GLVR-2. Using these antibodies, we were able to confirm the intracellular topology of the large hydrophilic domain between the proposed sixth and seventh transmembrane domains of the GLVR-2 protein. This assignment is in agreement with the fourth extracellular loop being outside the cell, consistent with the proposal that the fourth extracellular loop of GLVR-2 contains the envelope binding site.     

Thiosulfate Reduction, an Important Physiological Feature Shared by Members of the Order Thermotogales

Several members of the order Thermotogales in the domain Bacteria, viz., Thermotoga neapolitana, Thermotoga maritima, Thermosipho africanus, Fervidobacterium islandicum, and Thermotoga strain SEBR 2665, an isolate from an oil well, reduced thiosulfate to sulfide. This reductive process enhanced cellular yields and growth rates of all the members but was more significant with the two hyperthermophiles T. neapolitana and T. maritima. This is the first report of such an occurrence in this group of thermophilic and hyperthermophilic anaerobic bacteria. The results suggest that thiosulfate reduction is important in the geochemical cycling of sulfur in anaerobic thermal environments such as the slightly acidic and neutral-pH volcanic hot springs and oil reservoirs.     

Reevaluating the classification of Halobacteroides and Haloanaerobacter species based on sequence comparisons of the 16S ribosomal RNA gene

Abstract The 16S rRNA gene (rDNA) sequence analysis of four halophilic anaerobes: Halobacteroides halobius, H. lacunaris, Haloanaerobacter (Hb.) chitinovorans and H. acetoethylicus confirmed that they were all members of the family Haloanaerobiaceae. H. lacunaris and H. halobius were found to be more closely related to each other and were distantly related to Sporohalobacter lortetti and the members of the genera Haloanaerobium and Halothermothrix . These data are in agreement with their assignment to the genus Halobacteroides . Further analysis indicated that Hb. chitinovorans was closely affiliated to members of the genus Halobacteroides , and therefore we propose to transfer it to the genus Halobacteroides as H. chitinovorans comb. nov. This transfer would invalidate the genus Haloanaerobacter , as Hb. chitinovorans is the only member of this genus. The 16S rDNA sequence analysis of H. acetoethylicum indicated that it was very closely related to members of the genus Haloanaerobium , viz. Haloanaerobium (Ha.) praevalens, Ha. salsugo , and Ha. alcaliphilum , and hence we propose to transfer it to the genus Haloanaerobium as Ha. acetoethylicus comb. nov.     

Anaerobic digestion of sugar beet pulps

Summary Anaerobic digestion of sugar beet pulps was studied in a 70 l digestor with sequential feeding, after enzymatic hydrolysis by Trichoderma harzianum cellulases. During the 130 days feeding, 3.6 m³ of biogas were produced with an average content of 58% CH₄ from 270 l of hydrolysed pulps at 20 g VS/l. Average yield and production rate were respectively 0.67 m³/kg VS and 0.4 m³/ kg VS and 0.4 m³/m³ of digestor per day.     

An immunoblotting technique for the detection of bound and secreted bacterial Fc receptors

A rapid semiquantitative procedure that enables bacteria to be screened for surface or secreted receptors for the Fc region of human IgG is described. Surface Fc receptors were detected by direct transfer of bacterial colonies to nitrocellulose by electroblotting and then probing with ¹²⁵I-labeled human IgG in the presence of a two fold molar excess of unlabeled F(ab′)₂fragments. The blots were exposed to X-ray film and the intensity of the resulting autoradiograph was a measure of surface Fc receptors expression. This procedure reliably distinguished Staphylococcus aureus strains which expressed different levels of surface Fc receptors. When applied to the study of group A streptococci, a number of Fc receptor-positive strains were identified. Unlike the homogeneous Fc receptor expression on individual colonies of the staphylococcal strains, a wide variation in the level of Fc receptor expression was observed within a given streptococcal strain. Group A streptococcal substrains which expressed high and low levels of surface Fc receptors could be isolated from replica plates.Secreted Fc receptors were measured by a simple modification of the blotting procedure in which the nitrocellulose was placed on the opposite side of the agar from the bacterial colonies. Secreted Fc receptors was electroblotted through the agar onto nitrocellulose and probed as described above. This approach readily detected nanogram quantities of secreted type I Fc receptor (protein A) from the Staphylococcus aureus Cowan strain. None of the group A streptococcal strains tested were found to secrete detectable quantities of Fc receptors.     

Voltage-dependent proton fluxes in liposomes

Liposomes containing buffered KCl were prepared from bacterial lipids, were diluted into K+-free media and were treated with valinomycin to induce the formation of a diffusion potential (delta psi). Upon formation of such a potential, substantial proton influx was observed, as assayed by the quenching of 9-aminoacridine fluorescence. Complete reversal of fluorescence quenching occurred when the potential was collapsed by addition of KCl or when methylamine was added. Studies of proton influx as a function of the theoretical magnitude of the delta psi indicated that the phenomenon occurred only above a delta psi of about -60 mV. Establishment of a Na+ diffusion potential also resulted in proton influx. Treatment of K+-loaded liposomes with N,N'-dicyclohexylcarbodiimide did not reduce the delta psi-dependent proton influx. Moreover, proton influx could be demonstrated upon imposition of a diffusion potential in liposomes prepared from a synthetic lipid. The proton fluxes associated with generation of a diffusion potential in liposomes may complicate studies of reconstituted systems in which proton translocation should occur, and may affect the magnitude of the electrochemical proton gradient that is operant under some conditions.     

Similar growth pattern of mouse mammary epithelium cultivated in collagen matrix in vivo and in vitro

Mouse mammary ductal cells cultured in type I collagen gels give rise to three-dimensional multicellular outgrowths consisting of thin spikes which are often branched, and which may have pointed or blunt ends. The significance of these spikes to normal ductal morphogenesis has been unclear, since identical structures are not known to occur in vivo; conversely, it has not been possible to maintain in gel culture the highly structured end buds which are characteristic of ductal elongation in the animal. In order to evaluate whether the pattern of radiating spikes observed in collagen gel cultures results from chemical or physical peculiarities of the culture environment, a small volume of unpolymerized type I collagen solution was injected into mammary gland-free fat pads of young adult mice. After the bubble of collagen had polymerized, an implant of mammary ductal epithelium was introduced into the center of the gel. Histological examination of the implants after 3 to 6 days of growth revealed numerous small epithelial spikes, similar to those observed in gel culture, extending into the fibrous matrix. The early stages of regeneration of mammary implants placed in gland-free fat pads were then examined without the addition of exogenous collagen. In cases where the epithelium happened to contact a fibrous region of the fatty stroma, spikes were also seen to form in these natural collagenous substrates. Whether or not exogenous collagen was used, normal end buds were formed only when epithelial spikes contacted adipocytes. It was concluded that the three-dimensional pattern of radiating tubules in collagen gels in vitro is not merely an artifact of culture, but has a counterpart in vivo whereever regenerating mammary epithelium is surrounded by fibrous stroma. A model is presented in which the pattern of epithelial outgrowth is determined by the physical characteristics of the surrounding stroma; in collagen matrix a comparatively primitive and unspecialized type of morphogenesis occurs which may not require the participation of stromal cells. In contrast, epithelial-adipocyte interactions appear to be necessary for the formation of end buds and subsequent morphogenesis of fully structured mammary ducts.     

S-adenosylmethionine and morphogenesis inMucor racemosus

The intracellular concentration of S-adenosylmethionine (SAM) and the specific activity of S-adenosylmethionine synthetase (ATP:l-methionine S-adenosyltransferase, EC 2.5.1.6) were examined in wild-typeMucor racemosus, as well as a morphological mutant termedcoy, under conditions designed to prevent the morphogenesis of yeasts to hyphae. When the mutant was grown in a defined medium supplemented with methionine and induced to shift by exposure to air, there was an increase in intracellular SAM analogous to that previously reported with the wild type. However, when thecoy mutant was grown in the absence of methionine, the intracellular concentration decreased dramatically and the mutant failed to undergo the yeast to hypha transition. An inhibitor of SAM synthetase activity, cycloserine, was used to lower the intracellular concentration of SAM in the wild-type organisms. Under these conditions, wild-typeM. racemosus failed to undergo the transition from yeasts to hyphae when exposed to air.