The proα2 (V) collagen gene (COL5A2) maps to 2q14→2q32, syntenic to the proα1 (III) collagen locus (COL3A1)

Summary A recombinant probe specific for the pro2 chain of human Type V collagen has been used for the localization of the corresponding gene (COL5A2) to chromosome 2. Regional mapping by in situ hybridization and analysis of DNA from humanxrodent cell lines indicated that COL5A2 is confined within the segment 2q142q32, thus syntenic to the pro1 (III) collagen gene (COL3A1).     

Phage lambda and plasmid expression vectors with multiple cloning sites and lacZ alpha-complementation

Two new lambda vectors were constructed which permit cloning of genes that are potentially lethal if cloned in analogous plasmid vectors. lambda DL10 and lambda DL11 contain the alpha-complementing fragment of lacZ and multiple cloning sites found in the polylinker region of M13mp10 and M13mp11, respectively. DNA cloned into the unique cloning sites of these vectors can be detected by inactivation of alpha-complementation. These lambda vectors provide a lac promoter for expression of foreign genes as well as the ability to make fusion proteins. Two plasmid expression vectors, pPR110 and pPR111, were constructed from lambda DL10 and lambda DL11 respectively, and pCQV2. These plasmids, which express lacZ alpha-complementing activity from the lambda PR promoter, contain multiple cloning sites immediately downstream of the PR promoter. They allow cloning of genes under the control of the PR promoter and the plasmid-encoded thermosensitive (cI857) repressor, and allow easy detection of inserted fragments by inactivation of alpha-complementation.     

Photoaffinity Labeling of Benzodiazepine Receptors in Rat Brain with Flunitrazepam Alters the Affinity of Benzodiazepine Receptor Agonist But Not Antagonist Binding

Abstract: Recently, it was proposed that β-carbolines interact with a subset of benzodiazepine (BZD) binding sites in mouse brain. This postulate was based upon evidence showing changes in binding properties of the BZD receptor following photoaffinity labeling of membranes with flunitrazepam (FLU). Under conditions in which 80% of specific [³H]diazepam binding was lost in photolabeled membranes, specific [³H]propyl β-carboline-3-carboxylate ([³H]PCC) binding was spared. In this study, the binding of the BZD antagonists [³H]PCC, [³H]Ro15 1788 and [³H]CGS 8216 was examined in rat brain membranes following photoaffinity labeling with FLU. No significant changes in the apparent K_D and small reductions in the B_max of ³H antagonist binding were observed. However, in the same membranes, up to 89% of specific [³H]FLU binding was lost. When [³H]PCC (0.05 nM) was used to label the receptors in control and photolabeled membranes, the ability of BZD receptor agonists to inhibit [³H]PCC binding was greatly diminished in the photolabeled membranes. In contrast, the potency of BZD antagonists remained the same in both control and treated membranes. Based upon PCC/[³H]Ro15 1788 competition experiments, the ability of PCC to discriminate between BZD receptor subtypes was unaffected by photoaffinity labeling of cortical membranes. Overall, these findings suggest that β-carbolines do not interact with a subset of BZD binding sites per se, but may be a consequence of the differential interaction of BZD agonists and antagonists with BZD binding sites that have been photoaffinity labeled with FLU. A possible mechanism underlying this phenomenon is discussed. The ability of photolabeled membranes to differentiate between BZD agonists and antagonists provides a potential screen for agonist and antagonist activity in compounds that interact with the BZD receptor.     

Susceptibility of inbred mice to Leishmania tropica infection: correlation of susceptibility with in vitro defective macrophage microbicidal activities

Eleven mouse strains were inoculated in footpads with amastigotes of Leishmania tropica and observed for 12 weeks. Liver and spleen impression smears from infected mice were examined for the presence of intracellular parasites. Four strains (BALB/cJ, C57L/J, NZW/N, and P/J) failed to heal the subcutaneous lesion and showed evidence of systemic infection; the remaining seven strains (A/J, C3H/HeJ, C3H/HeN, C3HeB/FeJ, C57BL/6J, C57BL/10J, and C57BL/10ScN) were each resistant to infection and resolved their lesions by Week 10. Macrophages from the four susceptible strains could not be activated to kill L. tropica amastigotes by treatment with soluble lymphocyte products in vitro. In contrast, macrophages from all seven resistant strains responded to lymphokine treatment and eliminated 80-90% of intracellular parasites. These results suggest that in vitro macrophage microbicidal activities predict the course of systemic leishmanial disease.     

Association of newly synthesized major fl coat protein with infected host cell inner membrane

The bacteriophage fl major coat protein becomes associated with the host cell inner membrane very shortly after it is synthesized. Pulse-chase experiments suggest that the virus is never stably associated with the host cell outer membrane; we propose that it passes directly from the inner membrane to the growth medium.     

Properties of a prolactin receptor from the rabbit mammary gland

Receptors for human, simian, ovine, bovine and murine prolactin, human growth hormone and human placental lactogen have been identified in plasma-membrane-containing subcellular particles isolated from rabbit mammary glands. The association and dissociation of (125)I-labelled prolactin are time- and temperature-dependent processes, both being maximal at 37 degrees C. (125)I-labelled prolactin prepared by the enzymic iodination procedure with lactoperoxidase binds better to receptors than does the preparation obtained by using chloramine-t as the oxidizing agent. The binding of (125)I-labelled prolactin to receptors is strongly influenced by pH and ionic composition but not by many low-molecular-weight compounds tested, e.g. steroids, nucleotides and several drugs. Receptor activity is sensitive to trypsin and phospholipase C digestion, suggesting that protein and phospholipid moieties are essential for the binding of (125)I-labelled prolactin. The binding of (125)I-labelled prolactin to receptors is a saturable and reversible process. Scatchard and Lineweaver-Burk analyses suggest that (125)I-labelled prolactin has a high affinity for its receptor. Binding of (125)I-labelled prolactin to receptors does not result in the destruction of the hormone. Considerable prolactin-binding activity is also observed in subcellular fractions isolated from the adrenal gland, liver, ovary and kidney of the pregnant rabbit, a finding that is consistent with other reported actions of prolactin in these organs.     

Altered aggregational properties in a genetic variant of human glucose 6-phosphate dehydrogenase

The Tel Hashomer variant of human G6PD migrates as two prominent components during electrophoresis in several gel systems in which red cell G6PD from other males migrates predominantly as a single band. Since human males normally have but one X-chromosome, the extra band of this variant seemed an exception to earlier biochemical and genetic observations suggesting that human red cell G6PD is determined by a locus on the X chromosome. Results of the present studies indicate that the Tel Hashomer variant is unusually susceptible to the formation of a complex which has a higher molecular weight than normal G6PD and which represents the slow electrophoretic component. The conditions of formation and disruption of this complex in crude and purified Tel Hashomer preparations suggest that it results from the formation of disulfide bridges between molecules of Tel Hashomer G6PD.Supported by U.S. Public Health Service Research Grants AM-11065 and FR-5406 and Research Career Development Award 5 K3 AM 7992.     

Isologous Oxygen,Sulfur, and Selenium Compounds as Probes of Acetylcholine Receptors

Evidence is presented that while the conformations of acetylcholine and acetylthiolcholine are different, acetylthiolcholine and acetylselenolcholine are structurally and conformationally very similar. Experiments with sulfur and selenium isologs of acetylcholine, choline, and local anesthetics suggest that the active sites of receptors of the electroplax and of electric eel acetylcholinesterase are different, but are compatible with the postulate that acetylcholine receptors of axonal and synaptic excitable membranes are similar.     

Nature of Escherichia coli B(P1) Yielder Cells at the Time of Infection with Restricted T1

In the infection of Escherichia coli B(P1) with restricted T1, it was shown that yielder cells consist of both special and nonspecial cells. Special or predetermined yielders occurred only among the earliest yielders. In most instances, yielder-cell formation was most easily explained by assuming that the first step was a chance escape of the restricted phage DNA from the degrading enzyme of the restricting cell.