The existence of dimers and oligomers for many G protein-coupled receptors has been described by us and others. Since many G protein-coupled receptor subtypes are highly homologous to each other, we examined whether closely related receptors may interact with each other directly and thus have the potential to create novel signaling units. Using -and ␦-opioid receptors, we show that each receptor expressed individually was pharmacologically distinct and could be visualized following electrophoresis as monomers, homodimers, homotetramers, and higher molecular mass oligomers. When -and ␦-opioid receptors were coexpressed, the highly selective synthetic agonists for each had reduced potency and altered rank order, whereas endomorphin-1 and Leu-enkephalin had enhanced affinity, suggesting the formation of a novel binding pocket. No heterodimers were visualized in the membranes coexpressing -and ␦-receptors by the methods available. However, heterooligomers were identified by the ability to co-immunoprecipitate -receptors with ␦-receptors and vice versa using differentially epitope-tagged receptors. In contrast to the individually expressed -and ␦-receptors, the coexpressed receptors showed insensitivity to pertussis toxin and continued signal transduction, likely due to interaction with a different subtype of G protein.In this study, we provide, for the first time, evidence for the direct interaction of -and ␦-opioid receptors to form oligomers, with the generation of novel pharmacology and G protein coupling properties.Opioid receptors have distinct pharmacological profiles and discrete but overlapping distributions in brain. The relatively recent cloning of opioid receptors has established that the products of three genes form the known subtypes (the -, ␦-, and -opioid receptors) that interact with the complex family of endogenous opioid peptides (reviewed in Ref. 1). The endogenous opioid peptide-receptor systems mediate important physiological functions related to pain perception, locomotion, motivation, reward, autonomic function, immunomodulation, and hormone secretion.The analysis of the contribution of each receptor type to the various opioid functions documented has been limited by the selectivity and cross-reactivity of the available opioid ligands and the postulation that multiple receptor subtypes are present. Since the cloning of the opioid receptors, the individual pharmacological and biochemical profiles of the -, ␦-, and -opioid receptors have been better defined; however, there are many aspects of opioid receptor biology that still remain poorly understood. A major problem that still remains is that the pharmacology of opioid receptors in brain tissue predicts a greater number of receptor subsites than revealed by opioid receptor cloning (1, 2). One possible explanation may be that the precise receptor-effector interactions present in endogenous brain regions may have not been adequately replicated in the heterologous expression systems in which the cloned receptors have been studied or, alternativel...

Chronic infection by alginate-producing (mucoid) Pseudomonas aeruginosa is the leading cause of mortality among cystic fibrosis (CF) patients. During the course of sustained infection, the production of an alginate capsule protects the bacteria and allows them to persist in the CF lung. One of the key regulators of alginate synthesis is the algT (aIgU) gene encoding a putative alternative af factor (aE).AlgT was hyperproduced and purified from Escherichia coli. The N-terminal sequence of the purified protein matched perfectly with that predicted from the DNA sequence. The purified protein, in the presence of E. coli RNA polymerase core enzyme, was able to initiate transcription of an algT promoter. Deletion of the -35 region of this promoter abolished this activity in vitro as well as in vivo. These data indicate that the algT gene encodes a cr factor that is autoregulatory.

SummaryThe regulatory protein AlgR2 in Pseudomonas aeruginosa positively regulates nucleoside diphosphate kinase (Ndk) and succinyl-CoA synthetase, enzymes critical in nucleoside triphosphate (NTP) formation. AlgR2 positively regulates the production of alginate, GTP, ppGpp and inorganic polyphosphate (poly P). An algR2 mutant with low levels of these metabolites has them restored by introducing and overexpressing either the algR2 or the ndk gene into the algR2 mutant. Thus, Ndk is involved in the formation of these compounds and largely prevents the death of the algR2 mutant, which occurs early in the stationary phase. We demonstrate that the 12 kDa Ndk-pyruvate kinase (Pk) complex, previously shown to generate predominantly GTP instead of all the NTPs, has a low affinity for the deoxynucleoside diphosphates and cannot generate the dNTPs needed for DNA replication and cell division; this complex may thus be involved in regulating the levels of both NTPs and dNTPs that modulate cell division and survival in the stationary phase.

The serotonin (5-hydroxytryptamine (5-HT)) 1B and 1D receptor subtypes share a high amino acid sequence identity and have similar ligand binding properties. In this study, we demonstrate that both receptor subtypes exist as monomers and homodimers when expressed alone and as monomers and heterodimers when co-expressed. Gene expression studies have shown that there are brain regions where the 5-HT 1B and 5-HT 1D receptors are co-localized and where heterodimerization may occur physiologically. This is the first direct visualization of the physical association between G protein-coupled receptors of different subtypes.z 1999 Federation of European Biochemical Societies.