No abstract

Acute and chronic exposure to ethanol produces specific changes in several signal transduction cascades. Such alterations in signaling are thought to be a crucial aspect of the central nervous system's adaptive response, which occurs with chronic exposure to ethanol. We have recently identified and isolated several genes whose expression is specifically induced by ethanol in neural cell cultures. The product of one of these genes has extensive sequence homology to phosducin, a phosphoprotein expressed in retina and pineal gland that modulates trimeric guanine nucleotide-binding protein (G protein) function by binding to G-protein fly subunits. We identified from a rat brain cDNA library an isolate encoding the phosducin-like protein (PhLP), which has 41% identity and 65% amino acid homology to phosducin. PhLP cDNA is expressed in all tissues screened by RNA blot-hybridization analysis and shows marked evolutionary conservation on Southern, hybridization. We have identified four forms ofPhLP cDNA varying only in their 5' ends, probably due to alternative splicing. This 5'-end variation generates two predicted forms of PhLP protein that differ by 79 aa at the NH2 terminus.

The purple nonsulfur photosynthetic bacterium Rhodobacter capsulatus regulates synthesis of its photosystem in response to two environmental stimuli, oxygen tension and light intensity. Here we describe the identification and characterization of the trans-acting regulatory gene hvrA, which we show is involved in differentially controlling reaction center and light-harvesting gene expression in response to alterations in light intensity. An hvrA mutant strain is shown to lack the capability to trans-activate light-harvesting-I and reaction center gene expression but retain normal light-harvesting-II and photopigment regulation, in response to a reduction in light intensity. As a consequence of altered expression, hvrA mutant strains exhibit reduced photosynthetic growth capabilities under dim-light conditions. The results of this study and additional studies indicate that regulated synthesis of the photosystem involves complex sets of overlapping regulatory circuits that differentially control photosystem gene expression in response to environmental stimuli such as oxygen tension and light intensity.

Heterotrimeric guanine nucleotide-binding proteins (G proteins) play a major role in transmembrane signaling processes by transducing extracellular signals from the superfamily of heptahelical cell surface receptors to their appropriate intracellular effectors (1, 2). In its trimeric form, G␣␤␥ is inactive, and the G␣ subunit binds a molecule of GDP. Upon ligand binding, the receptor catalyzes the exchange of GDP for GTP on G␣ that causes its activation and dissociation from the tightly bound G␤␥ complex.1 Inactivation and reassociation of the heterotrimer is initiated by the hydrolysis of bound GTP into GDP by an intrinsic GTPase activity of the G␣ subunit. It is now known that both the free GTP-bound G␣ and the G␤␥ dimer can bind and regulate downstream effectors including adenylyl cyclases, phospholipases, and ion channels, and thereby modulate second messenger levels and ion flux (3).The discovery of several specific G␤␥ binding proteins has recently shed light on new roles for G␤␥ in the propagation and termination of cellular signaling. The dimer has been shown to recruit ␤-adrenergic receptor kinase (␤ARK) 2 to its membraneassociated receptor substrate and thus initiate receptor desensitization (4, 5). This process occurs via direct binding of G␤␥ to the C terminus of a putative pleckstrin homology domain on ␤ARK (6). Furthermore, the responsiveness of G protein-regulated signaling systems may be directly modulated through the interaction of G␤␥ subunits with intracellular regulatory proteins. For instance, phosducin, a phosphoprotein mainly expressed in the retina and pineal gland, inhibits the phototransduction cascade by scavenging ␤␥ subunits of the G protein transducin (G t ), thus preventing their reassociation with the G t ␣ subunit (7,8). Because phosducin has a higher affinity for G t ␤␥ than does G t ␣, it has been suggested that the formation of the phosducin/G t ␤␥ complex is a major factor regulating photoreceptor responsiveness (9). From in vitro binding and cotransfection assays, it was proposed that phosducin may also compete with other targets for G␤␥ binding, such as ␤ARK and phospholipase C type ␤2 (10, 11).We recently isolated a rat brain cDNA encoding a phosducinlike protein (PhLP), which has 65% amino acid homology to phosducin (12). We also described several 5Ј-end splice variants that generate two predicted isoforms of the protein: PhLP long (PhLP) of 301 amino acids containing the entire coding sequence and PhLP short (PhLP S ) of 218 amino acids missing the first 83 N-terminal residues of PhLP (12, 13). Based on sequence homology with phosducin, we have suggested that PhLP proteins regulate G␤␥ signaling in nonretinal tissues. In favor of this hypothesis, a recent report showed that recombinant PhLP S inhibits several G␤␥ functions in vitro (14). Interestingly, these authors suggested that unlike phosducin (11,15), the N terminus of PhLP was unlikely to contain a G␤␥-binding domain.To more directly characterize the interaction of PhLP with G␤␥, we studied PhLP binding to G␤␥ both...