The human glucagon receptor encoding gene: structure, cDNA sequence and chromosomal localization

Characterization of the human glucagon-receptor-encoding gene (GGR) should provide a greater understanding of blood glucose regulation and may reveal a genetic basis for the pathogenesis of diabetes. A cDNA encoding a complete functional human glucagon receptor (GGR) was isolated from a liver cDNA library by a combination of polymerase chain reaction and colony hybridization. The cDNA encodes a receptor protein with 80% identity to rat GGR that binds [¹²⁵I] glucagon and transduces a signal leading to increases in the concentration of intracellular cyclic adenosine 3′,5′-monophosphate. Southern blot analysis of human DNA reveals a hybridization pattern consistent with a single GGR locus. In situ hybridization to metaphase chromosome preparations maps the GGR locus to chromosome 17q25. Analysis of the genomic sequence shows that the coding region spans over 5.5 kb and is interrupted by 12 introns.     

White Ibis integument color during the breeding season

ABSTRACT.   Many birds undergo bare part color changes during the breeding season. Most investigators have focused on color as a signal of individual quality. An alternative, but not exclusive, function of bare part color may be signaling readiness to breed, especially in colonial, asynchronous breeders. White Ibises ( Eudocimus albus ) are colonial waterbirds that show vivid bare part colors on their bills and legs during reproduction. We quantified bill and leg colors to describe color changes and their possible relationship to reproductive status during the breeding season of White Ibises in the Florida Everglades from 1998 to 2001. We also examined the correlation between bare part colors and circulating concentrations of sex steroids to understand the factors that regulate bare part colors. During the display stage, male and female ibises developed dark pink bills and scarlet legs. As the breeding season progressed, bills and legs faded and developed a muted pink hue. The bare part colors of female ibises were correlated with testosterone concentrations, but those of male ibises were not correlated with any hormones. A discriminant function analysis based on principal component scores (representing variation in saturation and hue) and the amount of black on the bill successfully classified ibis reproductive stage 94% of the time. The use of bare part colors to determine reproductive status may be useful for studying reproduction in colonially nesting birds, where access to breeding sites can be difficult and potential for researcher disturbance is high.     

Developmental changes in the facial morphology of the Borneo orangutan (Pongo pygmaeus): possible signals in visual communication

Orangutans display remarkable developmental changes and sexual differences in facial morphology, such as the flanges or cheek-pads that develop only on the face of dominant adult males. These changes suggest that facial morphology is an important factor in visual communication. However, developmental changes in facial morphology have not been examined in detail. We studied developmental changes in the facial morphology of the Borneo orangutan (Pongo pygmaeus) by observing 79 individuals of various ages living in the Sepilok Orangutan Rehabilitation Centre (SORC) in Malaysia and in Japanese zoos. We also analyzed photographs of one captive male that were taken over a period of more than 16 years. There were clear morphological changes that occurred with growth, and we identified previously unreported sexual and developmental differences in facial morphology. Light-colored skin around the eyes and mouth is most prominent in animals younger than 3 years, and rapidly decreases in area through the age of approximately 7 years. At the same time, the scattered, erect hairs on the head (infant hair) become thick, dense hairs lying on the head (adult hair) in both sexes. The results suggest that these features are infant signals, and that adult signals may include darkened face color, adult hair, whiskers, and a beard, which begin to develop after the age of approximately 7 years in both sexes. In females, the eyelids remain white even after 10 years, and turn black at around the age of 20; in males, the eyelids turn black before the age of 10. The whiskers and beards of adults are thicker in males than in females, and are fully developed before the age of 10 in males, while they begin to develop in females only after approximately 20 years. White eyelids and undeveloped whiskers and beards may be visual signals that are indicative of young adult females. Our results also show that the facial morphology of the unflanged male is similar to that of the adult female, although it has also been pointed out that unflanged males resemble younger individuals.     

Discrepancy between GLUT4 translocation and glucose uptake after ischemia

Objective: Low-flow ischemia results in glucose transporter translocation and in increased glucose uptake. After total ischemia in rat heart, we found no increase in glucose uptake. Here we test the hypothesis that total ischemia is associated with decreased activation of GLUT4 despite translocation. Methods: Isolated working hearts (n=70, Sprague–Dawley rats) were perfused for 70 min at physiological workload with Krebs–Henseleit buffer containing [2-³H]glucose (5 mmol/l, 0.05 μCi/ml) with either oleate (0.4 mmol/l, 1%BSA) or pyruvate (5 mmol/l, 1%BSA). After 20 min, hearts were subjected to 15 min of total ischemia followed by 35 min of reperfusion. We measured glucose uptake and intracellular free glucose (IFG) using [2-³H]glucose and [¹⁴C]sucrose, and determined the distribution of GLUT4 by colocalization immunofluorescence with Na–K ATP-ase. Results: Cardiac power was 10.1 ± 0.90 mW before ischemia and did not differ between groups. Recovery was the same in both groups (55.7 ± 24.8$%). Glucose uptake did not differ between groups before ischemia, and did not increase during reperfusion. Despite evidence of GLUT4 translocation after reperfusion in both groups, IFG did not increase compared with before ischemia. Conclusion: We conclude that there is a discrepancy between glucose transporter availability and glucose uptake after ischemia, which may be due to inhibition of GLUT4 in the plasma membrane. (Mol Cell Biochem 278: 129–137, 2005)     

Cardiac syntrophin isoforms: Species-dependent expression, association with dystrophin complex and subcellular localization

Syntrophin is known to be a component of the dystrophin-glycoprotein complex (DGC), a membrane/cytoskeleton-anchoring structure that is essential for the maintenance of viability of sarcolemma. We purified DGC from hearts of human and several animal species, and compared their protein composition. While almost all components of DGC were present in various species, proteins with the apparent molecular mass of 50–65 kDa corresponding to syntrophin isoforms were very different among them. Three isoforms of syntrophin (1, 1, 2) were expressed in hamster, rat and canine ventricles, whereas only 1-isoform was mainly expressed in human and rabbit ventricles. Immunohistochemical analysis revealed that 1-and 2-syntrophins were co-localized in sarcolemma and in T-tubules of canine ventricles. However, despite membrane localization of most syntrophins, subcellular fractionation revealed that part of syntrophins were recovered in the cytosolic fraction devoid of other components of DGC, raising the possibility that syntrophins may play multiple roles in various intracellular sites of cardiac muscle cells. Species-dependent expression and unique subcellular localization of syntrophins in cardiac muscle may contribute to the variable severity of muscle dysgenesis caused by the same primary defect in components of DGC of human and other animal species. (Mol Cell Biochem 268: 59–66, 2005)     

Glutamate Regulates Dystrophin-71 levels in Glia Cells

Glial glutamate receptors are likely to be involved in neuronal differentiation, migration, and plasticity. Dystrophin, the protein defective in Duchenne muscular dystrophy (DMD) is widely expressed in the Central Nervous System. Activation of internal promoters of the DMD gene leads to the production of several proteins, the Dystrophin-71 (Dp-71) being the most abundant in the encephalon. This protein is known to stabilize neurotransmitter receptors in clusters and its absence has been correlated with cognitive deficits in a mouse model. Using cultured chick Bergmann glia cells and mouse cerebellar fusiform astrocytes, we demonstrate here that glutamate receptor activation results in a time and dose dependent decrease of Dp-71 levels. This effect is mediated through amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors. The present results suggest an involvement of Dp-71 in glutamate receptor signaling and possibly clustering and further support the notion of an active role of glia in the physiology of glutamatergic transmission.     

Determinants for nasal trigeminal detection of volatile organic compounds

We explored the influence of methodological and chemical parameters on the detection of nasal chemesthesis (i.e., trigeminal stimulation) evoked by volatile organic compounds (VOCs). To avoid odor biases, chemesthesis was probed via nasal pungency detection in anosmics and via nasal localization (i.e., lateralization) in normosmics, in both cases using forced-choice procedures. In the experiments with anosmics, 12 neat VOCs were selected based on previous reports of lack of chemesthetic response. Although none of the VOCs reached 100% detection, detectability and confidence of detection were higher when using a glass vessel system adapted with nosepieces to fit the nostrils tightly than when using wide-mouth glass jars. Half the stimuli were detected well above chance and half were not. When the latter were tested again after being heated to 37 degrees C, that is, body temperature (from room temperature, 23 degrees C), to increase their vapor concentration, only one, octane, significantly increased its detectability. Chemesthesis gauged with normosmics mirrored that with anosmics. Gas chromatography measurements showed that, even at 23 degrees C, the saturated vapor concentrations of the undetected stimuli, except vanillin, were well above the respective calculated nasal pungency threshold (NPT) from an equation that, in the past, had accurately described and predicted NPTs. We conclude that, except for octane and perhaps vanillin, the failure of the other four VOCs to precipitate nasal chemesthesis rests on a chemical-structural limitation, for example, the molecules lack a key property to fit a receptor pocket, rather than on a concentration limitation, for example, the vapor concentration is too low to reach a threshold value.     

Identification of an RNA-binding domain in human SRP72

The signal recognition particle (SRP) is a ribonucleoprotein complex that plays a crucial role during the delivery of secretory proteins from the ribosome to the cell membrane. Among the six proteins of the eukaryotic SRP, the 72 kDa protein (SRP72) is the largest and least characterized. Polypeptides corresponding to various regions of the entire human SRP72 sequence were expressed in Escherichia coli, purified, and partially proteolyzed. Human SRP RNA bound with high affinity to a 63 amino acid residue region near the C terminus of SRP72. Mild treatment of the fragment with chymotrypsin abolished its RNA-binding activity. A conserved sequence with the consensus PDPXRWLPXXER was identified within a 56 amino acid residue RNA-binding domain. Sucrose gradient centrifugation and filter-binding analysis using mutant SRP RNAs showed that SRP72 bound to the moderately conserved portion of SRP RNA helix 5. Nine tetratricopeptide-like repeats (TPRs) poised to interact with other SRP or ribosomal proteins were predicted in the NH2-terminal region. These identifications assign two important functions to a large portion of SRP72 and demonstrate the RNA-binding capacity of the protein.     

The core TatABC complex of the twin-arginine translocase in Escherichia coli: TatC drives assembly whereas TatA is essential for stability

Current models for the action of the twin-arginine translocation (Tat) system propose that substrates bind initially to the TatBC subunits, after which a separate TatA complex is recruited to form an active translocon. Here, we have studied the roles of individual subunits in the assembly and stability of the core TatBC-containing substrate-binding complex. Previous studies have shown that TatB and TatC are active when fused together; we show here that deletion of the entire TatB transmembrane span from this Tat(BC) fusion inactivates the Tat system but does not affect assembly of the core complex. In this mutated complex, TatA is present but more loosely bound, indicating a role for TatB in the correct binding of TatA. In the absence of TatA, the truncated TatBC fusion protein still assembles into a complex of the correct magnitude, demonstrating that the transmembrane spans of TatC are the only determinants within the membrane bilayer that specify assembly of this complex. Further studies on both the Tat(BC) construct and the wild-type TatBC subunits show that the TatBC complex is unstable in the absence of TatA, and we show that TatA stabilises the TatB subunit specifically within this complex. The results demonstrate a dual role and location for TatA: in the functioning/maintenance of the core complex, and as a separate homo-oligomeric complex.