Gary L. McKnight

IL-28, IL-29 and their class II cytokine receptor IL-28R

Cytokines play a critical role in modulating the innate and adaptive immune systems. Here, we have identified from the human genomic sequence a family of three cytokines, designated interleukin 28A (IL-28A), IL-28B and IL-29, that are distantly related to type I interferons (IFNs) and the IL-10 family. We found that like type I IFNs, IL-28 and IL-29 were induced by viral infection and showed antiviral activity. However, IL-28 and IL-29 interacted with a heterodimeric class II cytokine receptor that consisted of IL-10 receptor β (IL-10Rβ) and an orphan class II receptor chain, designated IL-28Rα. This newly described cytokine family may serve as an alternative to type I IFNs in providing immunity to viral infection.

Nucleotide sequence of the triosephosphate isomerase gene from Aspergillus nidulans: Implications for a differential loss of introns

A functional cDNA from Aspergillus nidulans encoding triosephosphate isomerase (TPI) was isolated by its ability to complement a tpi1 mutation in Saccharomyces cerevisiae. This cDNA was used to obtain the corresponding gene, tpiA. Alignment of the cDNA and genomic DNA nucleotide sequences indicated that tpiA contains five introns. The intron positions in the tpiA gene were compared with those in the TPI genes of human, chicken, and maize. One intron is present at an identical position in all four organisms, two other introns are located in similar positions in A. nidulans and maize, and the remaining two introns are unique to A. nidulans. These Aspergillus-specific introns are located in regions of the protein that were predicted to be interrupted by introns based on analysis of a Go plot of chicken TPI. These comparisons are discussed in relation to the evolution of introns within TPI genes.

Glucose-dependent insulinotropic polypeptide stimulation of lipolysis in differentiated 3T3-L1 cells: wortmannin-sensitive inhibition by insulin.

GIP is an important insulinotropic hormone (incretin) that has also been implicated in fat metabolism. There is controversy regarding the actions of GIP on adipocytes. In the current study, the existence of GIP receptors and effects of GIP on lipolysis were studied in differentiated 3T3-L1 cells. GIP receptor messenger RNA was detected by RT-PCR and RNase protection assay. Receptors were detected in binding studies (IC50 26.7 ± 0.7 nm). GIP stimulated glycerol release with an EC50 of 3.28 ± 0.63 nm. GIP (10−9–10−7 m) + IBMX increased cAMP production by 1180–2246%. The adenylyl cyclase inhibitor MDL 12330A (10−4 m) inhibited GIP-induced glycerol production by >90%, and reduced cAMP responses to basal. Preincubation of 3T3-L1 cells with insulin inhibited glycerol responses to GIP, and the inhibitory effect of insulin was blocked by the phosphatidylinositol 3′-kinase inhibitor, wortmannin. It is concluded that GIP stimulates glycerol release in 3T3-L1 cells primarily via stimulation of cAMP production, and t...

Canine galanin: sequence, expression and pancreatic effects.

To determine if dog galanin is a potent inhibitor of dog insulin secretion we determined its primary structure from its cloned cDNA, evaluated its expression in celiac ganglia and determined its effect on islet hormone secretion. The predicted amino acid sequence differs from the other known species of galanin by three to six amino acids in the C-terminal half of the molecule. In situ hybridization revealed the presence of dog progalanin mRNA in every neuronal cell body in the dog celiac ganglion. The predicted dog galanin peptide was synthesized and infused i.v. at 0.25, 2.5, 25 or 250 pmol/kg/min. It potently inhibited insulin secretion, less potently inhibited pancreatic somatostatin release and stimulated glucagon secretion, similar to the effects of porcine galanin in the dog. In summary, dog galanin is expressed in the neuronal cell bodies that innervate the pancreas and the sequence of the dog galanin preserves the potent insulin inhibitory part of the galanin molecule. These data support the hypothesis that galanin is a sympathetic neurotransmitter in dog pancreas.

Sequence of Human Galanin and Its Inhibition of Glucose-Stimulated Insulin Secretion From RIN Cells

Human progalanin cDNA was cloned with polymerase chain reaction techniques. The cDNA sequence predicts that the human form of galanin has a substitution of the glycine residue found at position 30 in other species and thus is likely to retain this residue during posttranslational processing and not be amidated at the COOH terminus. Furthermore, the cDNA sequence predicts three additional amino acid substitutions compared with known galanins. Human galanin was synthesized, and its bioactivity was compared with porcine and rat galanin based on inhibition of insulin release from a glucose-responsive rat insulinoma (RIN) cell line. Human galanin inhibited glucose-stimulated insulin secretion in adose-dependent manner in RIN cells. Human, porcine, and rat galanin exhibited similar activity with ED50 <1 nM.