In view of the importance of molecular sensing in the function of the gastrointestinal (GI) tract, we assessed whether signal transduction proteins that mediate taste signaling are expressed in cells of the human gut. Here, we demonstrated that the alpha-subunit of the taste-specific G protein gustducin (Galpha(gust)) is expressed prominently in cells of the human colon that also contain chromogranin A, an established marker of endocrine cells. Double-labeling immunofluorescence and staining of serial sections demonstrated that Galpha(gust) localized to enteroendocrine L cells that express peptide YY and glucagon-like peptide-1 in the human colonic mucosa. We also found expression of transcripts encoding human type 2 receptor (hT2R) family members, hT1R3, and Galpha(gust) in the human colon and in the human intestinal endocrine cell lines (HuTu-80 and NCI-H716 cells). Stimulation of HuTu-80 or NCI-H716 cells with the bitter-tasting compound phenylthiocarbamide, which binds hT2R38, induced a rapid increase in the intracellular Ca2+ concentration in these cells. The identification of Galpha(gust) and chemosensory receptors that perceive chemical components of ingested substances, including drugs and toxins, in open enteroendocrine L cells has important implications for understanding molecular sensing in the human GI tract and for developing novel therapeutic compounds that modify the function of these receptors in the gut.

We previously demonstrated the expression of bitter taste receptors of the type 2 family (T2R) and the ␣-subunits of the G protein gustducin (G␣ gust) in the rodent gastrointestinal (GI) tract and in GI endocrine cells. In this study, we characterized mechanisms of Ca 2ϩ fluxes induced by two distinct T2R ligands: denatonium benzoate (DB) and phenylthiocarbamide (PTC), in mouse enteroendocrine cell line STC-1. Both DB and PTC induced a marked increase in intracellular [Ca 2ϩ (3,13,19,60). Molecular sensing by GI cells plays a critical role in the control of multiple fundamental functions, including digestion, food intake, and metabolic regulation. Although these fundamental control systems have been known for a considerable time, the initial molecular recognition events that sense the chemical composition of the luminal contents have remained poorly understood. The gustatory system has been selected during evolution to detect nonvolatile nutritive and beneficial (sweet) compounds as well as potentially harmful (bitter) substances (24, 34). In particular, bitter taste has evolved as a central warning signal against the ingestion of potentially toxic substances, including plant alkaloids and other environmental toxins (21, 65). Specialized neuroepithelial taste receptor cells, organized within taste buds in human and rodent lingual epithelium, expressed a family of bitter taste receptors (referred as T2Rs) (1, 6, 46). These putative taste receptors belong to the guanine nucleotide-binding regulatory protein (G protein)-coupled receptor (GPCR) superfamily (1), which are characterized by seven transmembrane ␣-helices (32). Extensive genetic and biochemical evidence indicate that specific G proteins, gustducin and transducin, mediate bitter and sweet gustatory signals in the taste buds of the lingual epithelium (47,48,62,63,73). More recently, phospholipase C␤ 2 (PLC␤ 2 ) and TRPM5, a member (melastatin subtype 5) of the transient receptor potential (TRP) family (49), have been linked to bitter and sweet signal transduction (55, 56, 81). There is evidence for the activation of multiple second messenger pathways and ion channels in individual taste cells (1, 82). Clearly, taste signal transduction is complex and multifactorial and there is still much that is unknown about individual taste cell regulation.Outside the tongue, expression of the ␣-subunit of gustducin (G␣ gust ) has been also localized to gastric (28, 75) and pancreatic (27) cells, suggesting that a taste-sensing mechanism may also exist in the digestive system. Indeed, we demonstrated the expression of members of the bitter taste receptors of the T2R family in the mouse and rat GI tract and in mouse and rat enteroendocrine cells in culture (74,75). More recently, these results have been confirmed (45) and extended to the expression of the sweet taste receptors of the T1R family (15). Collectively, these findings demonstrated the expression of taste signal transduction pathways in cells of the GI tract of mice and rats.The intracellular signal transd...

Mammalian type 2 taste receptors (T2R) are a family of G protein-coupled receptors that mediate bitter signals in taste cells. In the present study, we compared the genomic organization of rodent T2R genes based on the recently completed mouse and rat genomes and examined tissue- and cell-specific expression of T2Rs. Both mouse and rat T2R families consist of 36 intact genes and at least 7 pseudogenes that are mapped to mouse chromosomes 15, 2, and 6 and to rat chromosomes 2, 3, and 4, respectively. All but two T2R genes are clustered on mouse chromosome 6 and rat chromosome 4 with virtually identical genomic organization. The orthologs of the first human T2R gene identified, mT2R119 and rT2R1, are located on mouse chromosome 15 and rat chromosome 2, whereas the novel rodent-specific T2R genes, mT2R134 and rT2R34, are located on mouse chromosome 2 and rat chromosome 3, respectively. Our results, using RT-PCR, demonstrate the presence of transcripts corresponding to the putative denatonium benzoate (DB) and phenylthiocarbamide (PTC) receptors in the antrum, fundus, and duodenum as well as in STC-1 and AR42J cells. The novel rodent-specific T2R gene (mT2R134 and rT2R34) was also expressed in these tissues and cell lines. The addition of DB, PTC, or cycloheximide to AR42J cells induced a rapid increase in the intracellular Ca2+ concentration. The specificity of these effects is shown by the fact that these bitter stimuli did not induce any detectable Ca2+ signaling in many other rodent or human cells that do not express receptors or G proteins implicated in bitter taste signaling. These results demonstrate that mouse and rat T2R genes are highly conserved in terms of genomic organization and tissue expression, suggesting that rodent T2Rs are evolved under similar dietary pressure and share bitter sensing functions in the lingual and gastrointestinal systems.

Scutellaria baicalensis (SB) and SB-derived polyphenols possess anti-proliferative activities in several cancers, including pancreatic cancer (PaCa). However, the precise molecular mechanisms have not been fully defined. SB extract and SB-derived polyphenols (wogonin, baicalin, and baicalein) were used to determine their anti-proliferative mechanisms. Baicalein significantly inhibited the proliferation of PaCa cell lines in a dose-dependent manner, whereas wogonin and baicalin exhibited a much less robust effect. Treatment with baicalein induced apoptosis with release of cytochrome c from mitochondria, and activation of caspase-3 and -7 and PARP. The general caspase inhibitor zVAD-fmk reversed baicalein-induced apoptosis, indicating a caspase-dependent mechanism. Baicalein decreased expression of Mcl-1, an anti-apoptotic member of the Bcl-2 protein family, presumably through a transcriptional mechanism. Genetic knockdown of Mcl-1 resulted in marked induction of apoptosis. The effect of baicalein on apoptosis was significantly attenuated by Mcl-1 over-expression, suggesting a critical role of Mcl-1 in this process. Our results provide evidence that baicalein induces apoptosis in pancreatic cancer cells through down-regulation of the anti-apoptotic Mcl-1 protein.