Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UMLS:C0043167 (pertussis)
 19,595 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Heterotrimeric G-proteins function as signal transducers for a variety of hormone-coupled enzyme systems in eukaryotic cells. In LLC-PK1 renal cells, vasopressin-stimulated adenylylcyclase activity is regulated in part, by the counterbalancing activity of stimulatory G-proteins (Gs) and inhibitory pertussis toxin-sensitive G-proteins (Gi). Two Gi genes encoding the Gi isoforms G alpha i-2 and G alpha i-3 are expressed in LLC-PK1 cells. In polarized cells, these isoforms are topographically segregated to different membranes, which allows for the selective inhibition of adenylylcyclase by G alpha i-2. The genes encoding these isoforms are similarly regulated in these cells during growth and differentiation but differ in response to steroid hormone signals (Holtzman, E.J., Kinane, T.B., West, K., Soper, B.W., Karga, H., Ausiello, D.A., and Ercolani, L. (1993) J. Biol. Chem. 268, 3964-3975). We now demonstrate after stimulating polarized LLC-PK1 cells with forskolin, which raises intracellular cAMP levels 50-fold, G alpha i-2 but not G alpha i-3 protein is increased 3-fold at 12 h and remains elevated above control values by 24 h. In cells stably transfected with G alpha i-2 or G alpha i-3 gene 5'-flanking sequences fused to firefly luciferase cDNA reporter gene, forskolin treatment increased G alpha i-2 transcription 3-fold but inhibited G alpha i-3 transcription by 50% at 12 h. In vivo footprinting of forskolin-treated cells was performed to examine the molecular basis for activation of the G alpha i-2 gene. Protected guanosines were identified in a 135-base pair (bp) area previously associated with enhancer activity of this gene in non-polarized cells. This DNA segment did not contain the classical cAMP response element 5'-TGACGTCA-3'. Utilizing the 135-bp DNA segment as a probe in mobility shift assays, which compared nuclear extracts from cells before and after forskolin treatment, an induced nuclear protein complex was identified. Following systematic reduction and mutation of this DNA segment, a "CCAAT" box motif was identified that bound the induced nuclear protein complex during forskolin-induced G alpha i-2 gene transcriptional activation. Induction of this nuclear protein complex was prevented in forskolin-treated cells by cycloheximide. To demonstrate functional activity of the CCAAT box motif, cells were transiently transfected with plasmids encoding either the minimal 135-bp segment or a multimerized CCAAT box segment fused to a Rous sarcoma minimal promoter/firefly luciferase reporter gene.(ABSTRACT TRUNCATED AT 400 WORDS)
 ...
PMID:cAMP regulates G-protein alpha i-2 subunit gene transcription in polarized LLC-PK1 cells by induction of a CCAAT box nuclear binding factor. 822 26

Heterotrimeric guanine nucleotide-binding proteins (G-proteins) function as signal transducers for a variety of hormone-coupled enzyme and ion transport systems in eukaryotic cells. The expression of pertussis toxin-sensitive G-proteins (Gi) which couple their cognate receptors and effectors are regulated by cell cycle-dependent events in porcine LLC-PK1 renal epithelial cells. G alpha i-2 and G alpha i-3 isoforms are detected in these cells, and like G alpha i-2 (Holtzman, E. J., Soper, B. W., Stow, L. L., Ausiello, D. A., and Ercolani, L. (1991) J. Biol. Chem. 266, 1763-1771), we now demonstrate that G alpha i-3 mRNA and protein is coordinately expressed in these cells during differentiation. To gain further insights into these events, the porcine G alpha i-3 gene minimal promoter was characterized and found 67 base pairs upstream from the major transcription start site. The 56-base pair minimal promoter lacked TATAAA and GC boxes but did contain a sequence GGAAGTG conserved in both the human and porcine gene that could potentially bind an adenovirus E4TF1 transcription factor. In cells stably transfected with G alpha i-2 or G alpha i-3 gene 5'-flanking sequences fused to firefly luciferase cDNA reporter, temporal 10-15-fold transcriptional activation of both genes occurred before cellular polarization. Utilizing mobility shift assays which compared nuclear extracts from cells before and after cell polarization, a motif in the 5' region of the gene promoter GTACTTCCGCT was identified that bound an induced nuclear protein complex during transcriptional activation. In polarized cells complemented with the human glucocorticoid receptor, dexamethasone decreased G alpha i-2 but increased G alpha i-3 basal transcription and mRNA content 3-fold. These studies demonstrate that both G alpha i genes are dynamically regulated in LLC-PK1 cells by both growth, differentiation, and hormone signals.
 ...
PMID:Transcriptional regulation of G-protein alpha i subunit genes in LLC-PK1 renal cells and characterization of the porcine G alpha 1-3 gene promoter. 844 Jun 88

A Chinese hamster ovary (CHO) cell line expressing the firefly luciferase gene under the control of six cAMP response elements (CREs) was stably transfected with the long form of the rat D2 dopamine receptor. Saturation binding analysis using [3H]spiperone showed that the receptor was expressed at low levels (Bmax = 96.5+/-15.8 fmol/mg), but with an affinity characteristic of the D2 receptor (Kd = 21.5+/-3.7 pM). Luciferase expression in this cell line was modified in a dose dependent manner with dopamine receptor agonists (N-propylapomorphine > apomorphine > quinpirole > dopamine) and antagonists (spiperone > (+)-butaclamol > D0710 > (-)-sulpiride > tiapride > remoxipride), according to their rank order of potency in binding and cAMP accumulation studies. Dopamine-mediated inhibition of forskolin-stimulated luciferase expression was pertussis toxin sensitive. This demonstrated the efficiency of the luciferase reporter gene assay for the functional testing of D2 dopamine receptors, which are negatively coupled to the adenylyl cyclase signaling pathway, when heterogously expressed at low levels in CHO cells.
 ...
PMID:Functional analysis of the D2L dopamine receptor expressed in a cAMP-responsive luciferase reporter cell line. 969 85

Regulation of the mitogen-activated protein kinase (MAPK) family by prolactin-releasing peptide (PrRP) in both GH3 rat pituitary tumor cells and primary cultures of rat anterior pituitary cells was investigated. PrRP rapidly and transiently activated extracellular signal-regulated protein kinase (ERK) in both types of cells. Both pertussis toxin, which inactivates G(i)/G(o) proteins, and exogenous expression of a peptide derived from the carboxyl terminus of the beta-adrenergic receptor kinase I, which specifically blocks signaling mediated by the betagamma subunits of G proteins, completely blocked the PrRP-induced ERK activation, suggesting the involvement of G(i)/G(o) proteins in the PrRP-induced ERK activation. Down-regulation of cellular protein kinase C did not significantly inhibit the PrRP-induced ERK activation, suggesting that a protein kinase C-independent pathway is mainly involved. PrRP-induced ERK activation was not dependent on either extracellular Ca(2+) or intracellular Ca(2+). However, the ERK cascade was not the only route by which PrRP communicated with the nucleus. JNK was also shown to be significantly activated in response to PrRP. JNK activation in response to PrRP was slower than ERK activation. Moreover, to determine whether a MAPK family cascade regulates rat prolactin (rPRL) promoter activity, we transfected the intact rPRL promoter ligated to the firefly luciferase reporter gene into GH3 cells. PrRP activated the rPRL promoter activity in a time-dependent manner. Co-transfection with a catalytically inactive form of a MAPK construct or a dominant negative JNK, partially but significantly inhibited the induction of the rPRL promoter by PrRP. Furthermore, co-transfection with a dominant negative Ets completely abolished the response of the rPRL promoter to PrRP. These results suggest that PrRP differentially activates ERK and JNK, and both cascades are necessary to elicit rPRL promoter activity in an Ets-dependent mechanism.
 ...
PMID:Prolactin-releasing peptide activation of the prolactin promoter is differentially mediated by extracellular signal-regulated protein kinase and c-Jun N-terminal protein kinase. 1065 64

The agonist-bound gonadotropin-releasing hormone (GnRH) receptor engages several distinct signaling cascades, and it has recently been proposed that coupling of a single type of receptor to multiple G proteins (G(q), G(s), and G(i)) is responsible for this behavior. GnRH-dependent signaling was studied in gonadotropic alphaT3-1 cells endogenously expressing the murine receptor and in CHO-K1 (CHO#3) and COS-7 cells transfected with the human GnRH receptor cDNA. In all cell systems studied, GnRH-induced phospholipase C activation and Ca(2+) mobilization was pertussis toxin-insensitive, as was GnRH-mediated extracellular signal-regulated kinase activation. Whereas the G(i)-coupled m2 muscarinic receptor interacted with a chimeric G(s) protein (G(s)i5) containing the C-terminal five amino acids of Galpha(i2), the human GnRH receptor was unable to activate the G protein chimera. GnRH challenge of alphaT3-1, CHO#3 and of GnRH receptor-expressing COS-7 cells did not result in agonist-dependent cAMP formation. GnRH challenge of CHO#3 cells expressing a cAMP-responsive element-driven firefly luciferase did not result in increased reporter gene expression. However, coexpression of the human GnRH receptor and adenylyl cyclase I in COS-7 cells led to clearly discernible GnRH-dependent cAMP formation subsequent to GnRH-elicited rises in [Ca(2+)](i). In alphaT3-1 and CHO#3 cell membranes, addition of [alpha-(32)P]GTP azidoanilide resulted in GnRH receptor-dependent labeling of Galpha(q/11) but not of Galpha(i), Galpha(s) or Galpha(12/13) proteins. Thus, the murine and human GnRH receptors exclusively couple to G proteins of the G(q/11) family. Multiple GnRH-dependent signaling pathways are therefore initiated downstream of the receptor/G protein interface and are not indicative of a multiple G protein coupling potential of the GnRH receptor.
 ...
PMID:Gonadotropin-releasing hormone receptor initiates multiple signaling pathways by exclusively coupling to G(q/11) proteins. 1073 55

Regulation of the PI3K-protein kinase B/Akt (serine/threonine kinase) cascade by PRL-releasing peptide (PrRP) and insulin in GH3 rat pituitary tumor cells was investigated. PrRP and insulin rapidly and transiently stimulated the activation of Akt, and the PI3K inhibitor wortmannin blocked the PrRP- or insulin-induced activation of Akt. Both pertussis toxin (10 ng/ml), which inactivates Gi/Go proteins, and expression of a peptide derived from the carboxyl terminus of the beta-adrenergic receptor kinase I, which specifically blocks signaling mediated by the betagamma subunits of G proteins, completely blocked the PrRP-induced Akt activation, suggesting that Gi/Go proteins are involved in PrRP-induced Akt activation, as they are in the activation of ERK by PrRP. Moreover, to determine whether a PI3K-Akt cascade regulates rat PRL (rPRL) promoter activity, we transfected the intact rPRL promoter ligated to the firefly luciferase reporter gene into GH3 cells. PrRP and insulin activated the rPRL promoter activity. Pretreatment with wortmannin or cotransfection with a dominant-negative Akt partially but significantly inhibited the induction of the rPRL promoter by PrRP or insulin. Cotransfection with a constitutively active Akt induced the rPRL promoter activity and cotransfection with a dominant-negative cAMP response element-binding protein (CREB) completely abolished the response of the rPRL promoter to the constitutively active Akt. Furthermore, either treatment with PrRP and insulin or transfection with the constitutively active Akt induced the phosphorylation of CREB. These results suggest that PrRP and insulin activate a PI3K-Akt cascade that is necessary to elicit rPRL promoter activity via a CREB-dependent mechanism.
 ...
PMID:Regulation of the PRL promoter by Akt through cAMP response element binding protein. 1175 85