CncC/Maf-mediated xenobiotic response pathway in insects.

Insects have evolved resistance to almost all insecticides developed for their control. Multiple mechanisms of resistance, including enhanced metabolism and excretion of insecticides, target-site insensitivity, reduced penetration of insecticides, and avoidance behavior, have been reported. The genes coding for proteins involved in resistance have been identified in numerous insects. The enzymes and transporters required for all three phases of insecticide metabolism and excretion including cytochrome P450 monooxygenases, glutathione S-transferases, UDP-glucuronosyltransferases, carboxylesterases, and ATP-binding cassette transmembrane transporters have been identified. Recent research in multiple insect species identified CNC-bZIP transcription factor superfamily members as regulators of genes coding for enzymes and transporters involved in insecticide metabolic resistance. The information on the pathway including reactive oxygen species, cap "n" collar isoform-C, and its heterodimer partner, muscle aponeurosis fibromatosis transcription factors involved in overexpression of enzymes and transporters involved insecticide resistance will be summarized.

No change in mRNA expression of immune-related genes in peripheral blood mononuclear cells challenged with Theileria annulata in Murrah buffalo (Bubalus bubalis).

Water buffaloes (Bubalus bubalis) act as carrier to Theileria annulata and show less clinical sign of tropical theileriosis as compared to indigenous and exotic cattle. Differential expression of immune-related genes such as major histocompatibility complex, class II, DQ alpha 1 (MHC-DQα), signal-regulatory protein alpha (SIRPA), prion protein (PRNP), Toll-like receptor 10 (TLR10), c-musculoaponeurotic fibrosarcoma oncogene homolog (cMAF) and V-maf avian musculoaponeurotic fibrosarcoma oncogene homolog B (MAFB) genes influence host resistance to this disease in exotic, crossbred and indigenous cattle. In the present study we examined the differential mRNA expression of the abovesaid immune-related genes in response to T. annulata infection in buffaloes. Peripheral blood mononuclear cells (PBMCs) harvested from blood samples of buffaloes were challenged with ground-up tick supernatant carrying T. annulata sporozoites in vitro. After 48h of in vitro challenge qPCR was employed to measure the relative mRNA expression of MHC-DQα, SIRPA, PRNP, TLR10, cMAF and MAFB genes in infected and control PBMCs. In the current study, the selected genes showed no change in mRNA expression after T.annulata infection which indicates that they have little role in providing host resistance to theileriosis in buffaloes.

Activation of PPARδ up-regulates the expression of insulin gene transcription factor MafA and ameliorates glucose-induced insulin secretion impaired by palmitate.

PPARδ, a member of the peroxisome proliferator-activated receptor superfamily, plays a key role in the transcriptional regulation of genes involved in cellular lipid and energy metabolism. Therefore, PPARδ may represent a new target for the treatment of obesity, hyperlipidemia, and type 2 diabetes. MafA is a ß-cell-specific and glucose-regulated transcriptional activator for insulin gene expression and plays a crucial role in pancreas development, ß-cell differentiation as well as maintenance of ß-cell function. However, little is known about how PPARδ regulates MafA and ameliorates glucose-stimulated insulin secretion impaired by free fatty acids (FFA). In the present study, we evaluated the basal insulin secretion (BIS), glucose-stimulated insulin secretion (GSIS), and insulin secretion index (ISI) of INS-1E cells that were cultured in media supplemented with or without 0.5 mM palmitate and treated with or without a PPARδ agonist (GW501516) or PPARδ siRNA. The expression of MafA, glucose transportor-2 (GLUT2), and insulin was found to be up-regulated in cells treated with GW501516. Finally, analysis of the level of JNK phosphorylation revealed that activated PPARδ could inhibit the activation of JNK and increase the expression of MafA. Accordingly, the insulin secretion dysfunction in lipotoxic INS-1E cells was improved. Collectively, these results demonstrate that activation of PPARδ improves insulin secretion impaired by palmitate and plays a role in the JNK-MafA-GLUT2 pathway.

A mechanistic rationale for MEK inhibitor therapy in myeloma based on blockade of MAF oncogene expression.

Modulating aberrant transcription of oncogenes is a relatively unexplored opportunity in cancer therapeutics. In approximately 10% of multiple myelomas, the initiating oncogenic event is translocation of musculoaponeurotic fibrosarcoma oncogene homolog (MAF), a transcriptional activator of key target genes, including cyclinD2. Our prior work showed that MAF is up-regulated in an additional 30% of multiple myeloma cases. The present study describes a common mechanism inducing MAF transcription in both instances. The second mode of MAF transcription occurred in myelomas with multiple myeloma SET domain (MMSET) translocation. MMSET knockdown decreased MAF transcription and cell viability. A small-molecule screen found an inhibitor of mitogen-activated protein kinase kinase (MEK), which activates extracellular signal-regulated kinase (ERK)-MAP kinases, reduced MAF mRNA in cells representing MMSET or MAF subgroups. ERK activates transcription of FOS, part of the AP-1 transcription factor. By chromatin immunoprecipitation, FOS bound the MAF promoter, and MEK inhibition decreased this interaction. MEK inhibition selectively induced apoptosis in MAF-expressing myelomas, and FOS inactivation was similarly toxic. Reexpression of MAF rescued cells from death induced by MMSET depletion, MEK inhibition, or FOS inactivation. The data presented herein demonstrate that the MEK-ERK pathway regulates MAF transcription, providing molecular rationale for clinical evaluation of MEK inhibitors in MAF-expressing myeloma.

Effect of PTTG on endogenous gene expression in HEK 293 cells.

BACKGROUND: Pituitary tumor transforming gene (PTTG), also known as securin, is highly expressed in various tumors including pituitary, thyroid, colon, ovary, testis, lung, and breast. An overexpression of PTTG enhances cell proliferation, induces cellular transformation in vitro, and promotes tumor development in nude mice. PTTG also inhibits separation of sister chromatids leading to aneuploidy and genetic instability. A great amount of work has been undertaken to understand the biology of PTTG and its expression in various tumors. However, mechanisms by which PTTG mediates its tumorigenic function are not fully understood. To utilize this gene for cancer therapy, identification of the downstream signaling genes regulated by PTTG in mediation of its tumorigenic function is necessary. For this purpose, we expressed PTTG in human embryonic kidney (HEK293) cells that do not express PTTG and analyzed the downstream genes using microarray analysis. RESULTS: A total of 22,277 genes printed on an Affymetrix HG-U133A 2.0 GeneChip array were screened with labeled cRNA prepared from HEK293 cells infected with adenovirus vector expressing PTTG cDNA (AdPTTG cDNA) and compared with labeled cRNA prepared from HEK293 cells infected with control adenovirus (control Ad) or adenovirus vector expressing GFP (AdGFP). Out of 22,277 genes, 71 genes were down-regulated and 35 genes were up-regulated with an FDR corrected p-value of < or = 0.05 and a fold change of > or =2. Most of the altered genes identified are involved in the cell cycle and cell apoptosis; a few are involved in mRNA processing and nitrogen metabolism. Most of the up-regulated genes belong to the histone protein family. CONCLUSION: PTTG is a well-studied oncogene for its role in tumorigenesis. In addition to its importance in regulation of the cell cycle, this gene has also been recently shown to play a role in the induction of cell apoptosis. The microarray analysis in the present study demonstrated that PTTG may induce apoptosis by down-regulation of oncogenes such as v-Jun and v-maf and up-regulation of the histone family of genes.

Identification of new Nerve Growth Factor-responsive immediate-early genes.

Stimulation of the PC12 pheochromocytoma cell line with the prototypical neurotrophin Nerve Growth Factor (NGF) induces a cellular response of neuronal differentiation and is therefore a widely used model to gain molecular insight into this process. Classically, the transcriptional response to extracellular stimuli such as NGF is divided in genes that require no protein synthesis prior to their induction (immediate-early genes) and genes that do (delayed-response genes). Because an increasing number of studies have reported important roles for immediate-early genes (IEGs) in neuronal differentiation, the goal of the present study was to identify previously unrecognized NGF-responsive IEGs. Stimulation with NGF for 15, 30, 60 and 120 min resulted in a typical transient induction of many known NGF-responsive IEGs. To identify candidate new genes, we analyzed 27000 measured expression profiles and selected 10 genes for further study. Five genes, including Cbp/p300-interacting transactivator 2 (Cited2), Kruppel-like factor 4 (Klf4), v-Maf musculoaponeurotic fibrosarcoma oncogene family, protein F (Maff), Kruppel-like factor 10 (Klf10 or Tieg) and Activating transcription factor 3 (Atf3) were selected and positively validated by qPCR. NGF-induced activation of all five genes seems to be mediated by MAPK and PI3K-mediated pathways. Additionally, we tested translation-independent induction and showed that NGF induced upregulation of these genes in both the subclonal Neuroscreen-1 PC12 and parental PC12 cell line. These 5 transcription factors have not been previously reported as NGF-responsive IEGs, however have previously been reported as important regulators of cell differentiation and proliferation in different systems. These observations may therefore provide important new information on the molecular mechanisms underlying NGF-induced differentiation.