cAMP-stimulated transcription of DGKθ requires steroidogenic factor 1 and sterol regulatory element binding protein 1

Abstract

( 4 ). DGK -null mice exhibit several neural abnormalities, including a higher resistance of electroconvulsive shock ( 5 ) and increased cyclooxygenase 2 and tyrosine hydroxylase expression ( 6 ), suggesting a role for DGK in regulating synaptic activity. Mice lacking DGK ␣ ( 7 ) or DGK ( 8 ) exhibit enhanced T cell function and demonstrate a role for these kinases in controlling DAG metabolism during the immune response. DGK isoforms have been implicated in various other cellular processes including inhibition of Rap1 signaling ( 9 ) and retinoblastoma-mediated cell cycle control ( 10 ). DGK is activated by nerve growth factor in PC12 cells ( 11 ) and thrombin in IIC9 fi broblasts ( 12, 13 ), whereas DGK promotes myogenesis in C2C12 cells ( 14 ) and DGK ␥ plays a role in regulating the cell cycle in CHO-K cells ( 15 ).To date, 10 mammalian DGKs have been identifi ed that are divided into fi ve groups based on functional domains ( 16,17 ). However, all isoforms contain cysteine-rich zinc fi nger-like structures, a conserved catalytic region ( 18-21 ). DGK , the sole member of group V, is comprised of three cysteine-rich domains (CRDs), a proline/glycine-rich domain at its N terminus, and a pleckstrin homology (PH) with an overlapping Ras-binding domain ( 22 ). While the functions of many of the other domains in DGK are unclear, the catalytic activity requires all domains of the enzyme ( 23 ). It has been postulated that the CRDs of the enzyme are required both for correct folding of the protein and for substrate presentation ( 23 ). Mutation of the CRD of DGK diminishes DAG-induced translocation of the enzyme to the plasma membrane ( 24 ); whereas the interaction between DGK and the nuclear receptor steroidogenic factor 1 (SF1) requires the PH domain ( 25 ). Abstract Diacylglycerol kinase (DGK) is

Lipin-1 Regulates Autophagy Clearance and Intersects with Statin Drug Effects in Skeletal Muscle

“…Triacylglycerol, free cholesterol, cholesteryl ester, and free fatty acid levels were determined using reagents from Wako (Richmond, VA). Phosphatidic acid (PA) in primary mouse embryonic fibroblasts (MEFs) or C2C12 myocytes was determined with a Total PA kit (Cayman, Ann Arbor, MI) (Zhang et al, 2012), and diacylgycerol (DAG) was assessed with a mouse DAG ELISA kit (MyBiosource Inc., San Diego, CA) (Cai and Sewer, 2013). …”

Diacylglycerol kinase θ couples farnesoid X receptor-dependent bile acid signalling to Akt activation and glucose homoeostasis in hepatocytes

“…(Manassas, VA, U.S.A.) and cultured in Eagle’s minimum essential medium (Mediatech) supplemented with 10 % FBS (Mediatech) and antibiotics (1× penicillin-streptomycin/amphotericin B; Lonza). DGK θ -knockdown cells were generated as described previously [38]. The following lots of primary human hepatocyte cultures were purchased from XenoTech: lot 1147 (35-year-old male), lot 1152 (69-year-old female), lot 1155 (43-year-old female) and lot 1157 (59-year-old female).…”

“…HepG2 cells were subcultured on to 24-well plates and transfected with 20 ng of pGL3-DGK θ [38], 0.5 ng of pRL-CMV (Promega) and 25 ng of pcDNA3.1-FXR (provided by Dr David D. Moore, Baylor College of Medicine, Houston, TX, U.S.A.) using Genejuice ® (Novagen). At 24 h after transfection, the cells were treated with 1 nM GW4064 for 24 h and the transcriptional activity of the DGK θ reporter gene was measured using a dual luciferase assay kit (Promega).…”

“…The purified chromatin solutions were immunoprecipitated using 5 μ g of anti-(acetyl-histone H3) (17-615, Millipore), anti-FXR (Santa Cruz Biotechnology) or anti-IgG (Millipore) antibodies and Protein A/G PLUS (Santa Cruz Biotechnology). Real-time PCR was carried out to amplify region − 1000 to − 700 bp of the DGK θ promoter [38] using the following primer sets: forward 5′-CAGAGTCCACAGCCCCCAGCCCCTTTCAGG-3′ and reverse 5′-CTGCCTCGTGCGCGCCACGGGTCTTGTTCA-3′. Output (immunoprecipitated promoter region) was normalized to input DNA.…”

Disentangling biological signaling networks by dynamic coupling of signaling lipids to modifying enzymes

“…Diacylglycerol kinase theta (DGKθ) is another lipid signaling enzyme known to generate phosphatidic acid from diacylglycerol (DAG) (Raben and Tu-Sekine, 2008). DGKθ interacts with SF-1 (Li et al, 2007) and DGKθ activity stimulates SF-1 gene expression programs in living cells (Cai and Sewer, 2013). Again, since IPMK requires an inositol headgroup to catalyze phosphorylation (Blind et al, 2012; Resnick et al, 2005), when DAG occupies SF-1, SF-1 must be decoupled from direct IPMK signaling.…”