Akt is a central player in the signal transduction pathways activated in response to many growth factors, hormones, cytokines, and nutrients and is thought to control a myriad of cellular functions including proliferation and survival, autophagy, metabolism, angiogenesis, motility, and exocytosis. Moreover, dysregulated Akt activity is being implicated in the pathogenesis of a growing number of disorders, including cancer. Evidence accumulated over the past 15 years has highlighted the presence of active Akt in the nucleus, where it acts as a fundamental component of key signaling pathways. For example, nuclear Akt counteracts apoptosis through a block of caspase-activated DNase: deoxyribonuclease and inhibition of chromatin condensation, and is also involved in cell cycle progression control, cell differentiation, mRNA: messenger RNA export, DNA repair, and tumorigenesis. In this review, we shall summarize the most relevant findings about nuclear Akt and its functions.

A diacylglycerol kinase cDNA was isolated from a rat brain cDNA library. This cDNA encoded an 801-amino acid protein of 90,287 Da. This 90-kDa diacylglycerol kinase showed 58% identity in deduced amino acid sequence with a previously isolated rat 80-kDa diacylglycerol kinase. EF-hand motifs, cysteine-rich zinc-finger-like sequences, and putative ATP-binding sites were all conserved between the two kinase species. However, mRNA encoding the 90-kDa kinase was confined to restricted neuronal populations such as the caudate-putamen, the accumbens nucleus, and the olfactory tubercle. Further, the 90-kDa kinase was found to exhibit high phosphorylation activity for long-chain diacylglycerols and was mainly associated with the membrane fraction when the cDNA was transfected into COS-7 ceils.In the process ofcell signal transduction, diacylglycerol (DG) kinase is thought to be involved in the resynthesis of phosphatidylinositol by converting the second messenger DG to phosphatidic acid (1, 2). DG kinase is thus regarded as an attenuator ofthe activity ofprotein kinase C (PKC) (3, 4). The importance of DG kinase in neurologic functions is indicated by the observation that the activity of this enzyme is lacking in the Drosophila retinal degeneration mutant (rdgA) (5).In mammals, a soluble DG kinase has been purified from porcine brain cytosol (6), and cDNA encoding the porcine 80-kDa DG kinase has been revealed to specify EF-hand motifs, cysteine-rich zinc-fmger-like sequences, and putative ATP-binding sites (7). Furthermore in situ hybridization followed by gene cloning of the rat homologue has demonstrated that mRNA for the 80-kDa DG kinase in brain is unexpectedly confined to oligodendrocytes, suggesting a regulatory involvement in myelin metabolism (8). To understand the functional significance of DG kinase in the process of neuronal signal transduction via the phosphatidylinositol cycle, it is crucial to identify neuronal species of DG kinase. Here we report the molecular cloning and expression ofa DG kinase that localizes in neurons and appears to be associated with the membrane.* MATERIALS AND METHODS cDNA Cloning. A rat brain cDNA library was screened with a 32P-labeled 1.0-kb Xba I fragment of rat 80-kDa DG kinase cDNA (8) under low-stringency conditions: 30% formamide/5 x standard saline citrate (SSC)/1 x Denhardt's solution/50 mM sodium phosphate, pH 7.2, with heatdenatured salmon sperm DNA (250 ug/ml) at 42°C. A single clone (4.7 kb, pNDGK1) showing weak signals was partially sequenced and found to be homologous to rat 80-kDa DG kinase. By rescreening a rat brain cDNA library with pNDGK1 as a probe under high-stringency conditions (8), three positive clones of 5.2, 5.9, and 2.7 kb (pNDGK2-4, The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

Isolation and characterization of a human cDNA demonstrated a novel lipoprotein receptor designated apolipoprotein E receptor 2 (apoER2). The new receptor consists of five functional domains resembling the low density lipoprotein (LDL) and very low density lipoprotein (VLDL) receptors. LDL receptor deficient Chinese hamster ovary cells expressing human apoER2 bound apoE rich ␤-migrating VLDL with high affinity and internalized. LDL was bound with much lower affinity to these cells. The 4.5-and 8.5-kb mRNAs for the receptor were most highly expressed in human brain and placenta. In rabbit tissues, multiple species of the mRNA with 4, 4.5, 5.5, 8.5, and 11 kb were detected most intensely in brain and testis and, to a much lesser extent, in ovary, but were undetectable in other tissues. In rat adrenal pheochromocytoma PC12 cells, the receptor mRNA was induced by treatment of the cells with nerve growth factor. The receptor transcripts were detectable most intensely in the cerebellar cortex, choroid plexus, ependyma, hippocampus, olfactory bulb and, to a much lesser extent, in the cerebral cortex as revealed by in situ hybridization histochemistry. In the cerebellar cortex, the receptor transcripts were densely deposited in Purkinje cell somata.Receptor-mediated endocytosis of plasma lipoproteins plays an important role in the metabolism of cholesterol and triglyceride in the body. The low density lipoprotein (LDL) 1 receptor, one of the best characterized cell surface receptors, mediates cholesterol homeostasis in the body (1). The LDL receptor binds apolipoprotein B-100 containing LDL and apolipoprotein E (apoE)-containing lipoproteins, whereas the recently found very low density lipoprotein (VLDL) receptor binds only apoEcontaining lipoproteins (2, 3). Both the LDL receptor (4 -7) and VLDL receptor (2, 8 -14) consist of five functional domains: (i) an amino-terminal ligand binding domain composed of multiple cysteine-rich repeats; (ii) an epidermal growth factor (EGF) precursor homology domain, which mediates the acid-dependent dissociation of the ligands from the LDL receptor (15); (iii) an O-linked sugar domain; (iv) a transmembrane domain; and (v) a cytoplasmic domain with a coated pit targeting signal (16). Genetic deficiencies of the LDL receptor give rise to familial hypercholesterolemia, one of the most common genetic diseases in humans (17). Mutations in the chicken VLDL receptor gene lead to the failure to produce offspring (13, 18).Lipoprotein metabolism in the central nervous system (CNS) has been poorly understood, despite the importance of lipids in some specialized neural membranes, such as myelin. Most of lipids in the CNS are actively synthesized in the CNS itself and deposited in large amounts during the early phase of development (19, 20). The rate of cholesterol and fatty acid synthesis in the brain is high during the myelinating period and declines thereafter (19,20). Although most of lipids in the brain are believed to be synthesized within the brain itself, small amounts of cholesterol (21)...

The cDNA corresponding to a fourth species of diacylglycerol (DG) kinase (EC 2.7.1.107) was isolated from cDNA libraries of rat retina and brain. This cDNA encoded a 929-aa, 104-kDa polypeptide termed DGK-IV. DGK-IV was different from previously identified mammalian DG kinase species, DGK-I, DGK-ll, and DGK-IH, in that it contained no EF-hand motifs but did contain four ankyrin-like repeats at the carboxyl terminus. These structural features of DGK-IV closely resemble the recently cloned, eye-specific DG kinase of Drosophila that is encoded by the retinal degeneration A (rdgA) gene. However, DGK-IV was expressed primarily in the thymus and brain with relatively low expression in the eye and intestine. Furthermore, the primary structure of the DGK-IV included a nuclear targeting motif, and immunocytochemical analysis revealed DGK-IV to localize in the nucleus of COS-7 cells transfected with the epitope-tagged cDNA, suggesting an involvement of DGK-IV in intranuclear processes.Phosphoinositide (PI) turnover produces two second messengers, diacylglycerol (DG) and inositol trisphosphate, in response to external stimuli (1, 2). DG acts as an activator of several forms of protein kinase C (PKC), whereas inositol trisphosphate mobilizes calcium ions from the endoplasmic reticulum (3, 4). DG is converted to phosphatidic acid by DG kinase. Although this conversion is regarded as the first step of the recycling of PI (5), several reports have recently shown that phosphatidic acid may be involved in the regulation of DNA synthesis; the induction of c-myc, c-fos, and plateletderived growth factor; and cAMP formation (6, 7). Furthermore, phosphatidic acid-dependent protein phosphorylation occurs in soluble preparations from several organs such as brain, spleen, and testis (6,8). Therefore, DG kinase may play a role not only in the attenuation of DG, but also in the production of a possible second messenger, phosphatidic acid. The importance of DG kinase in neurologic functions is also indicated by the observation that the activity of this enzyme is lacking in a Drosophila retinal degeneration mutant (rdgA) (9).In this study, representing the fourth of a series of our studies, we isolated cDNA for a fourth DG kinase, DGK-IV, from cDNA libraries of rat retina and brain. DGK-IV was distinct from DGK-I, DGK-II and DGK-III in primary structure and expression localization, but closely resembled the recently cloned eye-specific DG kinase of Drosophila encoded by the rdgA gene (10). These results suggest that DGK-IV is a mammalian homologue of the eye-specific DGK-rdgA. Furthermore, immunocytochemical analysis revealed DGK-IV to localize in the nucleus of COS-7 cells transfected with the epitope-tagged cDNA, suggesting that DGK-IV may be involved in intranuclear processes.MATERIALS AND METHODS cDNA Cloning. A rat retinal cDNA library (6 x 105 clones) was screened with mixed cDNA probes encoding the putative catalytic regions of rat DGK-I, DGK-II, and DGK-III under low stringency conditions as described (11-13). One positive ...

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Background-Diacylglycerol is a lipid second messenger that accumulates in cardiomyocytes when stimulated by Gq␣ protein-coupled receptor (GPCR) agonists such as angiotensin II, phenylephrine, and others. Diacylglycerol functions as a potent activator of protein kinase C (PKC) and is catalyzed by diacylglycerol kinase (DGK) to form phosphatidic acid and inactivated. However, the functional roles of DGK have not been previously examined in the heart. We hypothesized that DGK might prevent GPCR agonist-induced activation of diacylglycerol downstream signaling cascades and subsequent cardiac hypertrophy. Methods and Results-To test this hypothesis, we generated transgenic (DGK-TG) mice with cardiac-specific overexpression of DGK. There were no differences in heart size and heart weight between DGK-TG and wild-type littermate mice. The left ventricular function was normal in DGK-TG mice. Continuous administration of subpressor doses of angiotensin II and phenylephrine caused PKC translocation, gene induction of atrial natriuretic factor, and subsequent cardiac hypertrophy in WT mice. However, in DGK-TG mice, neither translocation of PKC nor upregulation of atrial natriuretic factor gene expression was observed after angiotensin II and phenylephrine infusion. Furthermore, in DGK-TG mice, angiotensin II and phenylephrine failed to increase cross-sectional cardiomyocyte areas and heart to body weight ratios. Phenylephrine-induced increases in myocardial diacylglycerol levels were completely blocked in DGK-TG mouse hearts, suggesting that DGK regulated PKC activity by controlling cellular diacylglycerol levels. Conclusions-These results demonstrated the first evidence that DGK negatively regulated the hypertrophic signaling cascade and resultant cardiac hypertrophy in response to GPCR agonists without detectable adverse effects in in vivo hearts. (Circulation. 2006;113:60-66.)

Diacylglycerol kinase (DGK) attenuates levels of second messenger diacylglycerol in cells and produces another (putative) messenger, phosphatidic acid. We have previously purified a 110-kDa DGK from rat brain (Kato, M., and Takenawa, T. (1990) J. Biol. Chem. 265, 794 -800). Here we report the cDNA cloning from human brain and retina cDNA libraries. The cDNA encodes a novel DGK isotype, termed DGK, of 941 amino acids with an apparent molecular mass of 110 kDa. DGK contains a C-terminal putative catalytic domain, which is present in all eukaryotic DGKs. In contrast to other DGK isotypes, DGK contains three cysteine-rich domains instead of two. The third cysteine-rich domain is most homologous to the second one in other DGK isotypes. This particular sequence homology extends C-terminally beyond the typical cysteine/histidine core structure and is DGKspecific. DGK furthermore contains various domains for protein-protein interaction, such as a proline-and glycine-rich domain with a putative SH3 domain-binding site and a pleckstrin homology domain with an overlapping Ras-associating domain. DGK is expressed in the brain and, to a lesser extent, in the small intestine, duodenum, and liver. In situ hybridization of DGK mRNA in adult rat brain reveals high expression in the cerebellar cortex and hippocampus. DGK activity in COS cell lysates is optimal toward diacylglycerols containing an unsaturated fatty acid at the sn-2 position.