Diosmetin Ameliorates Nonalcoholic Steatohepatitis through Modulating Lipogenesis and Inflammatory Response in a STAT1/CXCL10-Dependent Manner.

Nonalcoholic steatohepatitis (NASH) is an inflammatory lipotoxic disorder characterized by lipid accumulation and inflammation. Diosmetin (Dios), a flavonoid, has an active effect against nonalcoholic fatty liver disease, whereas its effect on NASH remains elusive. To investigate the effects of Dios on lipogenesis and inflammatory response and explore the molecular mechanisms of Dios on NASH, mice induced by high-fat diet (HFD), HepG2 cells stimulated by palmitic acid (PA), transcriptome sequencing, and molecular biological experiments were used. We show, by pathological analysis (HE, Oli Red O, and Masson staining) and biochemical parameters (TC, TG, LDL-C, ALT, and AST), Dios alleviated liver lipid accumulation and inflammatory injury. According to liver RNA-Seq analysis, CXCL10 and STAT1 were assumed to be the key target genes of Dios on NASH. Significantly, Dios regulated STAT1/CXCL10 signal pathway and further attenuated NASH via regulating the expression of LXRα/ß, SREBP-1c, CHREBP, and NF-κB. In conclusion, Dios is proposed to alleviate NASH through suppression of lipogenesis and inflammatory response via a STAT1/CXCL10-dependent pathway.

Protein arginine methyltransferase 5 promotes cholesterol biosynthesis-mediated Th17 responses and autoimmunity.

Protein arginine methyltransferase 5 (PRMT5) catalyzes symmetric dimethylation (SDM) of arginine, a posttranslational modification involved in oncogenesis and embryonic development. However, the role and mechanisms by which PRMT5 modulates Th cell polarization and autoimmune disease have not yet been elucidated. Here, we found that PRMT5 promoted SREBP1 SDM and the induction of cholesterol biosynthetic pathway enzymes that produce retinoid-related orphan receptor (ROR) agonists that activate RORγt. Specific loss of PRMT5 in the CD4+ Th cell compartment suppressed Th17 differentiation and protected mice from developing experimental autoimmune encephalomyelitis (EAE). We also found that PRMT5 controlled thymic and peripheral homeostasis in the CD4+ Th cell life cycle and invariant NK (iNK) T cell development and CD8+ T cell maintenance. This work demonstrates that PRMT5 expression in recently activated T cells is necessary for the cholesterol biosynthesis metabolic gene expression program that generates RORγt agonistic activity and promotes Th17 differentiation and EAE. These results point to Th PRMT5 and its downstream cholesterol biosynthesis pathway as promising therapeutic targets in Th17-mediated diseases.

Transcriptomic Profiling Provides Insights into Inbreeding Depression in Yesso Scallop Patinopecten yessoensis.

Inbreeding often causes a decline in biological fitness, known as inbreeding depression. In genetics study, inbreeding coefficient f gives the proportion by which the heterozygosity of an individual is reduced by inbreeding. With the development of high-throughput sequencing, researchers were able to perform deep approaches to investigate which genes are affected by inbreeding and reveal some molecular underpinnings of inbreeding depression. As one commercially important species, Yesso scallop Patinopecten yessoensis confront the same dilemma of inbreeding depression. To examine how inbreeding affects gene expression, we compared the transcriptome of two experimentally selfing families with inbreeding coefficient f reached 0.5 as well as one natural population (f ≈ 0) of P. yessoensis. A total of 24 RNA-Seq libraries were constructed using scallop adductor muscle, and eventually 676.56 M (96.85%) HQ reads were acquired. Based on differential gene analysis, we were able to identify nine common differentially expressed genes (DEGs) across the top-ranked 30 DEGs in both selfing families in comparation with the natural population. Remarkable, through weighted gene co-expression network analysis (WGCNA), five common DEGs were found enriched in the most significant inbreeding related functional module M14 (FDR = 1.64E-156), including SREBP1, G3BP2, SBK1, KIAA1161, and AATs-Glupro. These five genes showed significantly higher expression in self-bred progeny. Suggested by the genetic functional analysis, up-regulated SREBP1, G3BP2, and KIAA1161 may suggest a perturbing lipid metabolism, a severe inframammary reaction or immune response, and a stress-responsive behavior. Besides, the significant higher SBK1 and AATs-Glupro may reflect the abnormal cellular physiological situation. Together, these genetic aberrant transcriptomic performances may contribute to inbreeding depression in P. yessoensis, deteriorating the stress tolerance and survival phenotype in self-bred progeny. Our results would lay a foundation for further comprehensive understanding of bivalve inbreeding depression, which may potentially benefit the genetic breeding for scallop aquaculture.

Ginsenoside Rh2 reverses cyclophosphamide-induced immune deficiency by regulating fatty acid metabolism.

Ginsenoside Rh2 (G-Rh2) has well-established potent antitumor activity; yet, the effects of G-Rh2 on immune and metabolism regulation in cancer treatment, especially non-small cell lung cancer (NSCLC) remain unclear. We showed that G-Rh2 had a synergistic antitumor effect with cyclophosphamide (CY) on mice with NSCLC, and improved the immune deficiency caused by CY. Consistently, G-Rh2 exhibited no inhibitory effect on tumor growth of T cells-deficient nude mice. Furthermore, G-Rh2 treatment triggered the oxidative decomposition of fatty acid (FA), suppressed FA synthesis, increased ketone level, and decreased glucocorticoid (CORT) secretion. G-Rh2 significantly down-regulated the expression of fatty acid synthase (FASN). Of note, in liver-specific FASN knockout mice G-Rh2 failed to show the same immune enhancement effects. Further mechanistic exploration revealed that G-Rh2 suppressed the expression and nuclear translocation of sterol regulatory element binding protein 1 (SREBP-1), and disturbed the SREBP-1-FASN interaction in vitro.

Sterol regulatory element binding protein (SREBP) -1 mediates oxidized low-density lipoprotein (oxLDL) induced macrophage foam cell formation through NLRP3 inflammasome activation.

Macrophage foam cell formation (FCF) has long been known to play a critical role during atherosclerotic plaque development. In the presence of atherogenic molecules such as oxidized low-density lipoprotein (oxLDL) macrophages accumulate massive amounts of lipid through uptake. However, in the presence of oxLDL mechanism of dysregulated lipid homeostasis in the macrophages remains largely unknown. Herein we have investigated the role of Sterol regulatory element binding protein (SREBP)-1 in oxLDL-induced inflammation and altered lipid homeostasis in macrophages. The U937 monocytes and monocyte-derived macrophages (MDMs) were stimulated with different doses of oxLDL. MTT assay to study the effect of oxLDL on cell viability, Oil-Red-O (ORO) staining to observe cytosolic lipid accumulation, semi-quantitative PCR and Western blotting to analyze mRNA and protein expressions, respectively, and spectrophotometric assay to measure the lipid synthesizing enzyme's activity were performed. Our results indicate that oxLDL increased proliferation in monocytes and decreased the viability in MDMs in a time- and dose-dependent manner. The oxLDL (100 µg/ml) enhanced lipid accumulation via increased expressions of SREBP-1 and its downstream proteins such as fatty acid synthase (FAS) and 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR) at both RNA and protein levels in monocytes as well as in MDMs. Inhibiting SREBP-1 by a synthetic inhibitor prevented excessive lipid accumulation by downregulating the expression of its downstream proteins. Further, oxLDL increased reactive oxygen species (ROS) levels, NLRP3 inflammasome activation and active interleukin 1ß (IL-1ß) release in both the cell types. The oxLDL-induced NLRP3 could be responsible for SREBP-1 and downstream proteins overexpression as siRNA silencing of NLRP3 decreased SERBP-1 levels. In summary, we have demonstrated that SREBP-1 could be a key player in oxLDL-induced excessive lipid accumulation leading to macrophage FCF via ROS-mediated NLRP3/IL-1ß/SREBP-1 pathway.

Inhibition of P2X7R-NLRP3 Inflammasome Activation by Pleurotus citrinopileatus: A Possible Protective Role in Alcoholic Hepatosteatosis.

Pleurotus citrinopileatus (golden oyster mushroom) is a widely used edible mushroom. We investigated the inhibitory effect of P. citrinopileatus aqueous extract against alcoholic steatohepatitis and its underlying mechanism. Acute and chronic ethanol-feeding murine models were established by intragastrically administering ethanol or feeding an ethanol-containing Lieber-DeCarli liquid diet to male C57BL/6 mice. In both models, P. citrinopileatus decreased serum alanine aminotransferase (ALT), aspartate transaminase (AST), triglyceride (TG), and hepatic TG levels. Hematoxylin and eosin (HE) and Oil Red O staining confirmed that P. citrinopileatus ameliorated both acute and chronic alcoholic hepatosteatosis, characterized by regulation of lipid-metabolism-related proteins, including sirtuin 1 (SIRT1), AMP-activated kinase (AMPK), and sterol regulatory element-binding protein (SREBP1). P. citrinopileatus reversed inflammatory response via modulating purinergic receptor P2X ligand-gated ion channel 7 (P2X7R)-NOD-like receptor pyrin domain 3 (NLRP3) inflammasome. P. citrinopileatus restored the expressions of those proteins to a normal level. In addition, HepG2 cells were incubated with P. citrinopileatus prior to ethanol stimulation. P. citrinopileatus reduced ethanol exposure-induced lipid deposition. Concomitantly, P. citrinopileatus increased AMPK and SIRT1 expressions, which were reduced by ethanol treatment. P. citrinopileatus ameliorated alcoholic hepatic steatosis and accompanied inflammatory response via regulating SIRT1-AMPK and P2X7R-NLRP3 inflammasome activation, highlighting a promising strategy and utility of P. citrinopileatus for alcoholic steatohepatitis as dietary health supplements.

Lactoferrin attenuates high-fat diet-induced hepatic steatosis and lipid metabolic dysfunctions by suppressing hepatic lipogenesis and down-regulating inflammation in C57BL/6J mice.

Lactoferrin was reported to exert modulatory effects on lipid metabolism, but the regulatory mechanisms remain unclear. The present study investigated the beneficial effects of lactoferrin and their underlying mechanisms in high-fat diet-induced obese C57BL/6J mice. Oral administration of lactoferrin at 100 mg per body weight for 15 weeks significantly reduced weight gain, visceral adiposity, and serum glucose, leptin, and lipid levels in high-fat diet-induced obese mice. Hepatic steatosis in the obese mice was significantly improved. Expression of adipogenic and inflammation-related genes and proteins (SREBP-1c, FAS, MCP-1, leptin) was suppressed in the liver and epididymal adipose tissue of the obese mice. The present findings demonstrate that lactoferrin positively regulated lipid metabolism and improved hepatic steatosis in obese mice. The mechanisms of action for these effects may be attributed to suppression of lipogenic gene expression and amelioration of inflammation in the liver and epididymal adipose tissue.

Gut Microbiota-Derived Tryptophan Metabolites Modulate Inflammatory Response in Hepatocytes and Macrophages.

The gut microbiota plays a significant role in the progression of fatty liver disease; however, the mediators and their mechanisms remain to be elucidated. Comparing metabolite profile differences between germ-free and conventionally raised mice against differences between mice fed a low- and high-fat diet (HFD), we identified tryptamine and indole-3-acetate (I3A) as metabolites that depend on the microbiota and are depleted under a HFD. Both metabolites reduced fatty-acid- and LPS-stimulated production of pro-inflammatory cytokines in macrophages and inhibited the migration of cells toward a chemokine, with I3A exhibiting greater potency. In hepatocytes, I3A attenuated inflammatory responses under lipid loading and reduced the expression of fatty acid synthase and sterol regulatory element-binding protein-1c. These effects were abrogated in the presence of an aryl-hydrocarbon receptor (AhR) antagonist, indicating that the effects are AhR dependent. Our results suggest that gut microbiota could influence inflammatory responses in the liver through metabolites engaging host receptors.

Effect of short-term liver X receptor activation on epidermal barrier features in mild to moderate atopic dermatitis: A randomized controlled trial.

BACKGROUND: Liver X receptors (LXRs) are involved in maintaining epidermal barrier and suppressing inflammatory responses in model systems. The LXR agonist VTP-38543 showed promising results in improving barrier function and inflammatory responses in model systems. OBJECTIVE: To assess the safety, tolerability, cellular and molecular changes, and clinical efficacy of the topical VTP-38543 in adults with mild to moderate atopic dermatitis (AD). METHODS: A total of 104 ambulatory patients with mild to moderate AD were enrolled in this randomized, double-blind, vehicle-controlled trial between December 2015 and September 2016. VTP-38543 cream in 3 concentrations (0.05%, 0.15%, and 1.0%) or placebo was applied twice daily for 28 days. Pretreatment and posttreatment skin biopsy specimens were obtained from a subset of 33 patients. Changes in SCORing of Atopic Dermatitis, Eczema Area and Severity Index, Investigator's Global Assessment, and tissue biomarkers (by real-time polymerase chain reaction and immunostaining) were evaluated. RESULTS: Topical VTP-38543 was safe and well tolerated. VTP-38543 significantly increased messenger RNA (mRNA) expression of epidermal barrier differentiation (loricrin and filaggrin, P = .02) and lipid (adenosine triphosphate-binding cassette subfamily G member 1 and sterol regulatory element binding protein 1c, P < .01) measures and reduced epidermal hyperplasia markers (thickness, keratin 16 mRNA). VTP-38543 nonsignificantly suppressed cellular infiltrates and down-regulated mRNA expression of several TH17/TH22-related (phosphatidylinositol 3, S100 calcium-binding protein A12) and innate immunity (interleukin 6) markers. CONCLUSION: Topical VTP-38543 is safe and well tolerated. Its application led to improvement in barrier differentiation and lipids. Longer-term studies are needed to clarify whether a barrier-based approach can induce meaningful suppression of immune abnormalities. TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT02655679.

Leptin reduces microRNA-122 level in hepatic stellate cells in vitro and in vivo.

Obese patients, often accompanied by hyperleptinemia, are more likely to develop liver fibrosis. Leptin, an adipocyte-derived hormone, augments inflammatory in liver and promotes hepatic stellate cell (HSC) activation (a key step for liver fibrogenesis) and liver fibrosis. microRNA-122 (miR-122) is the most abundant liver-specific miRNA and can attenuate liver fibrosis. This study examined the effect of leptin on miR-122 level in HSCs in vivo and in vitro. Results demonstrated that leptin reduced the levels of both miR-122 (mature miR-122) and primary miR-122 (pri-miR-122). The effects of leptin on the levels of miR-122 and pri-miR-122 were through at least hedgehog pathway. Leptin-induced decrease in sterol regulatory element-binding protein-1c (SREBP-1c) has been shown to contribute to leptin-induced HSC activation. We revealed a mutual promotional effect between SREBP-1c and miR-122. Further experiments indicated that miR-122 inhibited leptin-induced liver fibrosis in leptin-deficient mouse model. These data have potential implications for clarifying the mechanisms of hepatic fibrogenesis associated with elevated leptin level in human such as obese patients.

Whole blood transcriptome analysis reveals potential competition in metabolic pathways between negative energy balance and response to inflammatory challenge.

Negative Energy Balance (NEB) is considered to increase susceptibility to mastitis. The objective of this study was to improve our understanding of the underlying mechanisms by comparing transcriptomic profiles following NEB and a concomitant mammary inflammation. Accordingly, we performed RNA-seq analysis of blood cells in energy-restricted ewes and control-diet ewes at four different time points before and after intra mammary challenge with phlogogenic ligands. Blood leucocytes responded to NEB by shutting down lipid-generating processes, including cholesterol and fatty acid synthesis, probably under transcriptional control of SREBF 1. Furthermore, fatty acid oxidation was activated and glucose oxidation and transport inhibited in response to energy restriction. Among the differentially expressed genes (DEGs) in response to energy restriction, 64 genes were also differential in response to the inflammatory challenge. Opposite response included the activation of cholesterol and fatty acid synthesis during the inflammatory challenge. Moreover, activation of glucose oxidation and transport coupled with the increase of plasma glucose concentration in response to the inflammatory stimuli suggested a preferential utilization of glucose as the energy source during this stress. Leucocyte metabolism therefore undergoes strong metabolic changes during an inflammatory challenge, which could be in competition with those induced by energy restriction.

Artemisia annua Leaf Extract Attenuates Hepatic Steatosis and Inflammation in High-Fat Diet-Fed Mice.

Artemisia annua L. (AA) is a well-known source of the antimalarial drug artemisinin. AA also has an antibacterial and antioxidant activity. However, the effect of AA extract on hepatic steatosis induced by obesity is unclear. We investigated whether AA extract prevents obesity-induced insulin resistance and hepatic steatosis in high-fat diet (HFD)-fed mice. Mice were randomly divided into groups that received a normal chow diet or HFD with or without AA for 12 weeks. We found that AA extract reduced insulin resistance and hepatic steatosis in HFD-fed mice. Western blot analysis showed that HFD-induced expression of nuclear sterol regulatory element-binding protein 1 and carbohydrate-responsive element-binding protein in the livers was decreased by AA extract. In particular, dietary administration of AA extract decreased hepatic high-mobility group box 1 and cyclooxygenase-2 expression in HFD-fed mice. AA extract also attenuated HFD-induced collagen deposition and fibrosis-related transforming growth factor-ß1 and connective tissue growth factor. These data indicate that dietary AA extract has beneficial effects on hepatic steatosis and inflammation in HFD-fed mice.

Age-related sensitivity to endotoxin-induced liver inflammation: Implication of inflammasome/IL-1ß for steatohepatitis.

Aging is associated with increased vulnerability to inflammatory challenge. However, the effects of altered inflammatory response on the metabolic status of tissues or organs are not well documented. In this study, we present evidence demonstrating that lipopolysaccharide (LPS)-induced upregulation of the inflammasome/IL-1ß pathway is accompanied with an increased inflammatory response and abnormal lipid accumulation in livers of aged rats. To monitor the effects of aging on LPS-induced inflammation, we administered LPS (2 mg kg(-1) ) to young (6-month old) and aged (24-month old) rats and found abnormal lipid metabolism in only aged rats with increased lipid accumulation in the liver. This lipid accumulation in the liver was due to the dysregulation of PPARα and SREBP1c. We also observed severe liver inflammation in aged rats as indicated by increased ALT levels in serum and increased Kupffer cells in the liver. Importantly, among many inflammation-associated factors, the aged rat liver showed chronically increased IL-1ß production. Increased levels of IL-1ß were caused by the upregulation of caspase-1 activity and inflammasome activation. In vitro studies with HepG2 cells demonstrated that treatment with IL-1ß significantly induced lipid accumulation in hepatocytes through the regulation of PPARα and SREBP1c. In summary, we demonstrated that LPS-induced liver inflammation and lipid accumulation were associated with a chronically overactive inflammasome/IL-1ß pathway in aged rat livers. Based on the present findings, we propose a mechanism of aging-associated progression of steatohepatitis induced by endotoxin, delineating a pathogenic role of the inflammasome/IL-1ß pathway involved in lipid accumulation in the liver.

Pancreastatin-dependent inflammatory signaling mediates obesity-induced insulin resistance.

Chromogranin A knockout (Chga-KO) mice exhibit enhanced insulin sensitivity despite obesity. Here, we probed the role of the chromogranin A-derived peptide pancreastatin (PST: CHGA(273-301)) by investigating the effect of diet-induced obesity (DIO) on insulin sensitivity of these mice. We found that on a high-fat diet (HFD), Chga-KO mice (KO-DIO) remain more insulin sensitive than wild-type DIO (WT-DIO) mice. Concomitant with this phenotype is enhanced Akt and AMPK signaling in muscle and white adipose tissue (WAT) as well as increased FoxO1 phosphorylation and expression of mature Srebp-1c in liver and downregulation of the hepatic gluconeogenic genes, Pepck and G6pase. KO-DIO mice also exhibited downregulation of cytokines and proinflammatory genes and upregulation of anti-inflammatory genes in WAT, and peritoneal macrophages from KO mice displayed similarly reduced proinflammatory gene expression. The insulin-sensitive, anti-inflammatory phenotype of KO-DIO mice is masked by supplementing PST. Conversely, a PST variant peptide PSTv1 (PST-NΔ3: CHGA(276-301)), lacking PST activity, simulated the KO phenotype by sensitizing WT-DIO mice to insulin. In summary, the reduced inflammation due to PST deficiency prevented the development of insulin resistance in KO-DIO mice. Thus, obesity manifests insulin resistance only in the presence of PST, and in its absence obesity is dissociated from insulin resistance.

Evidence of contribution of iPLA2ß-mediated events during islet ß-cell apoptosis due to proinflammatory cytokines suggests a role for iPLA2ß in T1D development.

Type 1 diabetes (T1D) results from autoimmune destruction of islet ß-cells, but the underlying mechanisms that contribute to this process are incompletely understood, especially the role of lipid signals generated by ß-cells. Proinflammatory cytokines induce ER stress in ß-cells and we previously found that the Ca(2+)-independent phospholipase A2ß (iPLA2ß) participates in ER stress-induced ß-cell apoptosis. In view of reports of elevated iPLA2ß in T1D, we examined if iPLA2ß participates in cytokine-mediated islet ß-cell apoptosis. We find that the proinflammatory cytokine combination IL-1ß+IFNγ, induces: a) ER stress, mSREBP-1, and iPLA2ß, b) lysophosphatidylcholine (LPC) generation, c) neutral sphingomyelinase-2 (NSMase2), d) ceramide accumulation, e) mitochondrial membrane decompensation, f) caspase-3 activation, and g) ß-cell apoptosis. The presence of a sterol regulatory element in the iPLA2ß gene raises the possibility that activation of SREBP-1 after proinflammatory cytokine exposure contributes to iPLA2ß induction. The IL-1ß+IFNγ-induced outcomes (b-g) are all inhibited by iPLA2ß inactivation, suggesting that iPLA2ß-derived lipid signals contribute to consequential islet ß-cell death. Consistent with this possibility, ER stress and ß-cell apoptosis induced by proinflammatory cytokines are exacerbated in islets from RIP-iPLA2ß-Tg mice and blunted in islets from iPLA2ß-KO mice. These observations suggest that iPLA2ß-mediated events participate in amplifying ß-cell apoptosis due to proinflammatory cytokines and also that iPLA2ß activation may have a reciprocal impact on ER stress development. They raise the possibility that iPLA2ß inhibition, leading to ameliorations in ER stress, apoptosis, and immune responses resulting from LPC-stimulated immune cell chemotaxis, may be beneficial in preserving ß-cell mass and delaying/preventing T1D evolution.

[Chicken SREBP1 gene antiserums preparation and its tissue expression characterization].

AIM: The aim of this study was to prepare the antiserums against chicken sterol regulatory element binding protein1 (SREBP1), and to analyze the expression of SREBP1 in chicken tissues. METHODS: The nuclear import sequence of SREBP1 was analyzed using the DNAStar programs to predict its major antigen epitopes, the fragment coding for SREBP1 major antigen epitopes (832-1 302 bp) was amplified by RT-PCR and inserted into pGEX-4T-1 to construct the expression vector pGEX-4T/SREBP1. The recombinant GST/SREBP1 was expressed in E.coli with IPTG induction and purified by Glutathione Sepharose 4B affinity chromatography. The purified recombinant GST/SREBP1 was used as immunogen in rabbits, and the titer and specificity of SREBP1 antiserums were detected by ELISA and Western blot. The tissue expression of chicken SREBP1 was analyzed with the antiserums. RESULTS: The titer of the antiserum determined by ELISA was 1:102 400, and the antiserums exhibited a high specificity through Western blot. Tissue expression analysis showed that the expression of chicken SREBP1 was much higher in abdominal adipose tissue and heart, but lower in liver, muscle stomach, duodenum, spleen, kidney, and no signal was detected in breast and leg muscle. The SREBP1 expression characteristics between fat and lean broiler lines was also detected, and the result showed that SREBP1 expressed significantly higher in the abdominal adipose tissue of fat line than that in lean line (P<0.05). CONCLUSION: The antiserum against chicken SREBP1 prepared in this study had high titer and specificity. SREBP1 was highly expressed in abdominal adipose tissue of broilers. Compared to lean broiler line, SREBP1 was expressed higher in the fat line.

Growth factor-induced phosphorylation of sterol regulatory element-binding proteins inhibits sumoylation, thereby stimulating the expression of their target genes, low density lipoprotein uptake, and lipid synthesis.

The destiny and activity of sterol regulatory element-binding proteins (SREBPs) in the nucleus are regulated by modification with ubiquitin, small ubiquitin-like modifier (SUMO), or phosphorus. ERK-dependent phosphorylation causes an increase in their transcriptional activity, whereas SUMO modification halts it. We hypothesized a causal linkage between phosphorylation and sumoylation because their sites are very closely located in SREBP-1 and -2 molecules. When Ser(455), a phosphorylation site in SREBP-2, was substituted with Ala, this SREBP-2 mutant was more efficiently modified by SUMO-1. On the other hand, substitution of Asp inhibited SUMO conjugation, mimicking phosphoserine. When cells were cultured with insulin-like growth factor-1, sumoylation of SREBP-2 was decreased with an increase in its phosphorylation, but SREBP-2(S455A) was continuously sumoylated. An ERK cascade inhibitor, U0126, inversely augmented SUMO modification of SREBP-2. Insulin-like growth factor-1 treatment stimulated the expression of SREBP target genes such as the low density lipoprotein (LDL) receptor, squalene synthase, and hydroxymethylglutaryl-CoA synthase genes. These results indicate that growth factor-induced phosphorylation of SREBP-2 inhibits sumoylation, thereby facilitating SREBP transcriptional activity. Glutathione S-transferase pulldown assays revealed that wild-type SREBP-2, but not a mutant lacking Lys(464), interacts with HDAC3 preferentially among the histone deacetylase family members. HDAC3 small interfering RNA induced gene expression of the LDL receptor and thereby augmented fluorescently labeled LDL uptake in HepG2 cells. In summary, growth factors inhibit sumoylation of SREBPs through their phosphorylation, thus avoiding the recruitment of an HDAC3 corepressor complex and stimulating the lipid uptake and synthesis required for cell growth.

Tom1l2 hypomorphic mice exhibit increased incidence of infections and tumors and abnormal immunologic response.

Studies have shown that the TOM1 family of proteins, including TOM1 and TOM1L1, are actively involved in endosomal trafficking and function in the immune response. However, much less is known about the function of TOM1L2. To understand the biological importance of TOM1L2 and the potential significance of its cellular role, we created and evaluated Tom1l2 gene-trapped mice with reduced Tom1l2 expression. Mice hypomorphic for Tom1l2 exhibited numerous infections and tumors compared to wild-type littermates. Associated with this increased risk for infection and tumor formation, apparently healthy Tom1l2 hypomorphs also had splenomegaly, elevated B- and T-cell counts, and an impaired humoral response, although at a reduced penetrance. Furthermore, cellular localization studies showed that a Tom1l2-GFP fusion protein colocalizes with Golgi compartments, supporting the role of Tom1l2 in cellular trafficking, while molecular modeling and bioinformatic analysis of Tom1l2 illustrated a structural basis for a functional role in trafficking. These results indicate a role for Tom1l2 in the immune response and possibly in tumor suppression.