HMGA2 promotes adipogenesis by activating C/EBPβ-mediated expression of PPARγ

AbstractAdipogenesis is orchestrated by a highly ordered network of transcription factors including peroxisome-proliferator activated receptor-gamma (PPARγ) and CCAAT-enhancer binding protein (C/EBP) family proteins. High mobility group protein AT-hook 2 (HMGA2), an architectural transcription factor, has been reported to play an essential role in preadipocyte proliferation, and its overexpression has been implicated in obesity in mice and humans. However, the direct role of HMGA2 in regulating the gene expression program during adipogenesis is not known. Here, we demonstrate that HMGA2 is required for C/EBPβ-mediated expression of PPARγ, and thus promotes adipogenic differentiation. We observed a transient but marked increase of Hmga2 transcript at an early phase of differentiation of mouse 3T3-L1 preadipocytes. Importantly, Hmga2 knockdown greatly impaired adipocyte formation, while its overexpression promoted the formation of mature adipocytes. We found that HMGA2 colocalized with C/EBPβ in the nucleus and was required for the recruitment of C/EBPβ to its binding element at the Pparγ2 promoter. Accordingly, HMGA2 and C/EBPβ cooperatively enhanced the Pparγ2 promoter activity. Our results indicate that HMGA2 is an essential constituent of the adipogenic transcription factor network, and thus its function may be affected during the course of obesity

Activation of Sp1-mediated transcription by Rta of Epstein–Barr virus via an interaction with MCAF1

Rta is a transcription factor encoded by BRLF1 of the Epstein–Barr virus (EBV). This factor is expressed during the immediate-early stage of the lytic cycle to activate the genes required for EBV lytic development. Although transcription activation by Rta is frequently associated with the binding of Rta to the Rta-response element (RRE) in promoters, Rta sometimes activates promoters without an RRE. Here we show that Rta interacts with an Sp1-interacting protein, MBD1-containing chromatin-associated factor 1 (MCAF1). This interaction is critical to the formation of an Sp1–MCAF1–Rta complex at Sp1 sites. Therefore, following lytic induction and the expression of Rta, Rta increases Sp1-mediated transcription. The genes that are thus activated include p16, p21, SNRPN and BRLF1. However, the binding of Rta to RRE prevents the interaction between Rta and MCAF1; therefore, transcription activation by RRE depends only on Rta, and not on MCAF1 or Sp1. Furthermore, this study finds that MCAF1 promotes the expression of Rta and Zta from EBV, indicating that MCAF1 participates EBV lytic activation. Our study documents the critical role of Rta in regulating the transcription of the genes that are mediated by Sp1

Seasonal Variation of the Content of Major Nutritional Elements in Leaves of Muscat Bailey A

1.1964年6月15日および8月1日に,岡山県山陽町でMuscat Bailey Aの28園について採葉して葉分析をおこなった. N含量については6月15日に2.63%(100)であったものが8月1日には2,16%(82)となっていることは不当な栽培法によるものと思われる. Mg含量は6月15日に0.19%(100),8月1日には0.33%(174)であるから,本品種は6月15日に“早期潜在的苦土欠乏”に陥っているということができる. 2.8月29日から10月16日までに4回にわたり,葉の片側から,その下方と上方から合計4切片(1切片当り1cm2)を打ち抜いた. 10月29日現在無処理の半面と他の半面との間でN,P,K,CaおよびMgの含量については,Ca以外にはほとんど差が認められなかった. 3.8月29日,9月29日および10月29日の葉内N含量は2.11%(100),1.90(90)〔100〕および1.49(71)〔78〕であって,10月末までに葉中N化合物が樹体内に移行する量は多くはない. Kは9月下旬の多雨による溶脱のためか,9月29日に1.10%〔100〕となったが,10月29日には1.87%〔170〕となった. 9月29日から10月29日の間のMg葉量の増大率(66%)はCaのそれ(21%)より大である. 葉内P含量は9月15日から10月29日の間でほとんど差がない. 4.10月29日現在,クロロシス発現葉の右側半分および左側半分の脈間部のMg含量は0.34％および0.30％であったが,健全葉のそれらは0.40％および0.33％であった. Muscat Bailey AのMg欠乏症発現についての8月の葉中Mg含量の安全限界濃度は0.30％と推定された

Functional Analysis of MeCP2 Mutations Associated with Rett Syndrome Using Transient Expression Systems

レット症候群は生後半年から1歳半ころに発症する重度の精神発達遅滞を伴う疾患で女児の1万人から1万5千人に1人に発症する頻度の高い遺伝子疾患である。この疾患の原因遺伝子が最近MeCP2遺伝子であることが判明した。レット症候群の患者でみられる変異がMeCP2の本来の機能にどのような影響を及ぼすかを理解することは、レット症候群の病態を解明する上での手がかりになる。MeCP2は2つの機能ドメインを持ち、一つはメチル化CpGに結合するメチル化結合ドメイン(MBD)で、もう一つはヒストン脱アセチル化酵素をリクルートするSin3Aと結合する転写抑制ドメイン(TRD)である。報告されている変異の中でミスセンス変異の多くは、この二つのドメイン内でみられ、特にMBD内での変異の割合は多い。MBD内のミスセンス変異のMeCP2機能への影響を把握するため、培養細胞を用いた遺伝子導入発現系を開発して解析を行った

Functional Characterisation of MeCP2 Mutatiions Found in Male Patients with X Linked Mental Retardation

MeCP2の遺伝子変異は、Rett症候群以外の疾患の患者でも見つかり、X染色体性の精神発達遅滞を伴う男性患者においても報告された。これらの患者ではMBD内の変異として137番目のGluからGlyと140番目AlaからValのアミノ酸変異が確認された。これらの変異に関して、開発した二つの機能解析系を用いて解析を行ったところ、140番目の変異では、メチル化DNAに対しての転写抑制活性は完全に維持されており、137番目の変異ではわずかに転写抑制活性の低下がみられる程度であった。また、マウス細胞のヘテロクロマチン親和性についても140番と137番目の変異は共に明らかな点状の像を示し、親和性は維持されていた。これらの遺伝性の精神発達遅滞を伴う男性患者でのMeCP2の変異は、その機能への影響がレット症候群の場合と比較して軽度であるため、Rett症候群とは異なる病態を呈する成因となっている可能性が示唆された

Potential Neuroprotective Effects of an LSD1 Inhibitor in Retinal Ganglion Cells via p38 MAPK Activity

Citation: Tsutsumi T, Iwao K, Hayashi H, et al. Potential neuroprotective effects of an LSD1 inhibitor in retinal ganglion cells via p38 MAPK activity. Invest Ophthalmol Vis Sci. 2016;57:6461-6473. DOI:10.1167/ iovs.16-19494 PURPOSE. The epigenetic mechanisms associated with ocular neurodegenerative diseases remain unclear. The present study aimed to determine the role of lysine-specific demethylase 1 (LSD1), which represses transcription by removing the methyl group from methylated lysine 4 of histone H3, in retinal ganglion cell (RGC) survival, and to investigate the details of the neuroprotective mechanism of tranylcypromine, a major LSD1 inhibitor. METHODS. The authors evaluated whether tranylcypromine contributes to neuronal survival following stress-induced damage using primary cultured rat RGCs and in vivo N-methyl-Daspartate (NMDA)-induced excitotoxicity. Additionally, the molecules associated with tranylcypromine treatment were assessed by microarray and immunoblot analysis. RESULTS. Tranylcypromine significantly suppressed neuronal cell death following glutamate neurotoxicity and oxidative stress. Microarray and immunoblot analyses revealed that p38 mitogen-activated protein kinase (MAPK)c was a key molecule involved in the neuroprotective mechanisms induced by tranylcypromine because the significant suppression of p38 MAPKc by glutamate was reversed by tranylcypromine. Moreover, although pharmacologic inhibition of the phosphorylation of the total p38 MAPKs interfered with neuroprotective effects of tranylcypromine, the specific inhibition of p38 MAPKa and p38 MAPKb did not influence RGC survival. This suggests that the non-p38 MAPKa/b isoforms have important roles in neuronal survival by tranylcypromine. Additionally, the intravitreal administration of tranylcypromine significantly saved RGC numbers in an in vivo glaucoma model employing NMDA-induced excitotoxicity. CONCLUSIONS. These findings indicate that tranylcypromine-induced transcriptional and epigenetic regulation modulated RGC survival via the promotion of p38 MAPKc activity. Therefore, pharmacologic treatments that suppress LSD1 activity may be a novel therapeutic strategy that can be used to treat neurodegenerative diseases

MCAF1 and synergistic activation of the transcription of Epstein–Barr virus lytic genes by Rta and Zta

Epstein–Barr virus (EBV) expresses two transcription factors, Rta and Zta, during the immediate-early stage of the lytic cycle. The two proteins often collaborate to activate the transcription of EBV lytic genes synergistically. This study demonstrates that Rta and Zta form a complex via an intermediary protein, MCAF1, on Zta response element (ZRE) in vitro. The interaction among these three proteins in P3HR1 cells is also verified via coimmunoprecipitation, CHIP analysis and confocal microscopy. The interaction between Rta and Zta in vitro depends on the region between amino acid 562 and 816 in MCAF1. In addition, overexpressing MCAF1 enhances and introducing MCAF1 siRNA into the cells markedly reduces the level of the synergistic activation in 293T cells. Moreover, the fact that the synergistic activation depends on ZRE but not on Rta response element (RRE) originates from the fact that Rta and Zta are capable of activating the BMRF1 promoter synergistically after an RRE but not ZREs in the promoter are mutated. The binding of Rta–MCAF1–Zta complex to ZRE but not RRE also explains why Rta and Zta do not use RRE to activate transcription synergistically. Importantly, this study elucidates the mechanism underlying synergistic activation, which is important to the lytic development of EBV

Nedd4 Regulates Egress of Ebola Virus-Like Particles from Host Cells

Ebola virus budding is mediated by two proline-rich motifs, PPxY and PTAP, within the viral matrix protein VP40. We have previously shown that a Nedd4-like protein BUL1, but not Nedd4, positively regulates budding of type D retrovirus Mason-Pfizer monkey virus (J. Yasuda, E. Hunter, M. Nakao, and H. Shida, EMBO Rep. 3:636-640, 2002). Here, we report that the cellular E3 ubiquitin ligase Nedd4 regulates budding of VP40-induced virus-like particles (VLPs) through interaction with the PPxY motif. Mutation of the active site cysteine (C894A), resulting in abrogation of ubiquitin ligase activity, impaired the function of Nedd4 on budding. In addition, the WW domains of Nedd4 are essential for binding to the viral PPxY motif, and a small fragment of Nedd4 containing only WW domains significantly inhibited Ebola VLP budding in a dominant-negative manner. Our findings suggest that the viruses containing PPxY as an L-domain motif specifically use E3 in the process of virus budding. We also examined the effects of overexpression of Tsg101 and its mutant. As expected, Tsg101 enhanced VP40-induced VLP release, and TsgΔC, which lacks its C-terminal half, inhibited VLP release. These results indicate that Nedd4, together with Tsg101, plays an important role in Ebola virus budding