ゲンチョ チョウセイ ノウカン ハンノウ ケンサ ノ リンショウテキ ケンキュウ 5ネンカン 273レイ ニ ツイテノ ケントウ

聴性脳幹反応(Auditory Brainstem Response,以下ABRと略す)は音刺激により誘発される聴覚伝導路由来の脳波であり,1970年にJewettらにより報告されて以来,他覚的聴力検査及び脳幹部の神経学的検査として臨床的に広く活用されている。我々は3kHzサイン波一波のクリックを刺激音とするABRを施行しているが,今回1985年から1989年までの5年間に施行した273例について検討した。症例は男性147例,女性126例であり,年齢的に純音聴力検査が不可能である0-4歳の幼児に多く施行した。幼児に対し施行した聴力検査症例152例のうち,85例に主訴となった聴力障害以外の何等かの基礎疾患があり,その半数が閾値上昇を示した。これらの疾患を有する症例では,難聴の合併率が高いばかりでなく,脳幹部を始めとする中枢神経系に異常を持つことがあるため,ABRがスクリーニング検査として非常に有用であり,早期及び経時的に施行することが必要であると思われた。また聴神経腫瘍を主とする小脳橋角部腫瘍の診断にあたっては,ABRのV波潜時の延長所見の検索は極めて有用であり,診断学的に高い特異性と感受性を有する検査手段のひとつとみなされる。さらに,他覚的聴力検査としてABRの記録は,機能性難聴の確定診断にも不可欠である事が確かめられた。journal articl

Measurement of the e+e-→D(*)+D(*)- cross sections

journal articl

KSHV Ref Genome GQ994935.1 with known annotations.zip

This is a Supplemental File for the scientific article entitled: KSHV 3.0: A State-of-the-Art Annotation of the Kaposi’s Sarcoma-Associated Herpesvirus Transcriptome Using Cross-Platform Sequencing.The .zip file contains .gb, .gff3 and .geneious files about the reannotated KSHV transcriptome made by the 3G research group.</p

LANA binds to the entire KSHV genome and is required for the recruitment of EZH2 onto the viral episome during <i>de novo</i> infection.

(A) Immunoblot analysis of LANA expression in BAC16-3xF-LANA infected SLK cells at 72 hpi using anti-LANA and anti-FLAG antibodies. (B and C) FLAG and LANA ChIP assays for testing LANA-binding to viral promoters in BAC16 and BAC16-3xF-LANA KSHV-infected SLK cells at 72 hpi. Cellular MYT1 promoter was used as a control. (D) ChIP-on-chip analysis of the genome-wide binding of LANA and EZH2 to the KSHV genome in WT and LANA KO KSHV-infected cells at 72 hpi.</p

LANA-Mediated Recruitment of Host Polycomb Repressive Complexes onto the KSHV Genome during <i>De Novo</i> Infection

<div><p>One of the hallmarks of the latent phase of Kaposi’s sarcoma-associated herpesvirus (KSHV) infection is the global repression of lytic viral gene expression. Following <i>de novo</i> KSHV infection, the establishment of latency involves the chromatinization of the incoming viral genomes and recruitment of the host Polycomb repressive complexes (PRC1 and PRC2) to the promoters of lytic genes, which is accompanied by the inhibition of lytic genes. However, the mechanism of how PRCs are recruited to the KSHV episome is still unknown. Utilizing a genetic screen of latent genes in the context of KSHV genome, we identified the latency-associated nuclear antigen (LANA) to be responsible for the genome-wide recruitment of PRCs onto the lytic promoters following infection. We found that LANA initially bound to the KSHV genome right after infection and subsequently recruited PRCs onto the viral lytic promoters, thereby repressing lytic gene expression. Furthermore, both the DNA and chromatin binding activities of LANA were required for the binding of LANA to the KSHV promoters, which was necessary for the recruitment of PRC2 to the lytic promoters during <i>de novo</i> KSHV infection. Consequently, the LANA-knockout KSHV could not recruit PRCs to its viral genome upon <i>de novo</i> infection, resulting in aberrant lytic gene expression and dysregulation of expression of host genes involved in cell cycle and proliferation pathways. In this report, we demonstrate that KSHV LANA recruits host PRCs onto the lytic promoters to suppress lytic gene expression following <i>de novo</i> infection.</p></div

Additional LANA expression is not sufficient for the early recruitment of EZH2 onto the KSHV genome.

<p><b>(A)</b> Time course analysis of LANA protein expression during <i>de novo</i> infection. <b>(B)</b> Time course ChIP analysis of LANA-binding to viral promoters during <i>de novo</i> infection. <b>(C)</b> Immunoblot analysis of LANA expression in empty lentiviral vector and 3xFLAG-LANA expressing lentivirus infected cells. <b>(D)</b> ChIP assays for binding of LANA and EZH2 to viral promoters in LANA-overexpressing cells during <i>de novo</i> KSHV infection.</p

Co-localization of KDM2B with LANA in latent KSHV-infected cells.

(A) Uninfected iSLK cells or (B) KSHV-infected iSLK cells (iSLKBAC16-3xFLAG-LANA) were subjected to immunofluorescence analysis for LANA (red) and KDM2B (green). Rabbit IgG served as a negative control. FLAG antibody was used to detect 3xFLAG-LANA expressed from KSHV. (C) Representative LANA puncta were connected by yellow line and the co-localization of LANA with KDM2B along the line was analyzed by ImageJ.</p

GATAD2B, KDM2B, and ETV6 repress the spontaneous lytic reactivation of KSHV in BCBL1 cells.

(A) Representative immunofluorescence images showing the expression of ORF6 and K8 lytic proteins in BCBL1 cells 72 hours after shRNA lentivirus infection. (B) Quantification of the number of K8, ORF6, and K8/ORF6 positive BCBL1 cells. The average is calculated from at least 10 randomly selected fields.</p

Co-localization and focal enrichment of polycomb proteins with LANA in KSHV-infected cells.

<p>KSHV-infected iSLK cells (iSLKBAC16-3xF-LANA) were subjected to confocal microscopic analysis for LANA (Green, false color) and polycomb proteins (Red). SPT5 and CYCT1 served as controls. FLAG antibody was used to detect the 3xF-LANA. Representative LANA puncta were connected by white lines and the co-localization of LANA with the indicated cellular proteins was measured by the image processing program ImageJ.</p

LANA-binding to the lytic KSHV promoters is required for the EZH2 recruitment during <i>de novo</i> infection.

<p><b>(A)</b> Immunoblot analysis of SLK cells infected with lentiviruses expressing 3xFLAG-tagged WT or mutant LANA using anti-FLAG antibody. <b>(B)</b> ChIP assays for the recruitment of LANA (using αLANA antibody) and EZH2 on viral promoters in RTA KO and LANA/RTA dKO KSHV-infected SLK cells as well as in LANA/RTA dKO KSHV-infected SLK cells expressing WT or mutant LANA. <b>(C)</b> Total lysates of 293T cells expressing either 3xF-LANA or 3xF-GMRRL mutant LANA were subjected to FLAG immunoprecipitation followed by immunoblotting with anti-FLAG and anti-EZH2 antibodies. Input lysates were included as controls. <b>(D)</b> ChIP assays for LANA and EZH2 occupancy on the KSHV promoters during <i>de novo</i> LANA/RTA dKO KSHV infection of WT- or GMRRL mutant LANA-expressing SLK cells. <b>(E)</b> Model showing LANA-mediated recruitment PRCs onto the KSHV genome during <i>de novo</i> infection.</p