Proteomics analysis of NAD+ depletion combined with reovirus infection in KMS12 multiple myeloma cells

Proteomics dataset of multiple myeloma cancer cell lines infected with reovirus and/or in the context of NAD+ synthesis inhibition. Experimentally, the human multiple myeloma cell line (KMS12PE) were infected or not (not-treated [NT]) with reovirus (Reo, multitude of infectivity of 10) and/or treated with NAMPT inhibitor FK866 (FK, 20 nM) and/or NAMPT product, NMN (100 uM) for 24 hours. Trypsin-digested peptides were labeled using TMT 11-plex reagents as described previously (doi.org/10.1074/mcp.M114.045849). TMT11-labeled samples were fractionated using high-pH reverse-phase chromatography performed with an Onyx monolithic 100 × 4.6-mm C18 column (Phenomenex). Fractions were desalted using homemade Stage Tips, lyophilized, and analyzed with an Orbitrap Velos Mass Spectrometer (Thermo Fisher Scientific) using an MS3 method as described previously (doi.org/10.1074/mcp.M114.045849). Protein identification was performed using a database search against a mouse proteome database (downloaded from UniProtKB in September 2014) concatenated to a mammalian orthoreovirus 3 (Dearing strain) database (downloaded from UniProtKB in September 2014). All FDR filtering and protein quantitation was performed as described previously (doi.org/10.1074/mcp.M114.045849). A protein was required to have a minimum total signal-to-noise ratio of 100 in all TMT reporter channels.

多发性骨髓瘤癌细胞系蛋白质组学数据集，包括受嗜神经病毒感染以及/或在NAD+合成抑制的背景下的样本。在实验中，人类多发性骨髓瘤细胞系（KMS12PE）或受感染（受病毒感染[Reo，感染性10倍]）或未经处理（未处理[NT]），以及/或用NAMPT抑制剂FK866（FK，20 nM）和/或NAMPT产物，NMN（100 μM）处理24小时。采用先前描述的方法（doi.org/10.1074/mcp.M114.045849）使用TMT 11-plex试剂对胰蛋白酶消化的肽段进行标记。利用Onyx单晶100 × 4.6-mm C18柱（Phenomenex）进行的高pH值反相液相色谱法对TMT标记样本进行分离。使用自制的Stage Tips对分馏物进行脱盐处理，冷冻干燥，并使用Orbitrap Velos质谱仪（Thermo Fisher Scientific）进行MS3方法分析，如先前所述（doi.org/10.1074/mcp.M114.045849）。蛋白质鉴定通过将小鼠蛋白质组数据库（2014年9月从UniProtKB下载）与哺乳动物正粘病毒3型（Dearing株）数据库（2014年9月从UniProtKB下载）连接，进行数据库搜索。所有FDR过滤和蛋白质定量均按照先前描述进行（doi.org/10.1074/mcp.M114.045849）。蛋白质在所有TMT报告通道中必须满足最小总信噪比为100的要求。