State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Veterinary Parasitology of Gansu Province, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, Gansu, China.
State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Veterinary Parasitology of Gansu Province, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, Gansu, China.
Exp Parasitol. 2022 Nov;242:108403. doi: 10.1016/j.exppara.2022.108403. Epub 2022 Oct 6.
The small ubiquitin-like modifier (SUMO) plays important roles, with the SUMOylation pathway as one of its core components. In the present work, a single SUMO gene was initially identified from Taenia pisiformis and designated as TpSUMO. Bioinformatic analysis showed that the TpSUMO gene contained a 309 bp open reading frame (ORF), encoding 102 amino acids, and had a predicted molecular weight of ∼12 kDa. The amino acid sequence of TpSUMO was deduced and it shared 44.00% identity with human SUMO2 (HsSUMO2) and exhibited more than 97.78% identity with SUMOs from Taenia and Echinococcus. TpSUMO possessed a putative non-consensus site (FKMG) within its N-terminus and a typical di-glycine (GG) motif at the C-terminus. Basic local alignment search tool (BLAST) analysis showed that only a single SUMO-related ortholog was present in each set of known genome data for fourteen tapeworm species. The precursor His-TpSUMO-FL, mature His-TpSUMO-GG and mutant His-TpSUMO-GG proteins (∼18 kDa) were expressed in Escherichia coli Rosseta (DE3), and rabbit polyclonal anti-TpSUMO was generated with a high titer of 1.28 × 10. In vitro SUMOylation assay results showed that TpSUMO multimer formation in the His-TpSUMO-GG reaction could be catalyzed by the human SAE1/SAE2 and UBC9 conjugation system, but K11R mutation disrupted TpSUMO chain synthesis. Quantitative real-time PCR (qRT-PCR) further revealed that TpSUMO was ubiquitously expressed in different stages of T. pisiformis and in higher levels during an early development phase (day 14) of adult worms. Immunofluorescence localization showed that TpSUMO was detected in the bladder wall of cysticerci, in the testis in immature segment, and within eggs in the gravid proglottids. These findings indicated that TpSUMO is a new member of the SUMO protein family and may play a vital role in regulation of functions within proteins involved in worm growth and development.
小泛素样修饰物 (SUMO) 发挥着重要作用，其 SUMO 化途径是其核心组成部分之一。本研究从带绦虫中首次鉴定出一个 SUMO 基因，命名为 TpSUMO。生物信息学分析表明，TpSUMO 基因含有一个 309 bp 的开放阅读框 (ORF)，编码 102 个氨基酸，预测分子量约为 12 kDa。推导的 TpSUMO 氨基酸序列与人类 SUMO2 (HsSUMO2) 具有 44.00%的同源性，与带绦虫和细粒棘球绦虫的 SUMO 具有超过 97.78%的同源性。TpSUMO 在其 N 端具有一个假定的非共识位点 (FKMG)，在 C 端具有一个典型的双甘氨酸 (GG) 基序。基本局部比对搜索工具 (BLAST) 分析表明，在 14 种绦虫的已知基因组数据中，每组仅存在一个 SUMO 相关直系同源物。原核表达系统中表达了 His-TpSUMO-FL、成熟 His-TpSUMO-GG 和突变 His-TpSUMO-GG 蛋白 (∼18 kDa)，并以 1.28×10 的高滴度产生了兔多克隆抗 TpSUMO。体外 SUMOylation 测定结果表明，TpSUMO 在 His-TpSUMO-GG 反应中的多聚体形成可被人 SAE1/SAE2 和 UBC9 缀合系统催化，但 K11R 突变破坏了 TpSUMO 链合成。实时定量 PCR (qRT-PCR) 进一步表明，TpSUMO 在带绦虫的不同发育阶段广泛表达，在成虫早期发育阶段 (第 14 天) 表达水平较高。免疫荧光定位显示，TpSUMO 存在于囊尾蚴的膀胱壁、未成熟节段的睾丸内和妊娠节片的卵内。这些发现表明，TpSUMO 是 SUMO 蛋白家族的一个新成员，可能在调节与蠕虫生长和发育相关的蛋白质功能中发挥重要作用。
