Alternative splicing is an essential molecular mechanism that increase the protein diversity of a species to regulate important biological processes. As a transcription factor, Interleukin-2 enhancer binding factor 2 (ILF2) regulates the functions of interleukin-2 (IL-2) at the levels of transcription, splicing and translation, and plays other critical roles in the immune system. ILF2 is well-documented in vertebrates, while little is currently known in crustacean species such as the Pacific white shrimp (Litopenaeus vannamei). In the present study, five cDNA for spliced isoforms of Lv-ILF2 were identified, in which four of them are the full-length long isoforms (Lv-ILF2-L1, Lv-ILF2-L2, Lv-ILF2-L3 and Lv-ILF2-L4) and one of them is a truncated short isoform (Lv-ILF2-S). The whole sequence of ILF2 gene from L. vannamei was obtained, which is 11,680 bp in length with 9 exons separated by 8 introns. All five isoforms contain a domain associated with zinc fingers (DZF). Two alternative splicing types (alternative 5′ splice site and alternative 3′ splice site) were identified in the five isoforms. The Lv-ILF2 mRNA showed a broad distribution in all detected tissues, and the Lv-ILF2-L transcript levels were higher than those of Lv-ILF2-S in corresponding tissues. The mRNA levels of Lv-ILF2-S in the hepatopancreas, heart, muscle and stomach, but not in the eyestalk, were significantly increased after challenges with Vibrio harveyi or lipopolysaccharide (LPS), while no significant changes were observed for the transcript levels of Lv-ILF2-L in these tissues under the same immune stimulants. On the contrary, the transcript levels of neither Lv-ILF2-S nor Lv-ILF2-L were affected by challenges of polyinosinic: polycytidylic acid [Poly (I:C)]. In addition, after knockdown of the Lv-ILF2 mRNA level by siRNA, the mortality of shrimp and the hepatopancreatic bacterial numbers were significantly increased under V. harveyi challenge, indicating that Lv-ILF2 might participate in the immune defenses against V. harveyi invasion. Collectively, our study here supplied the first evidence for a novel splicing mechanism of ILF2 transcripts, and provided a functional link between the Lv-ILF2 isoforms and the capacity against pathogenic Vibrio in penaeid shrimp.

选择性剪接是一种重要的分子机制，可增加物种的蛋白质多样性以调节重要的生物过程。作为一种转录因子，白介素 2 增强子结合因子 2 (ILF2) 在转录、剪接和翻译水平上调节白介素 2 (IL-2) 的功能，并在免疫系统中发挥其他关键作用。ILF2 在脊椎动物中有充分的记录，而目前在甲壳类动物如太平洋白虾 ( Litopenaeus vannamei ) 中知之甚少。在本研究中，鉴定了Lv- ILF2剪接同种型的 5 个 cDNA ，其中 4 个是全长长同种型（Lv -ILF2-L1、Lv -ILF2-L2、Lv -ILF2-L3 和Lv- ILF2-L4)，其中之一是截短的短异构体 ( Lv- ILF2-S)。获得南美白对虾ILF2基因全序列，全长11680 bp，9个外显子，8个内含子。所有五种异构体都包含一个与锌指 (DZF) 相关的结构域。在五种亚型中鉴定了两种替代剪接类型（替代 5' 剪接位点和替代 3' 剪接位点）。Lv-ILF2 mRNA在所有检测到的组织中分布广泛，Lv -ILF2-L转录本水平高于相应组织中的Lv-ILF2-S。Lv-ILF2-S的 mRNA 水平在用哈维弧菌或脂多糖 (LPS)攻击后，在肝胰腺、心脏、肌肉和胃中，但在眼柄中没有显着增加，而在这些组织中Lv-ILF2-L的转录水平没有观察到显着变化。相同的免疫刺激剂。相反，Lv-ILF2-S和Lv-ILF2-L的转录水平均不受聚肌苷挑战的影响：聚胞苷酸 [Poly (I:C)]。此外，通过 siRNA 敲低Lv-ILF2 mRNA 水平后，在哈维氏弧菌攻击下，虾的死亡率和肝胰腺细菌数量显着增加，表明Lv-ILF2 可能参与针对哈维氏弧菌入侵的免疫防御。总的来说，我们在此的研究为ILF2转录物的新剪接机制提供了第一个证据，并提供了 Lv-ILF2 同种型与对虾对致病性弧菌的能力之间的功能联系。