Leukocyte immune-type receptors (LITRs) are a multigene family of teleost immunoregulatory proteins that share structural, phylogenetic, and likely functional relationships with several innate immune receptor proteins in other vertebrates, including mammals. Originally discovered in channel catfish (Ictalurus punctatus), representative IpLITR-types have been shown to regulate diverse innate immune cell effector responses including phagocytosis, degranulation, and cytokine secretion. To date, IpLITRs have been primarily characterized using mammalian cell line expression systems, therefore many unanswered questions remain regarding their actual regulatory roles in fish immunity. In the present study, we report on the preliminary molecular characterization of five goldfish (Carassius auratus) CaLITR-types and the identification of several putative splice variants of these receptors cloned from various goldfish tissues and primary myeloid cell cultures. In general, CaLITR mRNA transcripts were detected in all goldfish tissues tested, and also in primary kidney macrophage and neutrophil cultures. Specifically, CaLITR1 is a functionally ambiguous receptor with no charged amino acids in its transmembrane (TM) segment and is devoid of tyrosine-based signaling motifs in its short cytoplasmic tail (CYT) region. CaLITR2 is a putative activating receptor-type that contains immunotyrosine-based activation motifs (ITAMs) within its long CYT region, and CaLITR3 has a positively charged TM segment, suggesting that it may recruit intracellular stimulatory adaptor signaling molecules. CaLITR4 and CaLITR5 appear to have diverse signaling capabilities since they contain various immunoregulatory signaling motifs within their CYT regions including putative Nck and STAT recruitment motifs as well as ITAM-like and ITIM sequences. We also identified putative CaLITR splice variants with altered extracellular Ig-like domain compositions and variable CYT regions. Interestingly, this suggests that alternative splicing-mediated diversification of CaLITRs can generate receptor forms with possible variable binding and/or intracellular signaling abilities. Overall, these findings reveal new information about the teleost LITRs and sets the stage for exploring how alternative splicing leads to the functional diversification of this complex multigene immunoregulatory receptor family.

白细胞免疫型受体（LITRs）是硬骨质免疫调节蛋白的多基因家族，与其他脊椎动物（包括哺乳动物）的几种先天免疫受体蛋白具有结构，系统发育和可能的功能关系。代表性的IpLITR类型最初发现于channel鱼（Ictalurus punctatus）中，可调节多种固有的免疫细胞效应反应，包括吞噬作用，脱粒作用和细胞因子分泌。迄今为止，已经使用哺乳动物细胞系表达系统对IpLITRs进行了主要鉴定，因此，关于其在鱼类免疫中的实际调节作用，还有许多未解决的问题。在本研究中，我们报告了五个金鱼（Car鱼）的初步分子表征CaLITR类型，并鉴定了从各种金鱼组织和原代骨髓细胞培养物中克隆的这些受体的几种假定剪接变体。通常，在所有测试的金鱼组织以及原代肾巨噬细胞和中性粒细胞培养物中均检测到CaLITR mRNA转录本。具体而言，CaLITR1是功能模糊的受体，在其跨膜（TM）段中没有带电荷的氨基酸，并且在其短细胞质尾（CYT）区域中没有基于酪氨酸的信号基序。CaLITR2是假定的激活受体类型，在其长CYT区域内包含基于免疫酪氨酸的激活基序（ITAM），CaLITR3具有带正电的TM片段，表明它可能募集细胞内刺激衔接子信号分子。CaLITR4和CaLITR5似乎具有多种信号传导功能，因为它们在其CYT区域内包含各种免疫调节信号传导基序，包括推定的Nck和STAT募集基序以及ITAM样和ITIM序列。我们还确定了推定的CaLITR剪接变异体，具有改变的细胞外Ig样结构域组成和可变的CYT区。有趣的是，这表明CaLITRs的选择性剪接介导的多样化可以产生具有可能的可变结合和/或细胞内信号传导能力的受体形式。总体而言，这些发现揭示了有关硬骨鱼LITR的新信息，并为探索选择性剪接如何导致这种复杂的多基因免疫调节受体家族功能多样化奠定了基础。