Transcriptomic and RNAi analyses reveal chloride channel 3-associated osmoregulation in Litopenaeus vannamei under low-salinity stress.

Ti Wang; Qijian Zhu; Xuenan Li; Yun Li

doi:10.1016/j.cbd.2026.101987

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Transcriptomic and RNAi analyses reveal chloride channel 3-associated osmoregulation in Litopenaeus vannamei under low-salinity stress.

Ti Wang·Qijian Zhu·Xuenan Li·Yun Li

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Abstract

Chloride channels and transporters are important for cellular volume regulation and salinity adaptation in euryhaline crustaceans, yet the intestinal transcriptional relationship between plasma-membrane and intracellular chloride pathways remains unclear in Litopenaeus vannamei. In this study, RNA interference of anoctamin 1 (ANO1) was combined with intestinal transcriptome sequencing under the production-relevant low-salinity condition of salinity 3. ANO1 silencing produced a focused transcriptional response, with 16 differentially expressed genes (DEGs) identified (11 upregulated and 5 downregulated). Functional enrichment indicated that these genes were associated with transporter activity, cytoskeletal organization, extracellular matrix-receptor interaction, membrane lipid metabolism, and vesicular processes. Notably, a transcript encoding chloride channel protein 3 (CLC-3) was significantly upregulated following ANO1 knockdown, suggesting a potential transcriptional relationship between ANO1 and CLC-3 in chloride homeostasis. Based on this finding, CLC-3 was selected for full-length cDNA cloning, sequence characterization, salinity-gradient expression analysis, and RNAi-based functional assessment. The cloned CLC-3 cDNA was 2883 bp in length and encoded an 850 amino acid protein containing a conserved voltage-gated chloride channel (Voltage-CLC) domain and two cystathionine β-synthase domains. Phylogenetic analysis placed LvCLC-3 within the intracellular CLC-c clade, and tissue distribution analysis showed the highest CLC-3 expression in the intestine. Intestinal CLC-3 expression responded nonlinearly to salinity variation, peaking at salinity 20. Under salinity 3, CLC-3 knockdown reduced ANO1, Na+/K+-ATPase alpha subunit, and Na+-K+-2Cl- cotransporter transcript levels, whereas glutamate-gated chloride channel expression increased. Mild hepatopancreatic structural alterations were also observed after CLC-3 knockdown. These findings suggest that CLC-3 is a salinity-responsive intracellular chloride-transporter candidate associated with intestinal ion-transport-related transcriptional responses after ANO1 suppression in L. vannamei, although the underlying physiological mechanism requires further validation.

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Animals Chloride Channels RNA Interference Penaeidae Osmoregulation Transcriptome Phylogeny Salinity Amino Acid Sequence Salt Stress Gene Expression Profiling ANO1 CLC-3 Chloride transport Litopenaeus vannamei Low salinity RNA interference Transcriptome sequencing Journal Article

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Ti Wang, Qijian Zhu, Xuenan Li, Yun Li. 2026-08-21. Transcriptomic and RNAi analyses reveal chloride channel 3-associated osmoregulation in Litopenaeus vannamei under low-salinity stress.. https://doi.org/10.1016/j.cbd.2026.101987

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Journal Article resource ANO1 subject Amino Acid Sequence subject Animals subject CLC-3 subject Chloride Channels subject Chloride transport subject Gene Expression Profiling subject Journal Article subject Litopenaeus vannamei subject Low salinity subject Osmoregulation subject Penaeidae subject

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