The inositol 1,4,5-trisphosphate receptor type 2 protein domains regulate calcium levels and Ion transport gene expression in laying ducks' uterus.

This study aimed to investigate the regulatory role of the inositol 1,4,5-trisphosphate receptor type 2 (IP3R2) protein domains in calcium ion (Ca2+) homeostasis and ion transport gene expression in duck uterine epithelial cells. Coding sequences corresponding to three IP3R2 protein domains-IP3R2-MIR, IP3R2-RYDR_ITPR (1), and IP3R2-RYDR_ITPR (2)-were cloned into the pCDNA3.1 vector with a Flag tag to generate expression constructs, which were subsequently transfected into primary duck uterine epithelial cells. Domain expression levels were quantified by ELISA, and intracellular Ca²⁺ concentrations were measured using the Fluo-3/AM fluorescent probe. Additionally, the mRNA expression of 20 ion transport-related genes was analyzed by RT-qPCR in cells overexpressing the domain-coding sequences to assess potential correlations with IP3R2 activity. Results demonstrated that overexpression of each IP3R2 protein domain decreased intracellular Ca²⁺ levels (P < 0.05), and Ca²⁺ concentrations were negatively correlated with domain expression levels (P < 0.05). Furthermore, the three IP3R2 protein domains exerted distinct regulatory effects on the expression of ion transport genes, and domain overexpression changed the correlation among the expression levels of these genes (P < 0.05). These findings provided new insights into the molecular mechanisms underlying eggshell mineralization and suggested a functional link between IP3R2-mediated Ca²⁺ signaling and ion transport gene networks in the duck uterus.