Multiple transcripts of anoctamin genes expressed in the mouse submandibular salivary gland
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Purpose: Salivary fluid formation is primarily driven by Ca2+-activated, apical efflux of chloride into the lumen of the salivary acinus. The anoctamin1 protein is an anion channel with properties resembling the endogenous calcium-activated chloride channels. In order to better understand the role of anoctamin proteins in salivary exocrine secretion, the ex¬pression of the ten members of the anoctamin gene family in the mouse submandibular gland was studied. Methods: Total RNA extracted from mouse submandibular salivary glands was reverse transcribed using primer pairs to amplify the full-length coding regions of each anoctamin gene and was subcloned into plasmid vectors for DNA sequencing. Alternative splice vari¬ants were also screened by polymerase chain reaction using primer pairs that amplified six overlapping regions of the complementary DNA of each anoctamin gene, spanning multi¬ple exons. Results: Multiple anoctamin transcripts were found in the mouse submandibular salivary gland, including full-length transcripts of anoctamin1, anoctamin3, anoctamin4, anocta¬min5, anoctamin6, anoctamin9, and anoctamin10. Exon-skipping splicing in the N-terminal exons of the anoctamins1, anoctamin5, and anoctamin6 genes resulted in multiple alterna¬tive splice variants. No expression of anoctamin2, anoctamin7, or anoctamin8 was found. Conclusions: The predominant anoctamin transcript expressed in the mouse submandibu¬lar gland is anoctamin1ac. The chloride channel protein produced by anoctamin1ac is likely responsible for the Ca2+-activated chloride efflux, which is the rate-limiting step in salivary exocrine secretion.
Abstract
Purpose: Salivary fluid formation is primarily driven by Ca2+-activated, apical efflux of chloride into the lumen of the salivary acinus. The anoctamin1 protein is an anion channel with properties resembling the endogenous calcium-activated chloride channels. In order to better understand the role of anoctamin proteins in salivary exocrine secretion, the ex¬pression of the ten members of the anoctamin gene family in the mouse submandibular gland was studied. Methods: Total RNA extracted from mouse submandibular salivary glands was reverse transcribed using primer pairs to amplify the full-length coding regions of each anoctamin gene and was subcloned into plasmid vectors for DNA sequencing. Alternative splice vari¬ants were also screened by polymerase chain reaction using primer pairs that amplified six overlapping regions of the complementary DNA of each anoctamin gene, spanning multi¬ple exons. Results: Multiple anoctamin transcripts were found in the mouse submandibular salivary gland, including full-length transcripts of anoctamin1, anoctamin3, anoctamin4, anocta¬min5, anoctamin6, anoctamin9, and anoctamin10. Exon-skipping splicing in the N-terminal exons of the anoctamins1, anoctamin5, and anoctamin6 genes resulted in multiple alterna¬tive splice variants. No expression of anoctamin2, anoctamin7, or anoctamin8 was found. Conclusions: The predominant anoctamin transcript expressed in the mouse submandibu¬lar gland is anoctamin1ac. The chloride channel protein produced by anoctamin1ac is likely responsible for the Ca2+-activated chloride efflux, which is the rate-limiting step in salivary exocrine secretion.
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