Jang Shyh-Ing, Ong Hwei Ling, Liu Xibao, Alevizos Ilias, Ambudkar Indu S
Sjögren's Syndrome and Salivary Gland Dysfunction Unit, Molecular Physiology and Therapeutics Branch, NIDCR/National Institutes of Health, Bethesda, Maryland 20892.
From the Secretory and Physiology Section and.
J Biol Chem. 2016 Apr 15;291(16):8709-20. doi: 10.1074/jbc.M115.701607. Epub 2016 Feb 22.
The signaling pathways involved in the generation and maintenance of exocrine gland acinar cells have not yet been established. Primary human salivary gland epithelial cells, derived from salivary gland biopsies, acquired an acinar-like phenotype when the [Ca(2+)] in the serum-free medium (keratinocyte growth medium, KGM) was increased from 0.05 mm (KGM-L) to 1.2 mm (KGM-H). Here we examined the mechanism underlying this Ca(2+)-dependent generation of the acinar cell phenotype. Compared with cells in KGM-L, those in KGM-H display enhancement of Orai1, STIM1, STIM2, and nuclear factor of activated T cells 1 (NFAT1) expression together with an increase in store-operated Ca(2+) entry (SOCE), SOCE-dependent nuclear translocation of pGFP-NFAT1, and NFAT-dependent but not NFκB-dependent gene expression. Importantly, AQP5, an acinar-specific protein critical for function, is up-regulated in KGM-H via SOCE/NFAT-dependent gene expression. We identified critical NFAT binding motifs in the AQP5 promoter that are involved in Ca(2+)-dependent up-regulation of AQP5. These important findings reveal that the Ca(2+)-induced switch of salivary epithelial cells to an acinar-like phenotype involves remodeling of SOCE and NFAT signaling, which together control the expression of proteins critically relevant for acinar cell function. Our data provide a novel strategy for generating and maintaining acinar cells in culture.
外分泌腺腺泡细胞生成和维持过程中涉及的信号通路尚未明确。源自唾液腺活检的原代人唾液腺上皮细胞,当无血清培养基(角质形成细胞生长培养基,KGM)中的[Ca(2+)]从0.05 mM(KGM-L)增加到1.2 mM(KGM-H)时,会获得腺泡样表型。在此,我们研究了这种Ca(2+)依赖性腺泡细胞表型生成的潜在机制。与KGM-L中的细胞相比,KGM-H中的细胞Orai1、STIM1、STIM2和活化T细胞核因子1(NFAT1)的表达增强,同时储存-操作性Ca(2+)内流(SOCE)增加,pGFP-NFAT1的SOCE依赖性核转位以及NFAT依赖性而非NFκB依赖性基因表达增加。重要的是,水通道蛋白5(AQP5)是一种对功能至关重要且具有腺泡特异性的蛋白质,在KGM-H中通过SOCE/NFAT依赖性基因表达上调。我们在AQP5启动子中鉴定出关键的NFAT结合基序,其参与了Ca(2+)依赖性的AQP5上调。这些重要发现表明,Ca(2+)诱导的唾液上皮细胞向腺泡样表型的转变涉及SOCE和NFAT信号通路的重塑,它们共同控制着与腺泡细胞功能密切相关的蛋白质的表达。我们的数据为在培养中生成和维持腺泡细胞提供了一种新策略。