Analysis of human sodium iodide symporter gene expression in extrathyroidal tissues and cloning of its complementary deoxyribonucleic acids from salivary gland, mammary gland, and gastric mucosa
Analysis of human sodium iodide symporter gene expression in extrathyroidal tissues and cloning of its complementary deoxyribonucleic acids from salivary gland, mammary gland, and gastric mucosa
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DOI：
10.1210/jc.83.5.1746
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发表时间：
1998-05-01
影响因子：
5.8
通讯作者：
Heufelder, AE
中科院分区：
文献类型：
--
作者：
Spitzweg, C;Joba, W;Heufelder, AE
浓缩碘化物的能力是正常功能的甲状腺组织的基本特性，是产生甲状腺激素的第一步。碘化物的吸收已在各种甲状腺外组织中得到证实，包括唾液腺、胃粘膜和哺乳期乳腺。最近，人类碘化钠同向转运蛋白（hNIS）的克隆和分子表征已被报道；然而，hNIS 基因在人体组织中的表达模式仍不清楚。为了检查人类 hNIS 基因在各种正常人体组织中的表达谱，我们使用 P-32 标记的 hNIS 特异性互补 DNA (cDNA) 探针（核苷酸 1184-1667）进行了高严格的 Northern 印迹分析。为了检测小组织样本中罕见的 hNIS 转录物，使用一对 hNIS 特异性寡核苷酸引物进行 RT-PCR，该引物设计用于扩增 hNIS 基因的一部分（核苷酸 1184-1667）。使用地高辛标记的内部 hNIS 特异性寡核苷酸探针（核苷酸 1460-1477）通过 Southern 杂交证实 hNIS 特异性转录物。为了监测 cDNA 的完整性和数量，并排除 DNA 污染和非法转录，所有样品均使用两对跨内含子的引物进行共扩增，这两对引物分别设计用于扩增人 β-肌动蛋白和甲状腺球蛋白基因的片段。使用 Northern 印迹分析，在甲状腺和腮腺中检测到约 4 kb 的 hNIS 转录本，但在广泛的内分泌和非内分泌组织中未检测到。 RT-PCR和Southern杂交显示hNIS基因在甲状腺、唾液腺、腮腺、颌下腺、垂体、胰腺、睾丸、乳腺、胃粘膜、前列腺和卵巢、肾上腺、心脏、胸腺和肺中表达。相比之下，在正常眼眶成纤维细胞、结肠和鼻咽粘膜中未检测到 hNIS 转录本。为了进一步分析腮腺、乳腺和胃粘膜中的hNIS基因序列，使用EXPAND高保真PCR系统和三组重叠的NIS寡核苷酸引物进行扩增和克隆。将所得PCR产物亚克隆到pBluescript-SK II(-)载体中，并对来自每个组织的至少两个独立的cDNA克隆进行自动测序。源自腮腺、乳腺和胃粘膜的 hNIS cDNA 的核苷酸序列与最近发表的源自人甲状腺的 NIS cDNA 序列完全一致。总之，我们的结果表明，几种甲状腺外组织中 hNIS 基因表达水平存在显着差异。尽管 hNIS 在这些组织中的生理作用有待进一步研究，但我们的结果表明，主动转运碘的能力可能是几种分泌和内分泌组织的共同特征。与甲状腺相比，甲状腺外组织（如腮腺、乳腺和胃粘膜）运输和浓缩碘的能力下降，这似乎并不是由 hNIS cDNA 一级结构改变引起的。正常甲状腺外组织中NIS基因表达水平的变异可能是由NIS基因转录活性的差异引起的。进一步的研究将解决这一假设并检查 NIS 基因表达的组织特异性调节机制。
The ability to concentrate iodide is a fundamental property of normally functioning thyroid tissue and represents the first step in the production of thyroid hormones. Iodide uptake has been demonstrated in various extrathyroidal tissues, including salivary gland, gastric mucosa, and lactating mammary gland. Recently, cloning and molecular characterization of the human sodium iodide symporter (hNIS) have been reported; however, the patterns of hNIS gene expression in human tissues have remained unidentified. To examine the profiles of human hNIS gene expression in various normal human tissues, we performed high-stringency Northern blot analysis using a P-32-labeled hNIS-specific complementary DNA (cDNA) probe (nucleotides 1184-1667). To detect rare hNIS transcripts in small tissue samples, RT-PCR was performed with a pair of hNIS-specific oligonucleotide primers designed to amplify a portion (nucleotides 1184-1667) of the hNIS gene. hNIS-specific transcripts were confirmed by Southern hybridization using a digoxigenin-labeled internal hNIS-specific oligonucleotide probe (nucleotides 1460-1477). To monitor cDNA integrity and quantity, and to rule out DNA contamination and illegitimate transcription, all samples were coamplified with two pairs of intron-spanning primers designed to amplify fragments of the human beta-actin and thyroglobulin genes, respectively. Using Northern blot analysis, hNIS transcripts of approximately 4 kb were detected in thyroid gland and parotid gland but not in a broad range of endocrine and nonendocrine tissues. RT-PCR and Southern hybridization revealed hNIS gene expression in thyroid gland, salivary gland, parotid gland, submandibular gland, pituitary gland, pancreas, testis, mammary gland, gastric mucosa, prostate and ovary, adrenal gland, heart, thymus, and lung. By contrast, hNIS transcripts were not detected in normal orbital fibroblasts, colon, and nasopharyngeal mucosa. To further analyze hNIS gene sequences in parotid gland, mammary gland, and gastric mucosa, the EXPAND High Fidelity PCR System and three sets of overlapping NIS oligonucleotide primers were used for amplification and cloning. The resulting PCR products were subcloned into pBluescript-SK II(-)vector, and at least two independent cDNA clones derived from each tissue were subjected to automated sequencing. The nucleotide sequences of hNIS cDNA derived from parotid gland, mammary gland, and gastric mucosa revealed full identity with the recently published human thyroid-derived NIS cDNA sequence. In conclusion, our results demonstrate markedly variable levels of hNIS gene expression in several extrathyroidal tissues. Although the physiological role of hNIS in these tissues awaits further study, our results suggest that the capacity to actively transport iodine may be a feature common to several secretory and endocrine tissues. The diminished capacity to transport and concentrate iodide in extrathyroidal tissues (such as parotid gland, mammary gland, and gastric mucosa), compared with thyroid gland, does not seem to be caused by an altered primary structure of the hNIS cDNA. Variability of NIS gene expression levels in normal extrathyroidal tissues may rather be caused by differences in NIS gene transcriptional activity. Further studies will address this hypothesis and examine the mechanisms of tissue-specific regulation of NIS gene expression.