| 摘要: | 甘丙胺激素是一種C端有乙醯化的神經內泌素，已知其具有調控脊椎動物行為與食慾的功能。為了對爬行類腦垂體甘丙胺激素的物種歧異度與基因之演化有更進一步之了解，本研究以中華鱉為實驗動物，利用快速核酸兩端放大之技術，定序腦垂體甘丙胺激素互補核苷酸序列，所得的序列共有672個核苷酸，5’-未轉錄區99 個；前原甘丙胺激素有324個核苷酸；3’-未轉錄區有249 個。前原甘丙胺激素有108個胺基酸；本體為29個胺基酸，與其他脊椎動物之間的同源性為72% ~ 96%。 半定量聚合酶連鎖反應實驗結果顯示，前原甘丙胺激素於腦、下丘腦、胃、肝臟、胰臟、睪丸、卵、子宮、大腸與小腸皆有表現，而在心臟、脾臟、肌肉與脂肪組織則無法偵測到其表現。雌性前原甘丙胺激素之表現量似乎高於雄性。 離體腦垂體組織培養研究顯示：雌二醇(10-6M~10-8M)、性釋素(10-7M~10-9M) 隨著濃度增加會明顯促進前原甘丙胺激素之表現；而瘦素(10-6M~10-8M)與神經月太 Y(10-6M~10-8M)隨著濃度增加，抑制甘丙胺激素基因表現之情形更趨明顯。 綜觀上述結果顯示脊椎動物之前原甘丙胺激素具高度之保守性，且與雌二醇、性釋素、瘦素及神經月太 Y在中華鱉腦垂體的生理功能上具交互作用。 Galanin is a C-terminally-amidated neurohormone known to regulate various aspects of behavior and appetite of vertebrates. To better understanding the phylogenetic diversity and evolution of galanin genes in reptilie, we here conducted an analysis of galanin cDNA from Chinese mud turtle pituitary. Using 3’- and 5’- RACE techniques, we have isolated and sequenced a novel cDNA derived from pituitary total RNA. Sequence analysis revealed a 672-bp cDNA containing a 99-bp 5’-untranslated region, a 324-bp preprogalanin coding region, and a 249-bp 3’-untranslated region. The deduced protein is a 108-amino acid preprogalanin molecule with [Ser23]-galanin-29 at the C-terminus of the coding region, making its homology to other vertebrates in the range of 72%?96%. Semiquantitative PCR revealed that preprogalanin was expressed in the brain, hypothalamus, pituitary, stomach, liver, pancreas, testis, ovary, uterus, small intestine and large intestine, but undetected in the heart, spleen, muscle and adipose tissue. Females seemed to express more preprogalanin than males. In vitro pituitary tissue culture study indicated that E2 (10-6M~10-8M) or GnRH (10-7M~10-9M) treatment significantly increased the levels of preprogalaninin a dose- dependent manner. However, Lep (10-6M~10-8M) or NPY (10-7M~10-9M) treatment decreased the levels of preprogalanin also in a dose-dependent manner. These results suggest that preprogalanin is conserved in vertebrates and that a physiological role of galanin interacting with E2, GnRH, Lep and/or NPY may exist in the pituitary of mud turtle. |
