Schussler N, Boularand S, Li J Y, Peillon F, Mallet J, Biguet N F
INSERM U. 223, Faculté de Médecine Pitié-Salpêtrière, Paris, France.
J Neurosci Res. 1995 Dec 15;42(6):846-54. doi: 10.1002/jnr.490420613.
Several regulatory neurofactors, classically associated with the hypothalamus, may be synthesized in the anterior pituitary (AP). Dopamine (DA) is the main prolactin-inhibiting factor. Its de novo synthesis in the normal AP has not been proved, although the TH transcript has been previously demonstrated by RT/PCR in the AP. We investigated tyrosine hydroxylase (TH) gene expression at both the protein and mRNA levels in the AP of normal random cycling female rats and in a catecholaminergic tissue, the adrenal gland (AG). The Western blot analysis of AP homogenates revealed two immunoreactive forms of TH in the AP, both differing from the TH present in the AG. RT/PCR products from AP and AG mRNA were subcloned and sequenced. In addition to the full-length form, we identified two TH transcripts generated by alternative splicing either involving the use of a new alternate splice-donor site within exon 2 or skipping exon 11. The form lacking exon 11 was not isolated from the AG. In the AP, all three forms were present. Although the AP contained the full-length TH mRNA, the expected size protein was not detected. Thus, there is alternative splicing of the TH primary transcript, and putative additional post-translational regulation may yield TH proteins with no enzymatic activity, at least in non-catecholaminergic tissues.
几种传统上与下丘脑相关的调节性神经因子可能在前垂体（AP）中合成。多巴胺（DA）是主要的催乳素抑制因子。尽管之前通过逆转录/聚合酶链反应（RT/PCR）已在前垂体中证实了酪氨酸羟化酶（TH）转录本的存在，但其在正常前垂体中的从头合成尚未得到证实。我们在正常随机发情的雌性大鼠的前垂体以及一种儿茶酚胺能组织——肾上腺（AG）中，从蛋白质和mRNA水平研究了酪氨酸羟化酶（TH）基因的表达。前垂体匀浆的蛋白质免疫印迹分析显示前垂体中有两种TH免疫反应形式，这两种形式均与肾上腺中存在的TH不同。对来自前垂体和肾上腺mRNA的RT/PCR产物进行亚克隆和测序。除了全长形式外，我们还鉴定出两种由可变剪接产生的TH转录本，一种涉及外显子2内新的可变剪接供体位点的使用，另一种是跳过外显子11。缺少外显子11的形式未从肾上腺中分离出来。在前垂体中，三种形式均存在。尽管前垂体中含有全长TH mRNA，但未检测到预期大小的蛋白质。因此，TH初级转录本存在可变剪接，并且推测至少在非儿茶酚胺能组织中可能存在额外的翻译后调控，从而产生无酶活性的TH蛋白。
