Pathogenesis of a transcription factor disease, combined pituitary hormone defficiency

Pit-1是一种垂体特异性转录因子，可激活生长激素（GH）、催乳素（PRL）、促甲状腺激素β（TSHb）基因的表达。突变的Pit-1可能会导致这三种激素的缺乏（联合垂体激素缺乏症），并且有几种Pit-1突变体确实被报道为联合垂体激素缺乏症的原因。我们分析了两个突变的Pit-1 s（P24 L和R271 W）的功能，这两个突变都被报道为垂体激素联合缺乏症的原因。在P24 L中，24位的脯氨酸被赖氨酸取代，而在R271 W中，271位的精氨酸被色氨酸取代。P24 L与Pit-1结合元件具有正常的结合活性，但与CBP失去结合。缺失构建体和双杂交分析表明，反式激活和POU结构域是必需的结合的Pit-1 CBP和CH 1和CH 3结构域的CBP的结合Pit-1。此外，P24 L，不像野生型Pit-1，不激活cAMP诱导的报告质粒含有Pit-1结合元件的表达。腺病毒E1 a抑制CBP，降低Pit-1和cAMP诱导的报告质粒的表达。这些发现表明CBP在Pit-1和cAMP诱导的基因表达中起作用。据报道，另一个Pit-1突变体R271 W是导致垂体激素联合缺乏症的原因。然而，在我们的研究中，R271 W和野生型Pit-1在不同的细胞和实验条件下以相似的程度增加GH、PRL报告质粒的表达。与我们的数据一致，有一份报告显示R271 W受试者没有任何垂体激素联合缺乏症的表型。需要进一步分析以得出结论，R271 W是垂体激素联合缺乏症的致病突变体。

A pituitary-specific transcription factor, Pit-1 activates growth hormone (GH), prolactin (PRL), TSHb gene expressions. Mutant Pit-1 may lead to the deficiency of the three hormones (combined pituitary hormone deficiency), and several Pit-1 mutants, indeed, were reported as a cause of combined pituitary hormone deficiency. We analyzed the function of two mutant Pit-1s (P24L and R271W), both of which have been reported as a cause of combined pituitary hormone deficiency. In P24L, proline at 24 is replaced with lysine, and arginine at 271 is replaced with tryptophan in R271W. The P24L had normal binding activity to Pit-1 binding element, but lost the binding to CBP. Deletion construct and two hybrid assay revealed that both transactivation and POU domains were required for the binding of Pit-1 to CBP and that CH1 and CH3 domains were needed for the binding of CBP to Pit-1. Furthermore, P24L, unlike wild type Pit-1, did not activate cAMP-induced expression of a reporter plasmid containing Pit-1 binding elements. Adenovirus E1a, which inhibit CBP, lowered Pit-1 and cAMP-induced expression of the reporter plasmid. These findings suggested that CBP plays a role in Pit-1 and cAMP-induced gene expression. It Is reported that another Pit-1 mutant R271W is responsible for combined pituitary hormone deficiency. However, R271W and wild type Pit-1 increased expression of GH, PRL reporter plasmids to similar extent in various cells and experimental conditions in our study. Consistent with our data, there is a report showing that a subject with R271W did nof have any phenotypes of combined pituitary hormone deficiency. Further analysis will be required for concluding that R271W is a causative mutant for combined pituitary hormone deficiency.