Mechanism of action of STAMP - a new comodulator of glucocorticoid receptors

STAMP是一种转录辅因子，可增强p160辅激活因子SRC-1和TIF 2在糖皮质激素受体调节的基因诱导和抑制中的活性。 在GR、共激活因子TIF 2和STAMP的三元复合物的早期研究中，我们发现每种蛋白质的不同区域可以选择性地不改变、改变一些或改变完整细胞中瞬时转染的报告基因的GR诱导的所有三个参数（Amax、EC 50和PAA）（Awasthi和Simons，2012，Mol Cell Endocrinol，355，121-134）。 然而，STAMP也被其他人描述为具有聚谷氨酰化活性的酪氨酸微管蛋白连接酶样家族成员（TTLL 5）。 此外，我们先前报道了STAMP关于几种细胞类型的生长的类固醇非依赖性活性（He等人，2010，BMC Cancer，10，128）。 本项目目前的目的是确定STAMP的整体动物活性。 这一目标已经通过在同质遗传背景中制备STAMP KO小鼠来实现。 制备了含有破坏的Stamp基因的小鼠，导致蛋白质中间的翻译提前终止。 纯合子靶向突变体（Stamptm/tm）雌性动物表现正常。 雄性Stamptm/tm看起来也很正常，除了它们几乎总是不育。 这种男性不育症似乎是由于精子形成的严重缺陷引起的，这种缺陷导致尾部的高频率卷曲和头部的沿着脱落，同时剩余的完整精子的活力降低。 这些异常与精子尾部轴丝的破坏有关，伴随着一个微管蛋白双联体的丢失以及微管蛋白多聚谷氨酰化的大量减少。 其他结构中的轴丝似乎不受影响，尽管Stamp是TTLL家族中唯一通过添加多聚谷氨酰化所需的第一个谷氨酰基团来有效修饰α-微管蛋白的成员。 尽管睾丸中wt STAMP mRNA的水平比检查的任何其他器官高20倍，但wt与Stamptm/tm雄性的精子发育器官没有明显变化。 这种男性生育能力的缺陷与先前确定的对精子功能重要的24个基因中的任何一个都无关。 因此，STAMP似乎参与了精子成熟和运动的独特途径，这可能与男性生育力有关。 总之，我们使用STAMP KO小鼠来确定KO动物的表型缺陷。 GR/TIF 2/STAMP三元复合物作用的详细研究已经确定GR调节诱导的三个参数（EC 50、PAA和Amax）不协调调节。 相反，这三种蛋白质的不同蛋白质表面可以选择性地调节各个参数，这表明可能存在可以改变一个、两个或所有三个参数的活性的药物。 这些综合研究结果有助于我们在分子水平上定义类固醇激素的作用并了解其在人体生理学中的作用的长期目标。

STAMP is a transcriptional cofactor that augments the activity of the p160 coactivators SRC-1 and TIF2 in glucocorticoid receptor-regulated gene induction and repression. In earlier studies of the ternary complex of GR, the coactivator TIF2, and STAMP, we found that different regions of each protein can selectively alter none, some, or all three parameters (Amax, EC50, and PAA) for GR induction of a transiently transfected reporter gene in intact cells (Awasthi and Simons, 2012, Mol Cell Endocrinol, 355, 121-134). However, STAMP has also been described by others as a tyrosine tubulin ligase-like family member (TTLL5) with polyglutamylation activity. Furthermore, we previously reported steroid-independent activity of STAMP regarding the growth of several cell types (He et al., 2010, BMC Cancer, 10, 128). The current aim of this project is to determine the whole animal activity of STAMP. This aim has been pursued by preparing the STAMP KO mice in a homogeneous genetic background. Mice containing a disrupted Stamp gene, resulting in premature termination of translation in the middle of the protein, were prepared. Homozygous targeted mutant (Stamptm/tm) females appear normal. Stamptm/tm males also seem normal except that they are almost always sterile. This male infertility appears to result from severe defects in sperm formation that cause a high frequency of coiling of the tails and detachment of the heads along with decreased motility in the remaining intact sperm. These abnormalities are associated with disruption of sperm tail axonemes that is accompanied by the loss of one tubulin doublet in conjunction with substantial reduction in α-tubulin polyglutamylation. The axonemes in other structures appear unaffected despite the fact that Stamp is the only member of the TTLL family that efficiently modifies a-tubulin by adding the first glutamyl group needed for polyglutamylation. There is no obvious change in the organs for sperm development of wt vs. Stamptm/tm males despite the levels of wt STAMP mRNA in the testes being 20-fold higher than in any other organ examined. This defect in male fertility is unrelated to any of the 24 genes previously identified as important for sperm function. Thus, STAMP appears to participate in a unique pathway of sperm maturation and motility that may be relevant for male fertility. In summary, we have used STAMP KO mice to determine the phenotypic defects of KO animals. Detailed studies of the action of the ternary complex of GR/TIF2/STAMP have established that the three parameters of GR-regulated induction (EC50, PAA, and Amax) are not coordinately regulated. Instead, different protein surfaces of the three proteins can selectively modulate individual parameters, which indicates that pharmaceuticals may exist that can alter the activities of one, two, or all three parameters. These combined findings contribute to our long-term goal of defining the action of steroid hormones at a molecular level and of understanding their role in human physiology.