Molecular mechanism of growth and differentiation of neutrophilic granulocyte mediated by G-CSF

我们的目标是了解由粒细胞集落刺激因子（G-CSF）介导的嗜中性粒细胞生长和分化的分子机制。为此，我们研究了骨髓特异性锌指蛋白MZF-2，它在粒细胞分化中起作用。对小鼠MZF-2的突变分析表明，MZF-2蛋白在N-末端携带转录抑制区，在分子中间携带反式激活结构域（TA结构域，50个氨基酸残基），在C-末端区域携带DNA结合锌指结构域。仅含有TA结构域的突变蛋白的过表达抑制了MZF-2介导的转录激活，表明TA结构域招募骨髓特异性转录共激活因子。已知G-CSF可以激活MAP激酶途径。为了阐明MAP激酶途径在G-CSF信号传导中的作用，我们分析了蛋白激酶MNK 1的体内功能，MNK 1是MAP激酶之一， 关于我们 激酶靶点。组成型活性MNK 1突变体或显性阴性突变体的表达分别导致真核翻译起始因子4 E（eIF-4 E）在Ser 209处的组成型磷酸化或组成型去磷酸化，表明eIF-4 E是MNK 1的生理靶点。用各种生长或应激刺激物处理细胞迅速诱导eIF-4 E通过MNK 1磷酸化，表明MNK 1在调节蛋白质合成中的生理功能。细胞生长受称为Cdks的细胞周期调节蛋白激酶家族调节。通过对细胞周期蛋白E/Cdk 2底物的磷酸化筛选，我们鉴定了一种新的细胞周期蛋白E/Cdk 2底物PRC 1，它与芽殖酵母蛋白Ase 1 p具有序列同源性。PRC 1蛋白在胞质分裂过程中定位于细胞中间体。显微注射抗PRC 1抗体到HeLa细胞中阻断细胞分裂，但不核分裂，表明PRC 1在胞质分裂过程中的功能作用。少

Our goal is to understand the molecular mechanism of growth and differentiation of neutrophilic granulocytes, which are mediated by granulocyte colony-stimulating factor (G-CSF). For this purpose, we have investigated the myeloid-specific zinc-finger protein MZF-2 that plays a role in granulocyte differentiation. Mutational analysis of mouse MZF-2 demonstrated that MZF-2 protein carries a transcription-inhibitory region at the N-terminus, a transactivation domain (TA domain, 50 amino-acid residues) in the middle of the molecule, and a DNA-binding zinc-finger domains in the C-terminal region. Overexpression of a mutant protein containing only the TA domain inhibited the MZF-2-mediated transcriptional activation, suggesting that the TA domain recruits a myeloid-specific transcriptional coactivator. G-CSF is known to activate the MAP kinase pathway. To elucidate the role of the MAP kinase pathway in G-CSF signalling, we analysed in vivo function of the protein kinase MNK1, one of the MAP … More kinase targets. Expression of a constitutively active MNK1 mutant or a dominant-negative mutant resulted in constitutive phosphorylation or constitutive dephosphorylation of eukaryotic translation initiation factor 4E (eIF-4E) at Ser209, respectively, indicating that eIF-4E is a physiological target for MNK1. Treatment of cells with various growth or stress stimuli rapidly induces phosphorylation of eIF-4E through MNK1, suggesting a physiological function of MNK1 in the regulation of protein synthesis. Cell growth is regulated by a family of cell-cycle regulating protein kinases called Cdks. By a phosphorylation screen for cyclin E/Cdk2 substrates, we have identified a novel cyclin E/Cdk2 substrate, PRC1, which has sequence homology to the budding yeast protein Ase1p. PRC1 protein localized to the cell midbody during cytokinesis. Microinjection of anti-PRC1 antibodies into HeLa cells blocked cellular cleavage, but not nuclear division, indicating a functional role for PRC1 in the process of cytokinesis. Less