Schmidt-Arras Dirk, Böhmer Sylvia-Annette, Koch Sina, Müller Jörg P, Blei Lutz, Cornils Hauke, Bauer Reinhard, Korasikha Sridhar, Thiede Christian, Böhmer Frank-D
Institute of Molecular Cell Biology, Center for Molecular Biomedicine, Friedrich Schiller University Jena, Jena, Germany.
Blood. 2009 Apr 9;113(15):3568-76. doi: 10.1182/blood-2007-10-121426. Epub 2009 Feb 9.
The mechanism of cell transformation by Fms-like tyrosine kinase 3 (FLT3) in acute myeloid leukemia (AML) is incompletely understood. The most prevalent activated mutant FLT3 ITD exhibits an altered signaling quality, including strong activation of the STAT5 transcription factor. FLT3 ITD has also been found partially retained as a high-mannose precursor in an intracellular compartment. To analyze the role of intracellular retention of FLT3 for transformation, we have generated FLT3 versions that are anchored in the perinuclear endoplasmic reticulum (ER) by appending an ER retention sequence containing a RRR (R3) motif. ER retention of R3, but not of corresponding A3 FLT3 versions, is shown by biochemical, fluorescence-activated cell sorting, and immunocytochemical analyses. ER anchoring reduced global autophosphorylation and diminished constitutive activation of ERK1/2 and AKT of the constitutively active FLT3 versions. ER anchoring was, however, associated with elevated signaling to STAT3. Transforming activity of the FLT3 D835Y mutant was suppressed by ER anchoring. In contrast, ER-anchored FLT3 ITD retained STAT5-activating capacity and was transforming in vitro and in vivo. The findings highlight another aspect of the different signaling quality of FLT3 ITD: It can transform cells from an intracellular location.
急性髓系白血病（AML）中Fms样酪氨酸激酶3（FLT3）导致细胞转化的机制尚未完全明确。最常见的激活型突变体FLT3 ITD表现出信号传导质量的改变，包括STAT5转录因子的强烈激活。FLT3 ITD还被发现部分保留为细胞内区室中的高甘露糖前体。为了分析FLT3细胞内滞留对转化的作用，我们通过附加包含RRR（R3）基序的内质网（ER）滞留序列，生成了锚定在核周内质网的FLT3变体。生化、荧光激活细胞分选和免疫细胞化学分析表明，R3变体而非相应的A3 FLT3变体能够在内质网中滞留。内质网锚定降低了组成型激活的FLT3变体的整体自磷酸化水平，并减弱了ERK1/2和AKT的组成型激活。然而，内质网锚定与STAT3信号增强有关。内质网锚定抑制了FLT3 D835Y突变体的转化活性。相比之下，内质网锚定的FLT3 ITD保留了激活STAT5的能力，并在体外和体内均具有转化能力。这些发现突出了FLT3 ITD不同信号传导质量的另一个方面：它可以从细胞内位置转化细胞。
