Institute of Physical and Theoretical Chemistry, Rheinische Friedrich Wilhelms University Bonn, Bonn, Germany.
Biophys J. 2011 Dec 7;101(11):2592-600. doi: 10.1016/j.bpj.2011.10.006.
The activation of STAT transcription factors is a critical determinant of their subcellular distribution and their ability to regulate gene expression. Yet, it is not known how activation affects the behavior of individual STAT molecules in the cytoplasm and nucleus. To investigate this issue, we injected fluorescently labeled STAT1 in living HeLa cells and traced them by single-molecule microscopy. We determined that STAT1 moved stochastically in the cytoplasm and nucleus with very short residence times (<0.03 s) before activation. Upon activation, STAT1 mobility in the cytoplasm decreased ∼2.5-fold, indicating reduced movement of STAT1/importinα/β complexes to the nucleus. In the nucleus, activated STAT1 displayed a distinct saltatory mobility, with residence times of up to 5 s and intermittent diffusive motion. In this manner, activated STAT1 factors can occupy their putative chromatin target sites within ∼2 s. These results provide a better understanding of the timescales on which cellular signaling and regulated gene transcription operate at the single-molecule level.
STAT 转录因子的激活是决定其亚细胞分布和调节基因表达能力的关键因素。然而，目前尚不清楚激活如何影响细胞质和细胞核中单个 STAT 分子的行为。为了研究这个问题，我们在活的 HeLa 细胞中注射了荧光标记的 STAT1，并通过单分子显微镜对其进行追踪。我们发现，在激活之前，STAT1 在细胞质和细胞核中随机移动，停留时间非常短（<0.03 秒）。激活后，细胞质中 STAT1 的迁移率降低了约 2.5 倍，表明 STAT1/importinα/β 复合物向细胞核的运动减少。在细胞核中，激活的 STAT1 显示出明显的跳跃式迁移，停留时间长达 5 秒，并有间歇性的扩散运动。通过这种方式，激活的 STAT1 因子可以在大约 2 秒内占据其假定的染色质靶位点。这些结果更好地理解了细胞信号转导和受调控的基因转录在单分子水平上的作用时间尺度。
