Reduced mobility of the alternate splicing factor (ASF) through the nucleoplasm and steady state speckle compartments.
Reduced mobility of the alternate splicing factor (ASF) through the nucleoplasm and steady state speckle compartments.
复制标题
通过核质和稳态斑点室降低了替代剪接因子（ASF）的迁移率。
DOI：
10.1083/jcb.150.1.41
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发表时间：
2000-07-10
影响因子：
7.8
通讯作者：
Hendzel, M J
中科院分区：
文献类型：
--
作者：
Kruhlak, M J;Lever, M A;Fischle, W;Verdin, E;Bazett-Jones, D P;Hendzel, M J
细胞核的区隔化现在被认为是影响特定核过程的一个重要调控水平。核空间因子组织和因子运动的机制尚未完全确定。例如，剪接因子已被证明以一种定向的方式作为大型完整结构从集中位点移动到活性转录位点，但剪接因子也被认为以自由扩散状态存在。在本研究中，我们使用延时显微镜和光漂白后荧光恢复技术（FRAP）检测了剪接因子ASF、绿色荧光融合蛋白（ASF - gfp）的运动。我们发现，当ASF-GFP与斑点相关时，其移动速度比自由扩散慢100倍，令人惊讶的是，当它分散在核质中时也是如此。ASF的迁移性与相对不迁移的核结合位点的频繁但短暂的相互作用是一致的。在RNA聚合酶II转录抑制剂的存在下，这种移动性略有增加，并且ASF分子进一步在斑点中富集。我们认为剪接因子的非随机组织反映了相对固定结合位点浓度的空间差异。
Compartmentalization of the nucleus is now recognized as an important level of regulation influencing specific nuclear processes. The mechanism of factor organization and the movement of factors in nuclear space have not been fully determined. Splicing factors, for example, have been shown to move in a directed manner as large intact structures from sites of concentration to sites of active transcription, but splicing factors are also thought to exist in a freely diffusible state. In this study, we examined the movement of a splicing factor, ASF, green fluorescent fusion protein (ASF–GFP) using time-lapse microscopy and the technique fluorescence recovery after photobleaching (FRAP). We find that ASF–GFP moves at rates up to 100 times slower than free diffusion when it is associated with speckles and, surprisingly, also when it is dispersed in the nucleoplasm. The mobility of ASF is consistent with frequent but transient interactions with relatively immobile nuclear binding sites. This mobility is slightly increased in the presence of an RNA polymerase II transcription inhibitor and the ASF molecules further enrich in speckles. We propose that the nonrandom organization of splicing factors reflects spatial differences in the concentration of relatively immobile binding sites.