Correlations of three-dimensional motion of chromosomal loci in yeast revealed by the double-helix point spread function microscope.
Correlations of three-dimensional motion of chromosomal loci in yeast revealed by the double-helix point spread function microscope.
复制标题
DOI：
10.1091/mbc.e14-06-1127
复制
发表时间：
2014-11-05
影响因子：
3.3
通讯作者：
Moerner WE
中科院分区：
文献类型：
--
作者：
Backlund MP;Joyner R;Weis K;Moerner WE
双螺旋点扩散函数显微镜用于在三个维度上追踪活酵母细胞中单对荧光标记的染色体位点。在抑制条件下，二倍体细胞中的 GAL 基因座对之间观察到增强的速度互相关性，并且发现普遍存在的次扩散指数接近 0.6-0.75。单粒子追踪已被应用于研究活细胞中的染色质运动，揭示了基因组材料的丰富动态行为，这些行为曾经被认为在整个细胞周期的大部分时间中相对静态。在这里，我们使用双色三维（3D）双螺旋点扩散函数显微镜来研究相同或不同芽殖酵母染色体上两个荧光标记基因位点之间运动的相关性。我们在激活和抑制条件下对二倍体细胞中 GAL 基因座的两个副本进行了快速 (10 Hz) 3D 跟踪。作为对照，我们追踪了同一染色体上不同间隔的基因座对，以及不同染色体上的转录正交基因。我们发现，在抑制条件下，GAL 位点表现出比激活条件下显着更高的速度互相关性。这种相对增加具有潜在的重要生物学意义，因为它可能表明通过共享沉默因子进行耦合或在激活时与解耦机制相关。我们还发现，在研究的时间尺度（∼0.1-30 s）上，基因座移动的亚扩散均方位移指数显着高于先前报道的，这对聚合物理论在真核生物染色质运动中的应用具有影响。
The double-helix point spread function microscope is used to track single pairs of fluorescently labeled chromosomal loci in live yeast cells in three dimensions. Enhanced velocity cross-correlations are observed between pairs of GAL loci in diploid cells under repressive conditions, and ubiquitous subdiffusive exponents are found to be near 0.6–0.75. Single-particle tracking has been applied to study chromatin motion in live cells, revealing a wealth of dynamical behavior of the genomic material once believed to be relatively static throughout most of the cell cycle. Here we used the dual-color three-dimensional (3D) double-helix point spread function microscope to study the correlations of movement between two fluorescently labeled gene loci on either the same or different budding yeast chromosomes. We performed fast (10 Hz) 3D tracking of the two copies of the GAL locus in diploid cells in both activating and repressive conditions. As controls, we tracked pairs of loci along the same chromosome at various separations, as well as transcriptionally orthogonal genes on different chromosomes. We found that under repressive conditions, the GAL loci exhibited significantly higher velocity cross-correlations than they did under activating conditions. This relative increase has potentially important biological implications, as it might suggest coupling via shared silencing factors or association with decoupled machinery upon activation. We also found that on the time scale studied (∼0.1–30 s), the loci moved with significantly higher subdiffusive mean square displacement exponents than previously reported, which has implications for the application of polymer theory to chromatin motion in eukaryotes.