The Spt6-tSH2 domain coordinates functional transitions of the RNA Polymerase II CTD during initiation and elongation.

The conserved transcription elongation factor Spt6 makes several contacts with the RNA Polymerase II (RNAPII) complex, including a high-affinity interaction between the Spt6 tandem SH2 domain (Spt6-tSH2) and phosphorylated residues in Rpb1 in a region between the catalytic core and the heptad repeats of its C-terminal domain (CTD). This interaction contributes to the global occupancy of Spt6 within transcription units, suggesting that it has a general role in tethering Spt6 to the elongation complex. However, we show here that disrupting this binding caused increases in some transcripts, revealing specific functional roles in regulating the expression of subsets of genes. These included loci whose regulation involves differential transcription start site selection, early termination of transcription, or efficient restoration of chromatin integrity after transcription. Loss of this interaction also caused a defect in splicing, and apparent pausing of RNAPII progression in regions requiring more complex processing of excised introns. The results support a global role for the Spt6-tSH2:Rpb1 interaction as one of several means of stabilizing the association of Spt6 with RNAPII, but they also reveal local functions at specific sites, especially those where dynamic decisions regarding initiation or termination are made, or where changes in the configuration of associated factors occur. We therefore propose that the Spt6-tSH2:Rpb1 interaction can provide a conduit for communication between RNAPII and the elongation factor function of Spt6, or with other factors associated with the Rpb1 CTD, supporting appropriate elongation through challenging templates and efficient co-transcriptional processing.保守的转录延伸因子Spt6可与RNA聚合酶II（RNA Polymerase II，RNAPII）复合物形成多重相互作用，其中包括Spt6串联SH2结构域（Spt6-tSH2）与Rpb1催化核心与其羧基末端结构域（C-terminal domain，CTD）七肽重复序列之间区域内的磷酸化残基之间的高亲和力结合。该相互作用有助于Spt6在转录单元内的全局占据，提示其在将Spt6锚定至转录延伸复合物的过程中发挥普遍作用。然而本研究显示，破坏该结合会导致部分转录本水平升高，揭示了其在调控特定基因子集表达中的特异性功能。此类靶位点包括那些调控涉及差异化转录起始位点选择、转录提前终止，或转录后有效恢复染色质完整性的基因座。该相互作用的缺失还会引发剪接缺陷，并使需要对剪接内含子进行更复杂加工的区域内的RNAPII行进出现明显暂停。研究结果表明，Spt6-tSH2与Rpb1的相互作用作为稳定Spt6与RNAPII结合的多种途径之一，发挥了全局层面的功能，但同时也揭示了其在特定位点的局部功能，尤其是那些涉及转录起始或终止的动态决策，或相关因子构象发生改变的位点。因此，我们提出Spt6-tSH2:Rpb1相互作用可作为RNAPII与Spt6延伸因子功能之间，或与Rpb1 CTD结合的其他因子之间的通信通路，助力其在复杂模板上顺利完成转录延伸，并实现高效的共转录加工。