Department of Microbiology and Immunology, Weill Cornell Medical College, New York, New York 10065, USA.
Gerstner Sloan Kettering Graduate School of Biomedical Sciences, New York, New York 10065, USA.
RNA. 2020 Oct;26(10):1334-1344. doi: 10.1261/rna.076463.120. Epub 2020 Jun 16.
Fission yeast Erh1 exists in a complex with RNA-binding protein Mmi1. Deletion of up-regulates the phosphate homeostasis gene , which is normally repressed by transcription in of a 5' flanking lncRNA. Here we present evidence that de-repression of by eΔ is achieved through precocious 3'-processing/termination of lncRNA synthesis, to wit: (i) Δ does not affect the activity of the or promoters per se; (ii) de-repression by Δ depends on CPF (cleavage and polyadenylation factor) subunits Ctf1, Dis2, Ssu72, Swd22, and Ppn1 and on termination factor Rhn1; (iii) de-repression requires synthesis by the Asp1 IPP kinase of inositol 1-pyrophosphates (1-IPPs); (iv) de-repression is effaced by mutating Thr4 of the RNA polymerase II CTD to alanine; and (v) Δ exerts an additive effect on de-repression in combination with mutating CTD Ser7 to alanine and with deletion of the IPP pyrophosphatase Aps1. These findings point to Erh1 as an antagonist of lncRNA termination in the axis. In contrast, in Δ cells there is a reduction in mRNA and increase in the formation of a read-through transcript, consistent with Mmi1 being an agonist of termination. We envision that Erh1 acts as a brake on Mmi1's ability to promote CPF-dependent termination during lncRNA synthesis. Consistent with this idea, Δ de-repression of was eliminated by mutating the Mmi1-binding sites in the lncRNA.
裂殖酵母 Erh1 与 RNA 结合蛋白 Mmi1 存在复合物。缺失上调磷酸盐稳态基因，该基因通常受转录抑制因子在 5'侧翼 lncRNA 的 区域抑制。在这里，我们提供的证据表明，eΔ 通过提前进行 3'加工/终止 lncRNA 合成来解除对 的抑制：（i）Δ 本身不影响 或 启动子的活性；（ii）Δ 对 的去抑制依赖于 CPF（切割和多聚腺苷酸化因子）亚基 Ctf1、Dis2、Ssu72、Swd22 和 Ppn1 和终止因子 Rhn1；（iii）去抑制需要由 Asp1 IPP 激酶合成肌醇 1-焦磷酸（1-IPPs）；（iv）将 RNA 聚合酶 II CTD 的 Thr4 突变为丙氨酸可消除去抑制；（v）Δ 与 CTD Ser7 突变为丙氨酸突变和 IPP 焦磷酸酶 Aps1 缺失组合对 去抑制具有附加效应。这些发现表明 Erh1 作为 lncRNA 终止在 轴线上的拮抗剂。相比之下，在 Δ 细胞中，减少了 mRNA 的表达并增加了 通读转录物的形成，这与 Mmi1 作为 终止的激动剂一致。我们设想 Erh1 作为 Mmi1 促进 lncRNA 合成过程中 CPF 依赖性终止的能力的制动。与这一观点一致，通过突变 lncRNA 中的 Mmi1 结合位点消除了 Δ 对 的去抑制。
