Henderson David J P, Miranda Jj L, Emerson Beverly M
The Salk Institute for Biological Studies, La Jolla, CA, USA.
Department of Cellular and Molecular Pharmacology, University of California, San Francisco, CA, USA.
Oncotarget. 2017 Aug 3;8(39):64698-64713. doi: 10.18632/oncotarget.19841. eCollection 2017 Sep 12.
Poly(ADP)ribosylation (PARylation) of the chromatin architectural protein CTCF is critical for CTCF-dependent regulation of chromatin boundary and insulator elements. Loss of CTCF PARylation results in epigenetic silencing of certain tumor suppressor genes through destabilization of nearby chromatin boundaries. We investigated the metabolic and mechanistic processes that regulate PARP-1-mediated CTCF PARylation in human cancer cell lines and discovered a key role for the expression and activity of β-NAD+ salvage enzymes, NAMPT and NMNAT-1. These enzymes are downregulated in cells that exhibit reduced CTCF PARylation, resulting in a decreased concentration of nuclear β-NAD+. In these cells, decreased NMNAT-1 expression is enforced by a proteasome-mediated feedback loop resulting in degradation of NMNAT-1, transcriptional repression of NAMPT, and suppression of PARP-1 activity. Interestingly, dePARylated CTCF is associated in a stable protein complex with PARP-1 and NMNAT-1 in cancer cells harboring silenced tumor suppressor genes. Although the metabolic context in these cells favors suppression of PARP-1 activity, CTCF PARylation can be restored by Protein Kinase C (PKC) signaling. PKC induces dissociation of the catalytically inactive PARP-1/NMNAT-1/CTCF protein complex and phosphorylation of NMNAT-1, which stimulates its proteasome-mediated degradation. Our findings suggest that CTCF PARylation is underpinned by a cellular metabolic context engendered by regulation of the β-NAD+ salvage pathway in which NMNAT-1 acts as a rheostat to control localized β-NAD+ synthesis at CTCF/PARP-1 complexes.
染色质结构蛋白CTCF的多聚（ADP）核糖基化（PARylation）对于依赖CTCF的染色质边界和绝缘子元件调控至关重要。CTCF PARylation的缺失会通过破坏附近染色质边界的稳定性导致某些肿瘤抑制基因的表观遗传沉默。我们研究了在人类癌细胞系中调节PARP-1介导的CTCF PARylation的代谢和机制过程，并发现β-NAD +补救酶NAMPT和NMNAT-1的表达和活性起关键作用。这些酶在CTCF PARylation降低的细胞中下调，导致核β-NAD +浓度降低。在这些细胞中，NMNAT-1表达的降低是由蛋白酶体介导的反馈环强制实现的，导致NMNAT-1降解、NAMPT转录抑制和PARP-1活性抑制。有趣的是，在携带沉默肿瘤抑制基因的癌细胞中，去PARylated的CTCF与PARP-1和NMNAT-1形成稳定的蛋白质复合物。尽管这些细胞中的代谢环境有利于抑制PARP-1活性，但蛋白激酶C（PKC）信号传导可恢复CTCF PARylation。PKC诱导催化无活性的PARP-1 / NMNAT-1 / CTCF蛋白质复合物解离并使NMNAT-1磷酸化，从而刺激其蛋白酶体介导的降解。我们的研究结果表明，CTCF PARylation由β-NAD +补救途径的调节所产生的细胞代谢环境所支撑，其中NMNAT-1作为变阻器来控制CTCF / PARP-1复合物处的局部β-NAD +合成。
