Goyal Ashish, Fiškin Evgenij, Gutschner Tony, Polycarpou-Schwarz Maria, Groß Matthias, Neugebauer Julia, Gandhi Minakshi, Caudron-Herger Maiwen, Benes Vladimir, Diederichs Sven
Division of RNA Biology & Cancer, German Cancer Research Center (DKFZ), 69120 Heidelberg, Germany.
Institute of Pathology, University Hospital Heidelberg, 69120 Heidelberg, Germany.
Nucleic Acids Res. 2017 Dec 1;45(21):12496-12508. doi: 10.1093/nar/gkx952.
Long non-coding RNAs (lncRNAs) have been proven to play important roles in diverse cellular processes including the DNA damage response. Nearly 40% of annotated lncRNAs are transcribed in antisense direction to other genes and have often been implicated in their regulation via transcript- or transcription-dependent mechanisms. However, it remains unclear whether inverse correlation of gene expression would generally point toward a regulatory interaction between the genes. Here, we profiled lncRNA and mRNA expression in lung and liver cancer cells after exposure to DNA damage. Our analysis revealed two pairs of mRNA-lncRNA sense-antisense transcripts being inversely expressed upon DNA damage. The lncRNA NOP14-AS1 was strongly upregulated upon DNA damage, while the mRNA for NOP14 was downregulated, both in a p53-dependent manner. For another pair, the lncRNA LIPE-AS1 was downregulated, while its antisense mRNA CEACAM1 was upregulated. To test whether as expected the antisense genes would regulate each other resulting in this highly significant inverse correlation, we employed antisense oligonucleotides and RNAi to study transcript-dependent effects as well as dCas9-based transcriptional modulation by CRISPRi/CRISPRa for transcription-dependent effects. Surprisingly, despite the strong stimulus-dependent inverse correlation, our data indicate that neither transcript- nor transcription-dependent mechanisms explain the inverse regulation of NOP14-AS1:NOP14 or LIPE-AS1:CEACAM1 expression. Hence, sense-antisense pairs whose expression is strongly-positively or negatively-correlated can be nonetheless regulated independently. This highlights the requirement of individual experimental studies for each antisense pair and prohibits drawing conclusions on regulatory mechanisms from expression correlations.
长链非编码RNA（lncRNA）已被证明在包括DNA损伤反应在内的多种细胞过程中发挥重要作用。近40%的注释lncRNA以与其他基因相反的方向转录，并且常常通过转录本或转录依赖性机制参与对这些基因的调控。然而，基因表达的负相关是否通常意味着基因之间存在调控相互作用仍不清楚。在这里，我们分析了肺癌和肝癌细胞在暴露于DNA损伤后lncRNA和mRNA的表达情况。我们的分析揭示了两对mRNA-lncRNA正义-反义转录本在DNA损伤后呈反向表达。lncRNA NOP14-AS1在DNA损伤后强烈上调，而NOP14的mRNA则下调，两者均以p53依赖性方式发生。对于另一对，lncRNA LIPE-AS1下调，而其反义mRNA CEACAM1上调。为了测试反义基因是否如预期那样相互调控从而导致这种高度显著的负相关，我们使用反义寡核苷酸和RNA干扰来研究转录本依赖性效应，并使用基于dCas9的CRISPRi/CRISPRa转录调控来研究转录依赖性效应。令人惊讶的是，尽管存在强烈的刺激依赖性负相关，但我们的数据表明，转录本依赖性和转录依赖性机制均无法解释NOP14-AS1:NOP14或LIPE-AS1:CEACAM1表达的反向调控。因此，表达呈强正相关或负相关的正义-反义对仍可能独立调控。这突出了对每一对反义基因进行单独实验研究的必要性，并禁止从表达相关性得出调控机制的结论。
