Identification of a Novel Noncoding RNA Transcript TISPL Upregulated by Stressors that Stimulate ATF4

Abstract

Cells sense, respond, and adapt to environmental conditions that cause stress. In a previous study using HeLa cells, we isolated reporter cells responding to the endoplasmic reticulum (ER) stress inducers, thapsigargin and tunicamycin, using a highly sensitive promoter trap vector system. Splinkerette PCR and 5′ rapid amplification of cDNA ends (5′ RACE) identified a novel transcript that is upregulated by ER stress. Its endogenous expression increased approximately 10-fold in response to thapsigargin and tunicamycin within 1 h, but was down-regulated after 4 h. Because the transcript starts from an intron of a long noncoding RNA known as LINC-PINT, we designated the newly identified transcript TISPL (transcript induced by stressors from LINC-PINT locus). TISPL was also expressed under several other stress conditions. It was particularly increased >10-fold upon glucose starvation and 7-fold by arsenite exposure. Furthermore, in silico analyses, including a ChIP-atlas search revealed that there is an ATF4-binding region with a c/ebp-Atf response element (CARE) downstream of the transcription start site of TISPL. Based on these results, we hypothesized that TISPL may be induced by the phospho-eIF2α and ATF4- axis of the integrated stress response pathway, which is known to be activated by the stress conditions listed above. As expected, knockout of ATF4 abolished the stress-induced upregulation of TISPL. Our results indicate that TISPL may be a useful biomarker for detecting stress conditions that activate ATF4. Our highly sensitive trap vector system proved beneficial in discovering new biomarkers.

Note:
Funding declaration: This study was supported in part by CBBD-OIL and the translational research programs from Fukushima Prefecture.

Conflict of Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Keywords: endoplasmic reticulum stress, long noncoding RNA, integrated stress response, ATF4, Biomarker