Naslov
WISP1-AS1, a long noncoding RNA, upregulated in human renal cells exposed to the mycotoxin ochratoxin A and in human renal cancer cells

Autori
Polović, Mirjana

Vrsta, podvrsta i kategorija rada
Ocjenski radovi, doktorska disertacija

Mjesto
Halle (Saale), Njemačka

Datum
31.07

Godina
2020

Stranica
140

Mentor
Michael Gekle, Gerald Schwerdt

Ključne riječi
lncRNA, antisense, WISP1, ochratoxin A, antiapoptotic, renal cell

Sažetak
INTRODUCTION: Ochratoxin A (OTA) is a worldwide spread mycotoxin that contaminates our food. Detrimental effects of OTA on human health are evident by damage to renal cells, causing nephropathies. Long non-coding RNAs (lncRNAs) are also implicated in the pathogenesis of renal nephropathies and cancers. Herein I report on a novel lncRNA, WISP1-AS1, an antisense transcript in the WISP1 locus. OTA is the only known inducer of WISP1-AS1. However, WISP1-AS1 functions in OTA-treated cells were undiscovered. This research was set to investigate WISP1-AS1 characteristics and functions in OTA toxicity and renal cancer cells and to understand the molecular mechanisms behind it. METHODS: WISP1-AS1 expression in normal and cancer renal cells was investigated by PCR methods and Northern blot. WISP1-AS1 knockdown was established by LNA™ GapmeR antisense oligonucleotides (ASOs). By RNAsequencing, influence of WISP1-AS1 on transcriptome was evaluated. Changes in transcription factor (TF) activities were analyzed by promoter activity assays. Glucose consumption and lactate production were determined by specific enzymatic assays. The effect of WISP1-AS1 on cell death (apoptosis and necrosis) was assessed by measuring caspase activities and lactate dehydrogenase (LDH) release. Nuclear fragmentation and mitochondrial membrane potential were analyzed by Cytation 3 Cell Imaging Multi-Mode Reader. RESULTS: WISP1-AS1 transcribes in antisense in comparison with WISP1 mRNA. Expression in antisense was confirmed by Northern blot. WISP1-AS1 transcription is completely inside the WISP1 locus and does not suppress WISP1 mRNA. WISP1-AS1 influences the activity of certain TFs. Promoter activity assays confirmed that the activity of TF EGR-1 was increased and E2F activity was suppressed in a WISP1-AS1-dependent manner. WISP1-AS1 expression was elevated in renal cancer cells and OTA further upregulated it. High WISP1-AS1 expression in renal cancer cells without OTA treatment indicates its implication in carcinogenesis. Functional experiments demonstrate that WISP1-AS1 might direct glucose towards mitochondrial respiration to obtain ATP. In WISP1-AS1 presence, caspase-3 activity was significantly low and the GAS6 gene, involved in caspase-3 suppression, was upregulated. CONCLUSIONS: WISP1-AS1 is an antisense noncoding transcript. WISP1-AS1 upregulates the activity of TF EGR-1 and suppresses the activity of E2F. It lowers caspase-3 activity and acts antiapoptotic, possibly by upregulating GAS6. In WISP1-AS1 presence, OTA-treated cells maintain normal energy metabolism.

Izvorni jezik
Engleski

Znanstvena područja
Biologija

POVEZANOST RADA