P0021LONG NON-CODING RNAS HOTAIR AND LINC00511 CAN EXPLAIN HUMAN RENAL STEM/PROGENITOR CELLS CAPACITY TO REPAIR DAMAGE INDUCED BY CISPLATIN

Abstract

Abstract Background and Aims The recent discovery of a complex system of human Adult renal stem/progenitor cells (ARPCs) dedicated to the renewal of epithelial renal cells has allowed scientists to better understand how the regenerative process can occur in the adult kidney. ARPCs have a great potential in view of developing future treatments for both acute and chronic renal injury. However, to completely take advantage of their capability it is essential to study the factors regulating the stem cell behavior. Recently, long noncoding RNAs (lncRNAs) have been recognized as a crucial class of gene expression regulators. lncRNAs have several distinguishing characteristics that provide particular regulatory functions, including cell- and tissue-specific expression and the ability to transduce higher-order spatial information. Several lncRNAs modulate some somatic stem cell renewal or differentiation, while others promote a differentiation program. Their functions are often helped by proteic co-factor that convey the ability to activate or repress gene expression or to post-transcriptionally regulate other RNAs. Furthermore, numerous lncRNAs have been shown to act as competing endogenous RNAs, where the lncRNAs are proposed to bind to and compete miRNAs away from cognate mRNA targets. The aim of the study was to evaluate the basal expression profile of lncRNA expressed specifically in ARPCs. Method lncRNA expression profile was obtained from ARPCs and renal proximal tubular cells (RPTECs) alone or following cisplatin damage induction. We used Agilent SurePrint G3 Human Gene Expression Microarrays providing comprehensive coverage of genes and transcripts using the latest annotation databases. Genespring and R software were used for the analysis. lncRNA expression was validated by Real-time PCR. CrispR/Cas9 system has been used to knock-down specific lncRNA. Results We compared lncRNA expression between ARPCs and RPTECs: 45 lncRNA were differently modulated in ARPCs vs RPTECs (Fold change 1.5; FDR <0.05). In particular, we found 13 lncRNA upregulated and 32 lncRNA downregulated. Classification analysis showed that most of lncRNA modulated in ARPCs interfere with WNT signaling pathway, immune cell activation, and G-protein signaling pathway. Moreover, the overrepresentation test showed their involvement in calcium-mediated signaling, cell cycle and protein glycosylation processes (p<0.005). Among most significantly modulated lincRNAs we found HOTAIR and CCND2-AS1 that were upregulated in ARPCs compared to RPTECs. When we knocked-down the HOTAIR lncRNA ARPCs were not able to normally proliferate in cell culture. However, following cisplatin damage induced in RPTECs, ARPCs upregulated the LINC00511 and the miR210-HG. LINC00511 binds histone methyltransferase enhancer of zeste homolog 2 (EZH2, the catalytic subunit of the polycomb repressive complex 2 - PRC2), a highly conserved protein complex that regulates gene expression by methylating lysine 27 on histone H3. It acts as a modular scaffold of EZH2/PRC2 complexes, coordinates their localization, and specifies the histone modification pattern on the target genes, including p57. LINC00511 can affect cell proliferation, invasiveness, and apoptosis. Conclusion ARPCs express specific lncRNAs that could explain some of the ARPC stemness properties and their capacity to repair damage induced by cisplatin. Our findings suggest that lncRNA may represent a novel therapeutic target in acute and chronic renal injury.