TY - JOUR

T1 - Human cell lines expressing modified smallnucleolar RNAs as a model for splicinginhibition

AU - Матвеева, Анна Геннадьевна

AU - Zhuravlev, Evgenii S.

AU - Semenov, Dmitriy V.

AU - Vlassov, Valentin V.

N1 - Human cell lines expressing modified smallnucleolar RNAs as a model for splicinginhibition / A. Matveeva, E. Zhuravlev, D. Semenov, V. Vlassov, G. Stepanov // FEBS Open Bio. - 2023. - Т. 13. № S2. - C. 154

PY - 2023

Y1 - 2023

N2 - Precursor mRNA (pre-mRNA) splicing presents one of the cru-cial stages of mRNA maturation in eukaryote cells. Alternativesplicing allows the formation of multiple isoforms thus providingtranscriptomic diversity and the assortment of biologicalfunctions. Hence, there is a need for the development of splicingmodulation tools in order to fine-tune gene expression withoutdisrupting gene structure. The current study focuses on smallnucleolar RNAs (snoRNAs) as a possible splicing regulator. Ineukaryote cells these small non-coding RNAs participate in thepost-transcriptional modification of ribosomal RNA (rRNA)nucleotides. Box-C/D-snoRNAs contain conservative boxes anda specific guide region in their structure that are responsible fortheir ability to guide 20-O-methylation. GAS5 intron-encodedbox-C/D-snoRNAs were selected as regulatory RNA modelssince the authors have already demonstrated the possibility ofediting corresponding SNORD genes in human cells withCRISPR/Cas9 (Previously published in: Filippova JA et al.(2019) Front Pharmacol, Nov 4;10:1246). IFITM3 gene contain-ing single intron was chosen to be a model gene of interest forre-targeting snoRNA activity at its pre-mRNA. Human cell linescarrying altered SNORD genes were obtained through CRISPR/Cas9-mediated homologous recombination in 293FT and A549cells. Cells were demonstrated to express novel modified snoRNAvariants carrying guide regions targeted at the IFITM3 pre-mRNA nucleotides that are essential for splicing. The specificdownregulation of mature IFITM3 mRNA (according to RNA-Seq data) as well as the decrease in the protein level were shown.The obtained results work in favor of the suggested snoRNA-me-diated splicing inhibition strategy. This work was supported bythe RFBR grant № 18-29-07073 and partially (in method devel-opment) by State Budget Program (№ 122022100238-7).

AB - Precursor mRNA (pre-mRNA) splicing presents one of the cru-cial stages of mRNA maturation in eukaryote cells. Alternativesplicing allows the formation of multiple isoforms thus providingtranscriptomic diversity and the assortment of biologicalfunctions. Hence, there is a need for the development of splicingmodulation tools in order to fine-tune gene expression withoutdisrupting gene structure. The current study focuses on smallnucleolar RNAs (snoRNAs) as a possible splicing regulator. Ineukaryote cells these small non-coding RNAs participate in thepost-transcriptional modification of ribosomal RNA (rRNA)nucleotides. Box-C/D-snoRNAs contain conservative boxes anda specific guide region in their structure that are responsible fortheir ability to guide 20-O-methylation. GAS5 intron-encodedbox-C/D-snoRNAs were selected as regulatory RNA modelssince the authors have already demonstrated the possibility ofediting corresponding SNORD genes in human cells withCRISPR/Cas9 (Previously published in: Filippova JA et al.(2019) Front Pharmacol, Nov 4;10:1246). IFITM3 gene contain-ing single intron was chosen to be a model gene of interest forre-targeting snoRNA activity at its pre-mRNA. Human cell linescarrying altered SNORD genes were obtained through CRISPR/Cas9-mediated homologous recombination in 293FT and A549cells. Cells were demonstrated to express novel modified snoRNAvariants carrying guide regions targeted at the IFITM3 pre-mRNA nucleotides that are essential for splicing. The specificdownregulation of mature IFITM3 mRNA (according to RNA-Seq data) as well as the decrease in the protein level were shown.The obtained results work in favor of the suggested snoRNA-me-diated splicing inhibition strategy. This work was supported bythe RFBR grant № 18-29-07073 and partially (in method devel-opment) by State Budget Program (№ 122022100238-7).

UR - https://febs.onlinelibrary.wiley.com/doi/epdf/10.1002/2211-5463.13646

M3 - Conference article

VL - 13

SP - P-04.2-02

JO - FEBS Open Bio

JF - FEBS Open Bio

SN - 2211-5463

IS - S2

ER -