High-throughput analysis revealed mutations’ diverging effects on SMN1 exon 7 splicing

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F00209775%3A_____%2F19%3AN0000022" target="_blank" >RIV/00209775:_____/19:N0000022 - isvavai.cz</a>

Nalezeny alternativní kódy

RIV/00216224:14740/19:00107593

Výsledek na webu

<a href="https://www.tandfonline.com/doi/full/10.1080/15476286.2019.1630796?scroll=top&needAccess=true" target="_blank" >https://www.tandfonline.com/doi/full/10.1080/15476286.2019.1630796?scroll=top&needAccess=true</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1080/15476286.2019.1630796" target="_blank" >10.1080/15476286.2019.1630796</a>

Jazyk výsledku

angličtina

Název v původním jazyce

High-throughput analysis revealed mutations’ diverging effects on SMN1 exon 7 splicing

Popis výsledku v původním jazyce

Splicing-affecting mutations can disrupt gene function by altering the transcript assembly. To ascertain splicing dysregulation principles, we modified a minigene assay for the parallel high-throughput evaluation of different mutations by next-generation sequencing. In our model system, all exonic and six intronic positions of the SMN1 gene’s exon 7 were mutated to all possible nucleotide variants, which amounted to 180 unique single-nucleotide mutants and 470 double mutants. The mutations resulted in a wide range of splicing aberrations. Exonic splicing-affecting mutations resulted either in substantial exon skipping, supposedly driven by predicted exonic splicing silencer or cryptic donor splice site (5′ss) and de novo 5′ss strengthening and use. On the other hand, a single disruption of exonic splicing enhancer was not sufficient to cause major exon skipping, suggesting these elements can be substituted during exon recognition. While disrupting the acceptor splice site led only to exon skipping, some 5′ss mutations potentiated the use of three different cryptic 5′ss. Generally, single mutations supporting cryptic 5′ss use displayed better pre-mRNA/U1 snRNA duplex stability and increased splicing regulatory element strength across the original 5′ss. Analyzing double mutants supported the predominating splicing regulatory elements’ effect, but U1 snRNA binding could contribute to the global balance of splicing isoforms. Based on these findings, we suggest that creating a new splicing enhancer across the mutated 5′ss can be one of the main factors driving cryptic 5′ss use.

Název v anglickém jazyce

High-throughput analysis revealed mutations’ diverging effects on SMN1 exon 7 splicing

Popis výsledku anglicky

Splicing-affecting mutations can disrupt gene function by altering the transcript assembly. To ascertain splicing dysregulation principles, we modified a minigene assay for the parallel high-throughput evaluation of different mutations by next-generation sequencing. In our model system, all exonic and six intronic positions of the SMN1 gene’s exon 7 were mutated to all possible nucleotide variants, which amounted to 180 unique single-nucleotide mutants and 470 double mutants. The mutations resulted in a wide range of splicing aberrations. Exonic splicing-affecting mutations resulted either in substantial exon skipping, supposedly driven by predicted exonic splicing silencer or cryptic donor splice site (5′ss) and de novo 5′ss strengthening and use. On the other hand, a single disruption of exonic splicing enhancer was not sufficient to cause major exon skipping, suggesting these elements can be substituted during exon recognition. While disrupting the acceptor splice site led only to exon skipping, some 5′ss mutations potentiated the use of three different cryptic 5′ss. Generally, single mutations supporting cryptic 5′ss use displayed better pre-mRNA/U1 snRNA duplex stability and increased splicing regulatory element strength across the original 5′ss. Analyzing double mutants supported the predominating splicing regulatory elements’ effect, but U1 snRNA binding could contribute to the global balance of splicing isoforms. Based on these findings, we suggest that creating a new splicing enhancer across the mutated 5′ss can be one of the main factors driving cryptic 5′ss use.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

10608 - Biochemistry and molecular biology

Projekt

Výsledek vznikl pri realizaci vícero projektů. Více informací v záložce Projekty.

Návaznosti

P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)

Rok uplatnění

2019

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Název periodika

RNA Biology

ISSN

1547-6286

e-ISSN

—

Svazek periodika

16

Číslo periodika v rámci svazku

10

Stát vydavatele periodika

US - Spojené státy americké

Počet stran výsledku

13

Strana od-do

1364-1376

Kód UT WoS článku

000472379600001

EID výsledku v databázi Scopus

2-s2.0-85067683899