Alternative splicing regulation of cell-cycle genes by SPF45/SR140/CHERP complex controls cell proliferation

Elena, Martin; Vivori, Claudia; Rogalska, Malgorzata; Vicente, Jorge Herrero; Valcárcel, Juan

doi:10.1261/rna.078935.121

Cited by 17 publications

(24 citation statements)

References 69 publications

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“…This discrepancy might be due to the different cell types used in both analyses. This possibility is supported by the recent findings that CHERP depletion in Hela cells frequently caused retained intron events [ 24 ] and implies that CHERP regulated the alternative splicing in a cell-specific manner. In our analysis, weak or suboptimal splice sites, high GC content in introns, and short intron lengths were important and common features of intron retention.…”

Section: Discussionsupporting

confidence: 62%

CHERP Regulates the Alternative Splicing of pre-mRNAs in the Nucleus

Yamanaka

Ishizuka

Fujita

et al. 2022

IJMS

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Calcium homeostasis endoplasmic reticulum protein (CHERP) is colocalized with the inositol 1,4,5-trisphosphate receptor (IP3R) in the endoplasmic reticulum or perinuclear region, and has been involved in intracellular calcium signaling. Structurally, CHERP carries the nuclear localization signal and arginine/serine-dipeptide repeats, like domain, and interacts with the spliceosome. However, the exact function of CHERP in the nucleus remains unknown. Here, we showed that poly(A)+ RNAs accumulated in the nucleus of CHERP-depleted U2OS cells. Our global analysis revealed that CHERP regulated alternative mRNA splicing events by interaction with U2 small nuclear ribonucleoproteins (U2 snRNPs) and U2 snRNP-related proteins. Among the five alternative splicing patterns analyzed, intron retention was the most frequently observed event. This was in accordance with the accumulation of poly(A)+ RNAs in the nucleus. Furthermore, intron retention and cassette exon choices were influenced by the strength of the 5′ or 3′ splice site, the branch point site, GC content, and intron length. In addition, CHERP depletion induced anomalies in the cell cycle progression into the M phase, and abnormal cell division. These results suggested that CHERP is involved in the regulation of alternative splicing.

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Section: Discussionsupporting

confidence: 62%

CHERP Regulates the Alternative Splicing of pre-mRNAs in the Nucleus

Yamanaka

Ishizuka

Fujita

et al. 2022

IJMS

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“…Among the OTCD mouse models, the mouse spf ash is characterized by the c.386G>A splicing mutation and it has been exploited to evaluate replacement gene therapy approaches (Moscioni et al 2006 ; Wang et al 2011 , 2012 ; Cunningham et al 2013 ). This animal model represents therefore an ideal pre-clinical platform to assess the therapeutic potential of RNA therapeutics based on engineered variants of the spliceosomal U1snRNAs, which were proven to be capable, both in cellular (Glaus et al 2011 ; Schmid et al 2013 ; Scalet et al 2017 , 2018 , 2019 ; Martínez-Pizarro et al 2018 ; Balestra et al 2019a , b ; Martín et al 2021 ) and animal (Balestra et al 2014 , 2016 , 2020b ; Dal Mas et al 2015 ; Rogalska et al 2016 ) models of human diseases, to counteract splicing mutations and force exon recognition, thus rescuing gene expression. Recently, a U1snRNA variant, upon delivery via an Adeno-Associated virus Vector in spf ash mice, partially restored OTC expression at RNA and protein level in liver (Balestra et al 2020a ).…”

Section: Discussionmentioning

confidence: 99%

“…Among molecules acting at RNA levels, engineered variants of the U1snRNA, the RNA component of the spliceosomal U1 ribonucleoprotein (U1snRNP) mediating 5′ss recognition and thus exon definition in the earliest splicing steps (De Conti et al 2013 ), have proven to be effective in modulating splicing for therapeutic purposes. In particular, U1snRNA variants with increased complementarity with the 5′ss of the defective exon (named compensatory U1snRNA), or targeting the downstream intronic sequences (named Exon specific U1snRNA, ExSpeU1), have demonstrated their ability to rescue exon skipping caused by different types of mutations in cellular and animal models of several human diseases (Donadon et al 2018 , 2019 ; Scalet et al 2018 , 2019 ; Yamazaki et al 2018 ; Balestra et al 2019a , 2020a ; Lee et al 2019 ; Donegà et al 2020 ; Martín et al 2021 ).…”

Section: Introductionmentioning

confidence: 99%

OTC intron 4 variations mediate pathogenic splicing patterns caused by the c.386G>A mutation in humans and spfash mice, and govern susceptibility to RNA-based therapies

Sacchetto

Peretto²,

Baralle³

et al. 2021

Mol Med

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Background Aberrant splicing is a common outcome in the presence of exonic or intronic variants that might hamper the intricate network of interactions defining an exon in a specific gene context. Therefore, the evaluation of the functional, and potentially pathological, role of nucleotide changes remains one of the major challenges in the modern genomic era. This aspect has also to be taken into account during the pre-clinical evaluation of innovative therapeutic approaches in animal models of human diseases. This is of particular relevance when developing therapeutics acting on splicing, an intriguing and expanding research area for several disorders. Here, we addressed species-specific splicing mechanisms triggered by the OTC c.386G>A mutation, relatively frequent in humans, leading to Ornithine TransCarbamylase Deficiency (OTCD) in patients and spfash mice, and its differential susceptibility to RNA therapeutics based on engineered U1snRNA. Methods Creation and co-expression of engineered U1snRNAs with human and mouse minigenes, either wild-type or harbouring different nucleotide changes, in human (HepG2) and mouse (Hepa1-6) hepatoma cells followed by analysis of splicing pattern. RNA pulldown studies to evaluate binding of specific splicing factors. Results Comparative nucleotide analysis suggested a role for the intronic +10-11 nucleotides, and pull-down assays showed that they confer preferential binding to the TIA1 splicing factor in the mouse context, where TIA1 overexpression further increases correct splicing. Consistently, the splicing profile of the human minigene with mouse +10-11 nucleotides overlapped that of mouse minigene, and restored responsiveness to TIA1 overexpression and to compensatory U1snRNA. Swapping the human +10-11 nucleotides into the mouse context had opposite effects. Moreover, the interplay between the authentic and the adjacent cryptic 5′ss in the human OTC dictates pathogenic mechanisms of several OTCD-causing 5′ss mutations, and only the c.386+5G>A change, abrogating the cryptic 5′ss, was rescuable by engineered U1snRNA. Conclusions Subtle intronic variations explain species-specific OTC splicing patterns driven by the c.386G>A mutation, and the responsiveness to engineered U1snRNAs, which suggests careful elucidation of molecular mechanisms before proposing translation of tailored therapeutics from animal models to humans.

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“…Similar to DNA and proteins, RNAs are also substrates for chemical modifications (24). N 6 methyladenosine (m 6 A), the most abundant internal modification in eukaryotic mRNA, has been shown to influence AS (25)(26)(27)(28). m 6 A is co-transcriptionally deposited by the methyltransferase-like 3 (METTL3) and METTL14 methyltransferase complex, which partially localizes to nuclear speckles, where splicing occurs (29)(30)(31).…”

Section: Introductionmentioning

confidence: 99%

“…N 6 methyladenosine (m 6 A), the most abundant internal modification in eukaryotic mRNA, has been shown to influence AS (25)(26)(27)(28). m 6 A is co-transcriptionally deposited by the methyltransferase-like 3 (METTL3) and METTL14 methyltransferase complex, which partially localizes to nuclear speckles, where splicing occurs (29)(30)(31). It has been shown that depletion of the Drosophila METTL3 methyltransferase homologue, results in altered AS patterns that influence sex determination (32)(33)(34).…”

Section: Introductionmentioning

confidence: 99%

METTL3 regulates breast cancer-associated alternative splicing switches

Achour

Groza

Bhattarai

et al. 2022

Preprint

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Alternative splicing (AS) enables differential inclusion of exons from a given transcript, thereby contributing to the transcriptome and proteome diversity. Aberrant AS patterns play major roles in the development of different pathologies, including breast cancer. N6-methyladenosine (m6A), the most abundant internal modification of eukaryotic mRNA, influences tumor progression and metastasis of breast cancer, and it has been recently linked to AS regulation. Here, we identify a specific AS signature associated with breast tumorigenesis in vitro. We characterize for the first time the role of METTL3 in modulating breast cancer-associated AS programs, expanding the role of the m6A-methyltransferase in tumorigenesis. Specifically, we find that both m6A deposition in splice site boundaries and in splicing and transcription factor transcripts, such as PHF5A and MYC, direct AS switches of specific breast cancer-associated transcripts. Finally, we show that five of the AS events validated in vitro are associated with a poor overall survival rate for patients with breast cancer, suggesting the use of these AS events as a novel potential prognostic biomarker.

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Alternative splicing regulation of cell-cycle genes by SPF45/SR140/CHERP complex controls cell proliferation

Cited by 17 publications

References 69 publications

CHERP Regulates the Alternative Splicing of pre-mRNAs in the Nucleus

CHERP Regulates the Alternative Splicing of pre-mRNAs in the Nucleus

OTC intron 4 variations mediate pathogenic splicing patterns caused by the c.386G>A mutation in humans and spfash mice, and govern susceptibility to RNA-based therapies

METTL3 regulates breast cancer-associated alternative splicing switches

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