The Splicing Factor hnRNPA1 Regulates Alternate Splicing of the <i>MYLK</i> Gene

Mascarenhas, Joseph B.; Tchourbanov, Alex Y.; Danilov, Sergei M.; Zhou, Tong; Wang, Ting; Garcia, Joe G.N.

doi:10.1165/rcmb.2017-0141oc

Cited by 15 publications

(11 citation statements)

References 43 publications

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“…The moxifloxacin treatment increased the amount of SRSF1 splicing factor in SMA fibroblasts, which affected the CASP9 exon cassette splice‐in; however, the effect on the inclusion of MAPT exon 10 was mild (Figure A,B). hnRNPA1 inhibits the inclusion of INSR exon 11 and promotes skipping of MYLK exon 11 . The reduced levels of hnRNPA1 after moxifloxacin treatment increased the inclusion of MYLK exon 11 as expected; however, in the case of the INSR splicing, the effect was significant and dose‐dependent but in contrast to the anticipated result (Figure C,D).…”

Section: Resultssupporting

confidence: 55%

“…hnRNPA1 inhibits the inclusion of INSR exon 11 43 and promotes skipping of MYLK exon 11. 44 The reduced levels of hnRNPA1 after moxifloxacin treatment increased the inclusion of MYLK exon 11 as expected; however, in the case of the INSR splicing, the effect was significant and dose-dependent but in contrast to the anticipated result ( Figure 8C,D). This can be caused by different cell lines than the one used in the original study, or more factors are implicated in the INSR exon 11 splicing.…”

Section: Moxifloxacin Treatment Modulates Levels Of Two Main Smn2 Ssupporting

confidence: 66%

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Drosophila SMN2minigene reporter model identifies moxifloxacin as a candidate therapy for SMA

Konieczny

Artero

2019

FASEB j.

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Spinal muscular atrophy is a rare and fatal neuromuscular disorder caused by the loss of alpha motor neurons. The affected individuals have mutated the ubiquitously expressed SMN1 gene resulting in the loss or reduction in the survival motor neuron (SMN) protein levels. However, an almost identical paralog exists in humans: SMN2.Pharmacological activation of SMN2 exon 7 inclusion by small molecules or modified antisense oligonucleotides is a valid approach to treat SMA. Here we describe an in vivo SMN2 minigene reporter system in Drosophila motor neurons that serves as a cost-effective, feasible, and stringent primary screening model for identifying chemicals capable of crossing the conserved Drosophila blood-brain barrier and modulating exon 7 inclusion. The model was used for the screening of 1100 drugs from the Prestwick Chemical Library, resulting in 2.45% hit rate. The most promising candidate drugs were validated in patient-derived fibroblasts where they proved to increase SMN protein levels. Among them, moxifloxacin modulated SMN2 splicing by promoting exon 7 inclusion. The recovery of SMN protein levels was confirmed by increased colocalization of nuclear gems with Cajal Bodies. Thus, a Drosophilabased drug screen allowed the discovery of an FDA-approved small molecule with the potential to become a novel therapy for SMA. K E Y W O R D S drug repurposing, SMA, SMN2, splicing 3022 | KONIECZNY aNd aRTERO | 3035 KONIECZNY aNd aRTEROtogether we have managed to use Drosophila as an in vivo tool to discover compounds with interesting properties on an SMN2 exon 7 reporter construct as evidenced by the ability of moxifloxacin to modulate exon 7 inclusion in the fly motorneurons.

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Section: Resultssupporting

confidence: 55%

Section: Moxifloxacin Treatment Modulates Levels Of Two Main Smn2 Ssupporting

confidence: 66%

Drosophila SMN2minigene reporter model identifies moxifloxacin as a candidate therapy for SMA

Konieczny

Artero

2019

FASEB j.

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“…Because pulmonary epithelial cell stretch is accompanied by increased mechanical stress in airways, cyclic stretch was used to imitate airway mechanical stress by Flex cell Tension System. Related studies showed that 18% elongation was widely applied to human pulmonary artery endothelial cells and pulmonary microvascular endothelial cells in ventilator-induced pulmonary injury [36–38]. 12% elongation was applied in airway smooth muscle hypertrophy using Flex cell Tension System [39].…”

Section: Discussionmentioning

confidence: 99%

Role of TRPC1 channels in pressure-mediated activation of airway remodeling

et al. 2019

Respir Res

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Background Bronchoconstriction and cough, a characteristic of the asthmatic response, leads to development of compressive stresses in the airway wall. We hypothesized that progressively pathological high mechanical stress could act on mechanosensitive cation channels, such as transient receptor potential channel 1 (TRPC1) and then contributes to airway remodeling. Methods We imitate the pathological airway pressure in vitro using cyclic stretch at 10 and 15% elongation. Ca 2+ imaging was applied to measure the activity of TRPC1 after bronchial epithelial cells exposed to cyclic stretch for 0, 0.5, 1, 1.5, 2, 2.5 h. To further clarify the function of channnel TRPC1 in the process of mechano-transduction in airway remodeling, the experiment in vivo was implemented. The TRPC1 siRNA and budesonide were applied separately to asthmatic models. The morphological changes were measured by HE and Massion method. The expression levels of TRPC1 were evaluated by real-time PCR, western blot and immunohistochemistry. The protein expression level of IL-13, TGF-β 1 and MMP-9 in BALF were measured by ELISA. Results The result showed that cyclic stretch for 15% elongation at 1.5 h could maximize the activity of TRPC1 channel. This influx in Ca 2+ was blocked by TRPC1 siRNA. Higher TRPC1 expression was observed in the bronchial epithelial layer of ovalbumin induced asthmatic models. The knockdown of TRPC1 with TRPC1 siRNA was associated with a hampered airway remodeling process, such as decreased bronchial wall thickness and smooth muscle hypertrophy/hyperplasia, a decreased ECM deposition area and inflammation infiltration around airway wall. Meantime, expression of IL-13, TGF-β 1 and MMP-9 in OVA+TRPC1 siRNA also showed reduced level. TRPC1 intervention treatment showed similar anti-remodeling therapeutic effect with budesonide. Conclusions These results demonstrate that most TRPC1 channels expressed in bronchial epithelial cells mediate the mechanotransduction mechanism. TRPC1 inducing abnormal Ca 2+ signal mediates receptor-stimulated and mechanical stimulus-induced airway remodeling. The inhibition of TRPC1 channel could produce similar therapeutic effect as glucocortisteroid to curb the development of asthmatic airway remodeling.

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“…hnRNP A1 belongs to the hnRNP family, a large family of ubiquitously expressed heterogeneous nuclear ribonucleoproteins (hnRNPs) that regulate several aspects of nucleic acid metabolism, including pre-mRNA packing, alternative splicing, and transportation of poly(A) mRNA 14 - 16 . hnRNP expression is altered in many types of cancers 17 .…”

Section: Introductionmentioning

confidence: 99%

SAM68 promotes tumorigenesis in lung adenocarcinoma by regulating metabolic conversion via PKM alternative splicing

Zhu¹,

Chen²,

Wang³

et al. 2021

Theranostics

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Background: A metabolic “switch” from oxidative phosphorylation to glycolysis provides tumor cells with energy and biosynthetic substrates, thereby promoting tumorigenesis and malignant progression. However, the mechanisms controlling this metabolic switch in tumors is not entirely clear. Methods: Clinical specimens were used to determine the effect of SAM68 on lung adenocarcinoma (LUAD) tumorigenesis and metastasis, and mouse models and molecular biology assays were performed to elucidate the function and underlying mechanisms in vitro and in vivo . Results: SAM68 mRNA levels were higher in LUAD tissue than in normal lung tissue, indicating that SAM68 expression is upregulated in LUAD. Patients with LUAD with SAM68 high (n = 257) had a higher frequency of tumor recurrence ( p = 0.025) and recurrence-free survival ( p = 0.013) than did those with SAM68 low (n = 257). Patients with SAM68 high mRNA levels (n = 257) were at a higher risk for cancer-related death ( p = 0.006), and had shorter overall survival ( p = 0.044) than did those with SAM68 low . SAM68 promotes tumorigenesis and metastasis of LUAD cells in vitro and in vivo by regulating the cancer metabolic switch. SAM68 drives cancer metabolism by mediating alternative splicing of pyruvate kinase (PKM) pre-mRNAs, and promoting the formation of PKM2. Mechanistically, SAM68 increased the binding of the splicing repressor hnRNP A1 to exon 9 of PKM , thereby enhancing PKM2 isoform formation and PKM2-dependent aerobic glycolysis and tumorigenesis. Conclusions: SAM68 promotes LUAD cell tumorigenesis and cancer metabolic programming via binding of the 351-443 aa region of SAM68 to the RGG motif of hnRNP A1, driving hnRNP A1-dependent PKM splicing, contributing to increased oncogene PKM2 isoform formation and inhibition of PKM1 isoform formation. SAM68 is therefore a promising therapeutic target for the treatment of LUAD.

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The Splicing Factor hnRNPA1 Regulates Alternate Splicing of the MYLK Gene

Cited by 15 publications

References 43 publications

Drosophila SMN2minigene reporter model identifies moxifloxacin as a candidate therapy for SMA

Drosophila SMN2minigene reporter model identifies moxifloxacin as a candidate therapy for SMA

Role of TRPC1 channels in pressure-mediated activation of airway remodeling

SAM68 promotes tumorigenesis in lung adenocarcinoma by regulating metabolic conversion via PKM alternative splicing

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