Rbm24 modulates neuronal RNA splicing to restrict cognitive dysfunction.

Synaptic dysfunction is associated with early neurodegenerative changes and cognitive deficits. Neuronal cell-specific alternative splicing (AS) programs exclusively encode unique neuron- and synapse-specific proteins. However, it remains unclear whether splicing disturbances in neurons influence the pathogenesis of cognitive impairment. Here, we observed that RNA-binding motif protein 24 (RBM24) expression was decreased in Alzheimer's disease (AD) patients. Using conditional RBM24 knockout mice, we demonstrated that deletion of RBM24 in the brain resulted in learning and memory impairment. Electrophysiological recordings from hippocampal slices from mice lacking RBM24 revealed multiple defects in excitatory synaptic function and plasticity. Furthermore, RNA sequencing and splicing analysis showed that RBM24 regulates a network of genes related to cognitive function. Deletion of RBM24 disrupted the AS of synapse-associated genes, including GluR2 and Prrt1, the major disease genes involved in cognitive impairment and memory loss, leading to cognitive dysfunction. Together, our results suggest that the regulation of mRNA splicing by RBM24 is a key process involved in maintaining normal synaptic function and provide novel mechanistic insights into the pathogenesis of AD.

YAP1-induced RBM24 promotes the tumorigenesis of triple-negative breast cancer through the ß-catenin pathway.

Triple-negative breast cancer (TNBC) is the most aggressive breast cancer subtype and refractory to current treatments. RBM24 is an RNA-binding protein and shows the ability to regulate tumor progression in multiple cancer types. However, its role in TNBC is still unclear. In this study, we analyzed publicly available profiling data from TNBC tissues and cells. Loss- and gain-of-function experiments were performed to determine the function of RBM24 in TNBC cells. The mechanism for RBM24 action in TNBC was investigated. RBM24 was deregulated in TNBC tissues and TNBC cells with depletion of SIPA1, YAP1, or ARID1A, three key regulators of TNBC. Compared to MCF10A breast epithelial cells, TNBC cells had higher levels of RBM24. Knockdown of RBM24 inhibited TNBC cell proliferation, colony formation, and tumorigenesis, while overexpression of RBM24 promoted aggressive phenotype in TNBC cells. YAP1 overexpression induced the expression of RBM24 and the RBM24 promoter-driven luciferase reporter. YAP1 was enriched at the promoter region of RBM24. Overexpression of RBM24 increased ß-catenin-dependent transcriptional activity. Most importantly, knockdown of CTNNB1 rescued RBM24 aggressive phenotype in TNBC cells. Collectively, the YAP1/RBM24/ß-catenin axis plays a critical role in driving TNBC progression. RBM24 may represent a novel therapeutic target for TNBC treatment.

Deciphering the genetic interactions between Pou4f3, Gfi1, and Rbm24 in maintaining mouse cochlear hair cell survival.

Mammals harbor a limited number of sound-receptor hair cells (HCs) that cannot be regenerated after damage. Thus, investigating the underlying molecular mechanisms that maintain HC survival is crucial for preventing hearing impairment. Intriguingly, Pou4f3-/- or Gfi1-/- HCs form initially but then rapidly degenerate, whereas Rbm24-/- HCs degenerate considerably later. However, the transcriptional cascades involving Pou4f3, Gfi1, and Rbm24 remain undescribed. Here, we demonstrate that Rbm24 expression is completely repressed in Pou4f3-/- HCs but unaltered in Gfi1-/- HCs, and further that the expression of both POU4F3 and GFI1 is intact in Rbm24-/- HCs. Moreover, by using in vivo mouse transgenic reporter assays, we identify three Rbm24 enhancers to which POU4F3 binds. Lastly, through in vivo genetic testing of whether Rbm24 restoration alleviates the degeneration of Pou4f3-/- HCs, we show that ectopic Rbm24 alone cannot prevent Pou4f3-/- HCs from degenerating. Collectively, our findings provide new molecular and genetic insights into how HC survival is regulated.

RBM24 Suppresses the Tumorigenesis of Glioblastoma by Stabilizing LATS1 mRNA.

The ribose nucleic acid (RNA)-binding motif protein 24 (RBM24) has been recognized as a critical regulatory protein in various types of tumors. However, its specific role in glioblastoma (GBM) has not been thoroughly investigated. The objective of this study is to uncover the role of RBM24 in GBM and understand the underlying mechanism. The expression of RBM24 in GBM was initially analyzed using the Gene Expression Profiling Interactive Analysis (GEPIA). Subsequently, the RBM24 expression levels in clinical samples of GBM were examined, and the survival curves of GBM patients were plotted based on high- and low-expression levels of RBM24 using Kaplan-Meier (KM) plotter. In addition, RBM24 knockdown cell lines and overexpression vectors were created to assess the effects on proliferation, apoptosis, and invasion abilities. Finally, the binding level of RBM24 protein to LATS1 messenger RNA (mRNA) was determined by RNA immunoprecipitation (RIP) assay, and the expression levels of RBM24 and LATS1 were measured through quantitative reverse-transcriptase-polymerase chain reaction (qRT-PCR) and Western blot (WB). Our data revealed a significant decrease in RBM24 mRNA and protein levels in GBM patients, indicating that those with low RBM24 expression had a worse prognosis. Overexpression of RBM24 led to inhibited cell proliferation, reduced invasion, and increased apoptosis in LN229 and U87 cells. In addition, knocking down LATS1 partially reversed the effects of RBM24 on cell proliferation, invasion, and apoptosis in GBM cells. In vivo xenograft model further demonstrated that RBM24 overexpression reduced the growth of subcutaneous tumors in nude mice, accompanied by a decrease in Ki-67 expression and an increase in apoptotic events in tumor tissues. There was also correlation between RBM24 and LATS1 protein expression in the xenograft tumors. RBM24 functions to stabilize LATS1 mRNA, thereby inhibiting the proliferation, suppressing invasion, and promoting apoptosis in GBM cells.

RNA-Binding Proteins in Cardiomyopathies.

The post-transcriptional regulation of gene expression plays an important role in heart development and disease. Cardiac-specific alternative splicing, mediated by RNA-binding proteins, orchestrates the isoform switching of proteins that are essential for cardiomyocyte organization and contraction. Dysfunctions of RNA-binding proteins impair heart development and cause the main types of cardiomyopathies, which represent a heterogenous group of abnormalities that severely affect heart structure and function. In particular, mutations of RBM20 and RBFOX2 are associated with dilated cardiomyopathy, hypertrophic cardiomyopathy, or hypoplastic left heart syndrome. Functional analyses in different animal models also suggest possible roles for other RNA-binding proteins in cardiomyopathies because of their involvement in organizing cardiac gene programming. Recent studies have provided significant insights into the causal relationship between RNA-binding proteins and cardiovascular diseases. They also show the potential of correcting pathogenic mutations in RNA-binding proteins to rescue cardiomyopathy or promote cardiac regeneration. Therefore, RNA-binding proteins have emerged as promising targets for therapeutic interventions for cardiovascular dysfunction. The challenge remains to decipher how they coordinately regulate the temporal and spatial expression of target genes to ensure heart function and homeostasis. This review discusses recent advances in understanding the implications of several well-characterized RNA-binding proteins in cardiomyopathies, with the aim of identifying research gaps to promote further investigation in this field.

RBM24 is required for mouse hair cell development through regulating pre-mRNA alternative splicing and mRNA stability.

As the sensory receptor cells in vertebrate inner ear and lateral lines, hair cells are characterized by the hair bundle that consists of one tubulin-based kinocilium and dozens of actin-based stereocilia on the apical surface of each hair cell. Hair cell development is tightly regulated, and deficits in this process usually lead to hearing loss and/or balance dysfunctions. RNA-binding motif protein 24 (RBM24) is an RNA-binding protein that is specifically expressed in the hair cells in the inner ear. Previously, we showed that RBM24 affects hair cell development in zebrafish by regulating messenger RNA (mRNA) stability. In the present work, we further investigate the role of RBM24 in hearing and balance using conditional knockout mice. Our results show that Rbm24 knockout results in severe hearing and balance deficits. Hair cell development is significantly affected in Rbm24 knockout cochlea, as the hair bundles are poorly developed and eventually degenerated. Hair bundle disorganization is also observed in Rbm24 knockout vestibular hair cells, although to a lesser extent. Consistently, significant hair cell loss is observed in the cochlea but not vestibule. RNAseq analysis identified several genes whose mRNA stability or pre-mRNA alternative splicing is affected by Rbm24 knockout. Among them are Cdh23, Pcdh15, and Myo7a, which have been shown to play important roles in stereocilia development as well as mechano-electrical transduction. Taken together, our present work suggests that RBM24 is required for mouse hair cell development through regulating pre-mRNA alternative splicing as well as mRNA stability.

Cellular activity of autophagy and multivesicular bodies in lens fiber cells during early lens development in rbm24a mutant of zebrafish: Ultrastructure analysis.

Use of zebrafish as animal model for various diseases during early developmental stages has been exponentially increased with the aim to achieve the best representative results in this transparent fish. Recent studies documented that Rbm24a mutant causes cataract formation and resulted in blindness using the zebrafish model. Therefore, correct interpretation of studies that aimed for molecular approaches, a description of comparative and in-depth analysis of development of lens in wildtype and mutant is crucial to obtain the correct conclusion. In this study, we use a gold standard method the Transmission Electron Microscopy (TEM) to analysis the lens development in rbm24a mutant zebrafish. Firstly, we compare the cellular structures at 16-20 h post fertilization (hpf), the lens placode in ectoderm indicated delay lens development in rbm24a mutant than wildtype (siblings) zebrafish. At 33 hpf, loosely appeared lens fiber cells showed heterogenous electron density with numbers of mitochondria in lens of rbm24a mutant, revealed the influence of gene mutation in lens development. A detail ultrastructure of lens of rbm24a mutant also presented at 33 hpf. Comparatively in wildtype (siblings) at 33 hpf, lens exhibited homogenous electron density in tightly packed lens fiber cells with few mitochondria. Furthermore, to characterize the lens in rbm24a mutant we obtained data of cellular structures on 25 hpf and 1.5 days' post fertilization (dpf). At 25 hpf in mutant zebrafish, the detached solid sphere lens mass from ectoderm showed karyorrhexis, mitophagy and vesicles (also multivesicular bodies), these cellular structures supposed to hamper the development of future fiber cells. Moreover, at 1.5 dpf in mutant, nuclear excisosome, multilamellar bodies and irregular shaped mitochondria in heterogenous electron dense cytoplasm of lens fiber cells, collectively shown affected lens transparency. In summary the ultrastructure results of lens of rbm24a mutant zebrafish expand our knowledge and give reflection of different cellular activities like autophagy, apoptosis, vesicles (multivesicular bodies) and nuclear excisosomes which play their role in transparency achievement.

RBM24 inhibits the translation of SARS-CoV-2 polyproteins by targeting the 5'-untranslated region.

SARS-CoV-2 is a betacoronavirus with single-stranded positive-sense RNA, which is a serious global threat to human health. Understanding the molecular mechanism of viral replication is crucial for the development of antiviral drugs. The synthesis of viral polyproteins is a crucial step in viral progression. The synthesis of viral polyproteins in coronaviruses is regulated by the 5'-untranslated region (UTR); however, the detailed regulatory mechanism needs further investigation. The present study demonstrated that the RNA binding protein, RBM24, interacts with the RNA genome of SARS-CoV-2 via its RNA recognition submotifs (RNPs). The findings revealed that RBM24 recognizes and binds to the GUGUG element at stem-loop 4 (SL4) in the 5'-UTR of SARS-CoV-2. The interaction between RBM24 and 5'-UTR prevents 80S ribosome assembly, which in turn inhibits polyproteins translation and the replication of SARS-CoV-2. Notably, other RNA viruses, including SARS-CoV, MERS-CoV, Ebolavirus, rhinovirus, West Nile virus, Zika virus, Japanese encephalitis virus, yellow fever virus, hepatitis C virus, and human immunodeficiency virus-1 also contain one or several G(U/C/A)GUG sequences in the 5'-UTR, which is also targeted by RBM24. In conclusion, the present study demonstrated that RBM24 functions by interacting with the 5'-UTR of SARS-CoV-2 RNA, and elucidated that RBM24 could be a host restriction factor for SARS-CoV-2 and other RNA viruses.

The RNA-binding protein RBM24 regulates lipid metabolism and SLC7A11 mRNA stability to modulate ferroptosis and inflammatory response.

Lipids play a critical role in many cellular processes by serving as structural components of cell membranes or functioning as energy fuel and signaling molecules. The RNA-binding proteins RBM24 and RBM38 share an identical RNA-binding domain and thereby, regulate a group of same targets, such as p21. However, it is not certain whether RBM24 and RBM38 participates in lipid homeostasis. Here, lipidomic analysis showed that a deficiency in RBM24 or RBM38 leads to altered lipid metabolism, with more profound alteration by loss of RBM24 in MCF7 cells. We also showed that mice deficient in RBM24 were prone to chronic inflammation and liver steatosis, but not spontaneous tumors. These data let us speculate whether RBM24 regulates ferroptosis, a programmed cell death that links inflammation and liver steatosis via lipid peroxidation. Indeed, we found that over-expression of RBM24 protected, whereas knockout of RBM24 sensitized, cells to Erastin-induced ferroptosis by modulating the mRNA stability of SLC7A11, a ferroptosis inhibitor. Moreover, we showed that knockdown of SLC7A11 reversed the effect of RBM24 on ferroptosis. Together, our study revealed that RBM24 regulates lipid metabolism and SLC7A11 mRNA stability to modulate ferroptosis and inflammatory response.

Mechanism of METTL14-mediated m6A modification in non-small cell lung cancer cell resistance to cisplatin.

Methyltransferase-like 14 (METTL14) mediates N6-methyladenosine (m6A) modification to influence cancer progression. This study aims to determine the mechanism of METTL14-mediated m6A in non-small cell lung cancer (NSCLC) cell resistance to cisplatin (DDP). METTL14, miR-19a-5p, RBM24, and AXIN1 expressions in NSCLC tissues/cells were determined. DDP-resistant cell line was obtained, followed by the interference of METTL14 expression. NSCLC cells were treated with DDP to establish a drug-resistant cell line, and METTL14 expression in cells was intervened. The IC50 of NSCLC cells to DDP was measured by CCK-8 assay. NSCLC cell proliferation and apoptosis were observed by clone formation assay and flow cytometry. The content of m6A in total RNA in tissues and cells of NSCLC patients was detected using m6A Methylation Quantification Kit. The expressions of DGCR8-bound pri-miR-19a and m6A-modified pri-miR-19a were detected. The binding relationships between miR-19a-5p and RBM24 and RBM24 and AXIN1 were validated using dual-luciferase assay and RNA immunoprecipitation. Finally, mouse xenograft tumor model was established to verify the role of METTL14 in vivo. METTL14 was highly expressed in NSCLC. METTL14 silencing diminished IC50 to DDP, repressed NSCLC cell proliferation, and enhanced apoptosis. METTL14-mediated m6A induced recognition and processing of pri-miR-19a by DGCR8, thus promoting the transition of pri-miR-19a to miR-19a-5p, repressing RBM24 expression, reducing the binding of RBM24 and AXIN1, and suppressing AXIN1 transcription. miR-19a-5p overexpression or RBM24/AXIN1 silencing abolished the effect of METTL14 silencing on NSCLC cell resistance to DDP. METTL14 silencing in vivo enhanced the suppressive role of DDP to tumor growth. Collectively, METTL14-mediated m6A modification facilitated NSCLC cell resistance to DDP via miR-19a-5p/RBM24/AXIN1 axis. KEY MESSAGES: • METTL14 is highly expressed NSCLC and further increased in DDP-resistant cells. • METTL14 silencing attenuates DDP resistance of NSCLC cells. • METTL14 promotes the nature of pri-miR-19a by upregulating pri-miR-19a m6A level. • miR-19a-5p targets RBM24, thus reducing the binding of RBM24 and AXIN1 and inhibiting AXIN1 transcription. • METTL14 silencing in vivo enhances the suppressive role of DDP to tumor growth.

RBM24 is localized to stress granules in cells under various stress conditions.

Stress granules (SGs) are formed when untranslated messenger ribonucleoproteins (mRNPs) accumulate in cells under stress, and are thought to minimize stress-induced damage and promote cell survival. RBM24 (RNA-binding motif protein 24) is an RNA-binding protein that plays pivotal roles in regulating the stability or translation initiation of target mRNAs as well as alternative splicing of target pre-mRNAs. Its important physiological functions are highlighted by the fact that Rbm24 knockout mice or zebrafish suffer from dysfunction of heart, eye, and inner ear. Here we show that RBM24 is recruited into SGs under various stress conditions, suggesting that it might protect its target RNAs in cells under stress. However, SG formation is unaffected when Rbm24 expression is down-regulated. Nevertheless, RBM24 overexpression in cultured cells is sufficient to induce SG formation, suggesting that RBM24 might play an important role in SG formation. In conclusion, our present work reveals that RBM24 is a SG component, which implies that RBM24 could protect its target mRNAs in stressed cells.

RBM24 in the Post-Transcriptional Regulation of Cancer Progression: Anti-Tumor or Pro-Tumor Activity?

RNA-binding proteins are critical post-transcriptional regulators of gene expression. They are implicated in a wide range of physiological and pathological processes by modulating nearly every aspect of RNA metabolisms. Alterations in their expression and function disrupt tissue homeostasis and lead to the occurrence of various cancers. RBM24 is a highly conserved protein that binds to a large spectrum of target mRNAs and regulates many post-transcriptional events ranging from pre-mRNA splicing to mRNA stability, polyadenylation and translation. Studies using different animal models indicate that it plays an essential role in promoting cellular differentiation during organogenesis and tissue regeneration. Evidence is also accumulating that its dysregulation frequently occurs across human cancers. In several tissues, RBM24 clearly functions as a tumor suppressor, which is consistent with its inhibitory potential on cell proliferation. However, upregulation of RBM24 in other cancers appears to promote tumor growth. There is a possibility that RBM24 displays both anti-tumor and pro-tumor activities, which may be regulated in part through differential interactions with its protein partners and by its post-translational modifications. This makes it a potential biomarker for diagnosis and prognosis, as well as a therapeutic target for cancer treatment. The challenge remains to determine the post-transcriptional mechanisms by which RBM24 modulates gene expression and tumor progression in a context- or background-dependent manner. This review discusses recent findings on the potential function of RBM24 in tumorigenesis and provides future directions for better understanding its regulatory role in cancer cells.

RNA-Binding motif protein 38 (RBM38) mediates HBV pgRNA packaging into the nucleocapsid.

The binding of HBV polymerase (Pol) and the epsilon stem loop (ε) on the 5' terminal region of pgRNA is required for pgRNA packaging and HBV replication. Previous research has demonstrated that RNA binding motif protein 24 (RBM24) is involved in pgRNA packaging by mediating the interaction between HBV polymerase (Pol) and the ε element. Here, we demonstrate that RBM38 interacts with ε, pol, RBM24 and HBV core which mediate pgRNA packaging. RBM38 directly binds to the lower bulge of ε via RNA recognition submotifs (RNPs) and interacts with HBV Pol in an RNA-independent manner. RBM38 interacts with RBM24 and forms heterogeneous oligomers, which mediate Pol-ε binding and the formation of the Pol-RBM38/RBM24-ε complex. More important, RBM38 also binds to the HBV core via the C-terminal region (ARD domain), which facilitates the combination of Pol-ε with the HBV core protein. In conclusion, RBM38 facilitates the Pol-ε interaction and mediates Pol-ε in combining with the HBV core, triggering pgRNA packaging for reverse transcription and DNA synthesis. This study provides new insights into pgRNA encapsidation.

RNA-binding proteins and post-transcriptional regulation in lens biology and cataract: Mediating spatiotemporal expression of key factors that control the cell cycle, transcription, cytoskeleton and transparency.

Development of the ocular lens - a transparent tissue capable of sustaining frequent shape changes for optimal focusing power - pushes the boundaries of what cells can achieve using the molecular toolkit encoded by their genomes. The mammalian lens contains broadly two types of cells, the anteriorly located monolayer of epithelial cells which, at the equatorial region of the lens, initiate differentiation into fiber cells that contribute to the bulk of the tissue. This differentiation program involves massive upregulation of select fiber cell-expressed RNAs and their subsequent translation into high amounts of proteins, such as crystallins. But intriguingly, fiber cells achieve this while also simultaneously undergoing significant morphological changes such as elongation - involving about 1000-fold length-wise increase - and migration, which requires modulation of cytoskeletal and cell adhesion factors. Adding further to the challenges, these molecular and cellular events have to be coordinated as fiber cells progress toward loss of their nuclei and organelles, which irreversibly compromises their potential for harnessing genetically hardwired information. A long-standing question is how processes downstream of signaling and transcription, which may also participate in feedback regulation, contribute toward orchestrating these cellular differentiation events in the lens. It is now becoming clear from findings over the past decade that post-transcriptional gene expression regulatory mechanisms are critical in controlling cellular proteomes and coordinating key processes in lens development and fiber cell differentiation. Indeed, RNA-binding proteins (RBPs) such as Caprin2, Celf1, Rbm24 and Tdrd7 have now been described in mediating post-transcriptional control over key factors (e.g. Actn2, Cdkn1a (p21Cip1), Cdkn1b (p27Kip1), various crystallins, Dnase2b, Hspb1, Pax6, Prox1, Sox2) that are variously involved in cell cycle, transcription, cytoskeleton maintenance and differentiation in the lens. Furthermore, deficiencies of these RBPs have been shown to result in various eye and lens defects and/or cataract. Because fiber cell differentiation in the lens occurs throughout life, the underlying regulatory mechanisms operational in development are expected to also be recruited for the maintenance of transparency in aged lenses. Indeed, in support of this, TDRD7 and CAPRIN2 loci have been linked to age-related cataract in humans. Here, I will review the role of key RBPs in the lens and their importance in understanding the pathology of lens defects. I will discuss advances in RBP-based gene expression control, in general, and the important challenges that need to be addressed in the lens to define the mechanisms that determine the epithelial and fiber cell proteome. Finally, I will also discuss in detail several key future directions including the application of bioinformatics approaches such as iSyTE to study RBP-based post-transcriptional gene expression control in the aging lens and in the context of age-related cataract.

RNA-binding protein RBM24 represses colorectal tumourigenesis by stabilising PTEN mRNA.

BACKGROUND: RNA-binding motif protein 24 (RBM24) functions as a splicing regulator, which is critical for organ development and is dysregulated in human cancers. Here, we aim to uncover the biological function of RBM24 in colorectal tumourigenesis. METHODS: Xenograft tumour model, Rbm24 knockout and Apcmin/+ mouse models were utilised. Colorectal cancer cells overexpressing or silencing RBM24 were established. RNA immunoprecipitation (RIP) assay was conducted to detect protein-RNA associations. Gene expression was measured by immunohistochemistry, western blotting, or quantitative PCR (qPCR). RESULTS: Rbm24-knockout mice developed spontaneous colorectal adenomas with lower expression of phosphatase and tensin homolog (PTEN). Immunohistochemical staining for the proliferation markers Ki-67 and pHH3 and BrdU assay showed intestinal hyperplasia in Rbm24-knockout mice compared to wild-type mice. RBM24 expression in colorectal adenoma tissues of Apcmin/+ mouse was downregulated compared with adjacent normal samples and was positively correlated with PTEN expression. In vitro, RBM24 overexpression suppressed cell proliferation, migration, invasion and increased sensitivity to 5-FU or cisplatin in CRC cells. Mechanistically, RBM24 maintained PTEN mRNA stability by directly binding to the GT-rich region at positions 8101-8251 in the 3'-UTR of PTEN mRNA, prolonging the half-life of PTEN mRNA, thereby increasing PTEN expression. Hence, low expression of RBM24 downregulated PTEN mRNA, causing the activation of PI3K-Akt signalling in CRC cells. Furthermore, RBM24 expression in CRC tissues was lower than adjacent normal samples. RBM24 expression was positively correlated with PTEN expression and negatively correlated with Ki-67 level. CRC patients with high RBM24 expression had a favourable outcome. CONCLUSIONS: Taken together, RBM24 expression is markedly lower in colorectal tumours than in para-carcinoma tissues. Rbm24-knockout mice develop spontaneous colorectal adenomas. RBM24 directly binds and stabilises PTEN mRNA, which could cause the suppression of CRC cell proliferation, migration and invasion, thereby repressing colorectal tumourigenesis. These findings support the tumour-suppressive role of RBM24. Targeting RBM24 holds strong promise for the diagnosis and treatment of CRC.

miR­383 increases the cisplatin sensitivity of lung adenocarcinoma cells through inhibition of the RBM24­mediated NF­κB signaling pathway.

The expression of microRNA­383 (miR­383) is downregulated in a variety of tumor tissues, and it exhibits antiproliferative activity in non­small cell lung cancer cells. In the present study, an association between the downregulation of miR­383 expression and the deletion of chr8p22 in patients with lung adenocarcinoma was identified. The promoting effect of miR­383 on cisplatin sensitivity was verified both in vivo and in vitro. Additionally, it was revealed that the expression of RNA binding motif protein 24 (RBM24) protein was regulated by and negatively correlated with miR­383 expression. Ectopic expression of RBM24 or inhibition of miR­383 decreased the chemosensitivity of parental A549 cells, whereas knockdown of RBM24 in cisplatin­resistant A549 cells increased chemosensitivity. Mechanistically, miR­383 interfered with the activation of nuclear factor κB (NF­κB) signaling through repression of RBM24­mediated phosphorylation of Rel­like domain­containing protein A and inhibitor α of NF­κB. Taken together, the downregulation of miR­383 induced RBM24 expression, which was mediated through the activation of NF­κB signaling, to contribute to chemotherapy resistance in lung adenocarcinoma cells. The results of the present study highlight potential therapeutic targets for the clinical reversal of the chemotherapy resistance in lung adenocarcinoma.

TPRG1-AS1 induces RBM24 expression and inhibits liver cancer progression by sponging miR-4691-5p and miR-3659.

BACKGROUND & AIMS: Noncoding RNAs (ncRNAs) play critical roles in hepatocellular carcinoma (HCC) progression. Here, by performing RNA-sequencing (RNA-Seq) profiling, we sought to identify novel ncRNAs that potentially drive the heterogeneous progression of liver cancers. METHODS: RNA-Seq profiles were obtained from 68 HCC specimens and 10 samples of adjacent non-tumour liver tissues. The functional significance of the potential driver ncRNAs was evaluated by cell experiments. RESULTS: TPRG1-AS1 was identified as a potential driver noncoding RNA that promotes heterogeneous liver cancer progression. TPRG1-AS1 induced tumour suppressor RNA-binding motif protein 24 (RBM24), suppressing tumour growth by activating apoptotic tumour cell death. In addition, we report that TPRG1-AS1 acts as a competing endogenous RNA (ceRNA) for RBM24, sponging miR-4691-5p and miR-3659 to interfere with their binding to RBM24. CONCLUSIONS: We suggest that TPRG1-AS1 is a novel ceRNA sponging miR-4691-5p and miR-3659, resulting in RBM24 expression and suppression of liver cancer growth. Our results provide new insights into the functions of ncRNAs in heterogeneous HCC progression.

Mosaic CRISPR-stop enables rapid phenotyping of nonsense mutations in essential genes.

CRISPR-stop converts protein-coding sequences into stop codons, which, in the appropriate location, results in a null allele. CRISPR-stop induction in one-cell-stage zygotes generates Founder 0 (F0) mice that are homozygous mutants; this avoids mouse breeding and serves as a rapid screening approach for nonlethal genes. However, loss of function of 25% of mammalian genes causes early lethality. Here, we induced CRISPR-stop in one of the two blastomeres of the zygote, a method we name mosaic CRISPR-stop, to produce mosaic Atoh1 and Sox10 F0 mice; these mice not only survived longer than regular Atoh1/Sox10 knockout mice but also displayed their recognized cochlear phenotypes. Moreover, by using mosaic CRISPR-stop, we uncovered a previously unknown role of another lethal gene, Rbm24, in the survival of cochlear outer hair cells (OHCs), and we further validated the importance of Rbm24 in OHCs by using our Rbm24 conditional knockout model. Together, our results demonstrated that mosaic CRISPR-stop is reliable and rapid, and we believe this method will facilitate rapid genetic screening of developmentally lethal genes in the mouse inner ear and also in other organs.

Functional Role of the RNA-Binding Protein Rbm24a and Its Target sox2 in Microphthalmia.

Congenital eye defects represent a large class of disorders affecting roughly 21 million children worldwide. Microphthalmia and anophthalmia are relatively common congenital defects, with approximately 20% of human cases caused by mutations in SOX2. Recently, we identified the RNA-binding motif protein 24a (Rbm24a) which binds to and regulates sox2 in zebrafish and mice. Here we show that morpholino knockdown of rbm24a leads to microphthalmia and visual impairment. By utilizing sequential injections, we demonstrate that addition of exogenous sox2 RNA to rbm24a-deplete embryos is sufficient to suppress morphological and visual defects. This research demonstrates a critical role for understanding the post-transcriptional regulation of genes needed for development.

Rbm24 regulates inner-ear-specific alternative splicing and is essential for maintaining auditory and motor coordination.

Tissue-specific alternative splicing (AS) is emerging as one of the most exciting types of mechanisms associated with organ development and disease. In the auditory system, many hearing-related genes undergo AS, and errors in this process result in syndromic or non-syndromic hearing loss. However, little is known about the factors and mechanisms directing AS in the inner ear. In the present study, we identified a novel RNA-binding protein, Rbm24, which was critically involved in regulating inner-ear-specific AS. Rbm24 deletion resulted in hearing loss and defects in motor coordination. Global splicing analysis showed Rbm24 was required for correct splicing of a subset of pre-mRNA transcripts with essential roles in stereocilia integrity and survival of hair cells. Furthermore, we identified that Rbm24 directly regulated the splicing of Cdh23, a known disease gene responsible for human Usher syndrome 1D and non-syndromic autosomal recessive deafness DFNB12. In conclusion, our findings demonstrated that Rbm24 was a critical factor in regulating inner-ear-specific splicing and maintaining the hearing and motor coordination function of the inner ear. Our data not only offer mechanistic insights but also provide functional annotation of Rbm24 splicing targets that contribute to hearing loss.