Optimizing retinal ganglion cell nuclear staining for automated cell counting.

The retinal ganglion cells (RGCs) serve as the critical pathway for transmitting visual information from the retina to the brain, yet they can be dramatically impacted by diseases such as glaucoma. When investigating disease processes affecting RGCs in mouse models, accurately quantifying affected cells becomes essential. However, the use of pan RGC markers like RBPMS or THY1 presents challenges in accurate total cell counting. While Brn3a serves as a reliable RGC nuclear marker for automated counting, it fails to encompass all RGC subtypes in mice. To address this limitation and enable precise automated counting, our research endeavors to develop a method for labeling nuclei in all RGC subtypes. Investigating RGC subtypes labeled with the nuclear marker POU6F2 revealed that numerous RGCs unlabeled by Brn3a were, in fact, labeled with POU6F2. We hypothesize that using antibodies against both Brn3a and POU6F2 would label virtually all RGC nuclei in the mouse retina. Our experiments confirmed that staining retinas with both markers resulted in the labeling of all RGCs. Additionally, when using the cell body marker RBPMS known to label all mouse RGCs, all RBPMS-labeled cells also exhibited Brn3a or POU6F2 labeling. This combination of Brn3a and POU6F2 antibodies provides a pan-RGC nuclear stain, facilitating accurate automated counting by labeling cell nuclei in the retina.

RBPMS-AS1 sponges miR-19a-3p to restrain cervical cancer cells via enhancing PLCL1-mediated pyroptosis.

Cervical cancer (CC) poses a threat to human health. Enhancing pyroptosis can prevent the proliferation and epithelial-mesenchymal transition (EMT) of tumor cells. This study aims to reveal the candidates that modulate pyroptosis in CC. Accordingly, the common microRNAs (miRNAs/miRs) that were sponged by RBPMS antisense RNA 1 (RBPMS-AS1) and could target Phospholipase C-Like 1 (PLCL1) were intersected. The expression of PBPMS-AS1/miR-19a-3p (candidate miRNA)/PLCL1 was predicted in cervical squamous cell carcinoma (CESC), by which the expression location of RBPMS-AS1 and the binding between RBPMS-AS1/PLCL1 and miR-19a-3p were analyzed. The targeting relationship between RBPMS-AS1/PLCL1 and miR-19a-3p was confirmed by dual-luciferase reporter assay. After the transfection, cell counting kit-8 assay, colony formation assay, quantitative reverse transcription PCR, and Western blot were implemented for cell viability and proliferation analysis as well as gene and protein expression quantification analysis. Based on the results, RBPMS-AS1 and PLCL1 were lowly expressed, yet miR-19a-3p was highly expressed in CESC. RBPMS-AS1 overexpression diminished the proliferation and expressions of N-cadherin, vimentin, and miR-19a-3p, yet enhanced those of E-cadherin, PLCL1, and pyroptosis-relevant proteins (inteleukin-1ß, caspase-1, and gasdermin D N-terminal). However, the above RBPMS-AS1 overexpression-induced effects were counteracted in the presence of miR-19a-3p. There also existed a targeting relationship and negative interplay between PLCL1 and miR-19a-3p. In short, RBPMS-AS1 sponges miR-19a-3p and represses the growth and EMT of CC cells via enhancing PLCL1-mediated pyroptosis.

Loss of dmrt1 restores zebrafish female fates in the absence of cyp19a1a but not rbpms2a/b.

Sex determination and differentiation is a complex process regulated by multiple factors, including factors from the germline or surrounding somatic tissue. In zebrafish, sex-determination involves establishment of a bipotential ovary that undergoes sex-specific differentiation and maintenance to form the functional adult gonad. However, the relationships among these factors are not fully understood. Here, we identify potential Rbpms2 targets and apply genetic epistasis experiments to decipher the genetic hierarchy of regulators of sex-specific differentiation. We provide genetic evidence that the crucial female factor rbpms2 is epistatic to the male factor dmrt1 in terms of adult sex. Moreover, the role of Rbpms2 in promoting female fates extends beyond repression of Dmrt1, as Rbpms2 is essential for female differentiation even in the absence of Dmrt1. In contrast, female fates can be restored in mutants lacking both cyp19a1a and dmrt1, and prolonged in bmp15 mutants in the absence of dmrt1. Taken together, this work indicates that cyp19a1a-mediated suppression of dmrt1 establishes a bipotential ovary and initiates female fate acquisition. Then, after female fate specification, Cyp19a1a regulates subsequent oocyte maturation and sustains female fates independently of Dmrt1 repression.

Maternal RNA binding protein with multiple splicing 2 (RBPMS2) is involved in mouse blastocyst formation through the bone morphogenetic protein pathway.

RESEARCH QUESTION: Is early embryo development in mice influenced by RNA binding protein with multiple splicing 2 (RBPMS2), a maternal factor that accumulates and is stored in the cytoplasm of mature oocytes? DESIGN: The expression patterns of RBPMS2 in mouse were analysed using quantitative real-time PCR (qRT PCR) and immunofluorescence staining. The effect of knockdown of RBPMS2 on embryo development was evaluated through a microinjection of specific morpholino or small interfering RNA. RNA sequencing was performed for mechanistic analysis. The interaction between RBPMS2 and the bone morphogenetic protein (BMP) pathway was studied using BMP inhibitor and activator. The effect on the localization of E-cadherin was determined by immunofluorescence staining. RESULTS: Maternal protein RBPMS2 is highly expressed in mouse oocytes, and knockdown of RBPMS2 inhibits embryo development from the morula to the blastocyst stage. Mechanistically, RNA sequencing showed that the differentially expressed genes were enriched in the transforming growth factor-ß (TGF-ß) signalling pathway. BMPs are members of the TGF-ß superfamily of growth factors. It was found that the addition of BMP inhibitor to the culture medium led to a morula-stage arrest, similar to that seen in RBPMS2 knockdown embryos. This morula-stage arrest defect caused by RBPMS2 knockdown was partially rescued by BMP activator. Furthermore, the localization of E-cadherin to the membrane was impaired in response to a knockdown of RBPMS2 or inhibition of the BMP pathway. CONCLUSION: This study suggests that RBPMS2 activates the BMP pathway and thus influences the localization of E-cadherin, which is important for early mouse embryo development during blastocyst formation.

Loss of RBPMS in ovarian cancer compromises the efficacy of EGFR inhibitor gefitinib through activating HER2/AKT/mTOR/P70S6K signaling.

RBPMS may be a tumor suppressor in cancer, but its impact in modulation of drug sensitivity is unclear. This study aimed to investigate the regulatory role of RBPMS in cellular response to EGFR inhibitor gefitinib in ovarian cancer (OC). By western blotting assay, we revealed RBPMS was down-regulated in epithelial ovarian cancer tissues compared to normal control ovarian epithelial tissues. Overexpression of RBPMS inhibited cell viability and proliferation, and conferred gefitinib sensitivity, accompanied by reduced expression of p-EGFR, and vice versa. Proteomic analysis and flow cytometry experiments showed that RBPMS induced S-stage cell cycle arrest in gefitinib-treated OC cells. Co-IP assay suggested that HER2 was a downstream target of RBPMS, and RBPMS negatively regulated HER2 expression. HER2 counteracted the stimulation of RBPMS to cell growth blocking, gefitinib sensitivity and cell cycle arrest. We further demonstrated that RBPMS overexpression suppressed the activation of p-AKT, p-mTOR and p-P70S6K, which was rescued by up-regulation of HER2. The combination of AKT inhibitor MK2206 and gefitinib had a synergistic effect on OC cells with high level of RBPMS. In conclusion, through the direct inhibition of HER2/AKT/mTOR/P70S6K pathway, RBPMS may be a potential therapeutic target for improving gefitinib sensitivity in OC.

Increased Expression of the RBPMS Splice Variants Inhibits Cell Proliferation in Ovarian Cancer Cells.

RNA-Binding Protein with Multiple Splicing (RBPMS) is a member of family proteins that bind to nascent RNA transcripts and regulate their splicing, localization, and stability. Evidence indicates that RBPMS controls the activity of transcription factors associated with cell growth and proliferation, including AP-1 and Smads. Three major RBPMS protein splice variants (RBPMSA, RBPMSB, and RBPMSC) have been described in the literature. We previously reported that reduced RBPMS levels decreased the sensitivity of ovarian cancer cells to cisplatin treatment. However, little is known about the biological role of the RBPMS splice variants in ovarian cancer cells. We performed RT-PCR and Western blots and observed that both RBPMSA and RBPMSC are reduced at the mRNA and protein levels in cisplatin resistant as compared with cisplatin sensitive ovarian cancer cells. The mRNA and protein levels of RBPMSB were not detectable in any of the ovarian cancer cells tested. To better understand the biological role of each RBPMSA and RBPMSC, we transfected these two splice variants in the A2780CP20 and OVCAR3CIS cisplatin resistant ovarian cancer cells and performed cell proliferation, cell migration, and invasion assays. Compared with control clones, a significant reduction in the number of colonies, colony size, cell migration, and invasion was observed with RBPMSA and RBPMSC overexpressed cells. Moreover, A2780CP20-RBPMSA and A2780CP20-RBPMSC clones showed reduced senescence-associated ß-galactosidase (ß-Gal)-levels when compared with control clones. A2780CP20-RBPMSA clones were more sensitive to cisplatin treatment as compared with A2780CP20-RBPMSC clones. The A2780CP20-RBPMSA and A2780CP20-RBPMSC clones subcutaneously injected into athymic nude mice formed smaller tumors as compared with A2780CP20-EV control group. Additionally, immunohistochemical analysis showed lower proliferation (Ki67) and angiogenesis (CD31) staining in tissue sections of A2780CP20-RBPMSA and A2780CP20-RBPMSC tumors compared with controls. RNAseq studies revealed many common RNA transcripts altered in A2780CP20-RBPMSA and A2780CP20-RBPMSC clones. Unique RNA transcripts deregulated by each RBPMS variant were also observed. Kaplan-Meier (KM) plotter database information identified clinically relevant RBPMSA and RBPMSC downstream effectors. These studies suggest that increased levels of RBPMSA and RBPMSC reduce cell proliferation in ovarian cancer cells. However, only RBPMSA expression levels were associated with the sensitivity of ovarian cancer cells to cisplatin treatment.

Prognostic value of noggin protein expression in patients with resected gastric cancer.

BACKGROUND: Noggin and RNA-binding protein for multiple splicing 2 (RBPMS2) are known to regulate the expression of smooth muscle cells, endothelial cells, and osteoblasts. However, the prognostic role of combined Noggin and RBPMS2 expression in resected gastric cancer (GC) is unclear. METHODS: A total of 163 patients with GC who underwent gastrectomy were included in this study. The expression of Noggin and RBPMS2 proteins in tumor cells at the tumor center and invasive front of resected GC was evaluated by immunohistochemistry, and in conjunction with clinicopathological parameters the patient survival was analyzed. RESULTS: RBPMS2 protein expression was high at the tumor center (n = 86, 52.8%) and low at the invasive front (n = 69, 42.3%), while Noggin protein expression was high in both tumor center (n = 91, 55.8%) and the invasive front (n = 90, 55.2%). Noggin expression at the invasive front and tumor center was significantly decreased in advanced T stage, non-intestinal-type (invasive front, P = 0.008 and P <  0.001; tumor center lesion, P = 0.013 and P = 0.001). RBPMS2 expression at the invasive front was significantly decreased in non-intestinal-type and positive lymphatic invasion (P <  0.001 and P = 0.013). Multivariate analysis revealed that high Noggin protein expression of the invasive front was an independent prognostic factor for overall survival (hazard ratio [HR], 0.58; 95% confidence interval [CI]; 0.35-0.97, P <  0.036), but not at the tumor center (HR, 1.35; 95% CI; 0.81-2.26, P = 0.251). CONCLUSIONS: Our study indicates that high Noggin expression is a crucial prognostic factor for favorable outcomes in patients with resected GC.

Exploring the Activities of RBPMS Proteins in Myocardial Biology.

Numerous RNA-binding proteins (RBPs) are expressed in the heart, and mutations in several RBPs have been implicated in cardiovascular disease through genetic associations, animal modeling, and mechanistic studies. However, the functions of many more cardiac RBPs, and their relevance to disease states, remain to be elucidated. Recently, we have initiated studies to characterize the functions of the RBPs RBPMS and RBPMS2 in regulating myocardial biology in zebrafish and higher vertebrate species. These studies began when we learned, using an unbiased gene discovery approach, that rbpms2a and rbpms2b in zebrafish are robust markers of embryonic myocardium. This observation, which is consistent with published data, suggests that the encoded proteins are likely to be performing critical functions in regulating one or more aspects of cardiomyocyte differentiation, proliferation, survival, and/or contractility. This notion is supported by recent reports demonstrating that zebrafish embryos with disrupted Rbpms2 function exhibit gross signs of cardiac distress. Interestingly, a 20-year-old study determined that myocardial tissue from the frog, chick, and mouse also express high levels of Rbpms and/or Rbpms2, which is suggestive of evolutionary conservation of function. In this review, we will provide a historical account of how RBPMS and RBPMS2 genes were discovered, attempt to clarify some potentially confusing nomenclature, and summarize published observations that inform our ongoing studies.

Expanding the spectrum of pediatric NTRK-rearranged fibroblastic tumors to the central nervous system: A case report with RBPMS-NTRK3 fusion.

We report a case of a 20-month-old male presenting with seizures who was found to have a hyperintense lesion on T2-weighted images of magnetic resonance imaging in the left medial temporal lobe that was initially clinically and radiologically thought to be either low-grade glioma or focal cortical dysplasia. Histologic, immunohistochemical and molecular evaluation (array comparative genomic hybridization, Archer fusion panel) of the resection specimen demonstrated a highly infiltrative fibroblastic spindle cell neoplasm with mild nuclear atypia and an RBPMS-NTRK3 fusion. NTRK-fused mesenchymal tumors are known to involve extracranial sites but, to our knowledge, have not been described within the central nervous system. Accurate and timely diagnosis of this entity has important prognostic and therapeutic implications, as NTRK-fused tumors may recur locally and may respond to selective kinase inhibitor therapies.

Oocyte polarity requires a Bucky ball-dependent feedback amplification loop.

In vertebrates, the first asymmetries are established along the animal-vegetal axis during oogenesis, but the underlying molecular mechanisms are poorly understood. Bucky ball (Buc) was identified in zebrafish as a novel vertebrate-specific regulator of oocyte polarity, acting through unknown molecular interactions. Here we show that endogenous Buc protein localizes to the Balbiani body, a conserved, asymmetric structure in oocytes that requires Buc for its formation. Asymmetric distribution of Buc in oocytes precedes Balbiani body formation, defining Buc as the earliest marker of oocyte polarity in zebrafish. Through a transgenic strategy, we determined that excess Buc disrupts polarity and results in supernumerary Balbiani bodies in a 3'UTR-dependent manner, and we identified roles for the buc introns in regulating Buc activity. Analyses of mosaic ovaries indicate that oocyte pattern determines the number of animal pole-specific micropylar cells that are associated with an egg via a close-range signal or direct cell contact. We demonstrate interactions between Buc protein and buc mRNA with two conserved RNA-binding proteins (RNAbps) that are localized to the Balbiani body: RNA binding protein with multiple splice isoforms 2 (Rbpms2) and Deleted in azoospermia-like (Dazl). Buc protein and buc mRNA interact with Rbpms2; buc and dazl mRNAs interact with Dazl protein. Cumulatively, these studies indicate that oocyte polarization depends on tight regulation of buc: Buc establishes oocyte polarity through interactions with RNAbps, initiating a feedback amplification mechanism in which Buc protein recruits RNAbps that in turn recruit buc and other RNAs to the Balbiani body.

Temporal expression of CD184(CXCR4) and CD171(L1CAM) identifies distinct early developmental stages of human retinal ganglion cells in embryonic stem cell derived retina.

Human retinal ganglion cells (RGCs) derived from pluripotent stem cells (PSCs) have anticipated value for human disease study, drug screening, and therapeutic applications; however, their full potential remains underdeveloped. To characterize RGCs in human embryonic stem cell (hESC) derived retinal organoids we examined RGC markers and surface antigen expression and made comparisons to human fetal retina. RGCs in both tissues exhibited CD184 and CD171 expression and distinct expression patterns of the RGC markers BRN3 and RBPMS. The retinal progenitor cells (RPCs) of retinal organoids expressed CD184, consistent with its expression in the neuroblastic layer in fetal retina. In retinal organoids CD184 expression was enhanced in RGC competent RPCs and high CD184 expression was retained on post-mitotic RGC precursors; CD171 was detected on maturing RGCs. The differential expression timing of CD184 and CD171 permits identification and enrichment of RGCs from retinal organoids at differing maturation states from committed progenitors to differentiating neurons. These observations will facilitate molecular characterization of PSC-derived RGCs during differentiation, critical knowledge for establishing the veracity of these in vitro produced cells. Furthermore, observations made in the retinal organoid model closely parallel those in human fetal retina further validating use of retinal organoid to model early retinal development.

The RNA-binding protein RBPMS1 represses AP-1 signaling and regulates breast cancer cell proliferation and migration.

The activator protein-1 (AP-1) transcription factor complex plays a crucial role in tumor growth and progression. However, how AP-1 transcriptional activity is repressed is not fully understood. Here, we show that RNA-binding protein with multiple splicing 1 (RBPMS1) physically and functionally interacts with AP-1 in vitro and in vivo. The RNA-recognition motif (RRM) and C-terminus of the RBPMS1 isoforms RBPMS1A and RBPMS1C, but not RBPMS1B, interacted with cFos, a member of the AP-1 family that dimerizes with cJun to stimulate AP-1 transcriptional activity. RBPMS1 did not associate with Jun proteins. RBPMS1A and RBPMS1C bound to the basic leucine zipper (bZIP) domain of cFos that mediates dimerization of AP-1 proteins. In addition, RBPMS1A-C interacted with the transcription factor Smad3, which was shown to interact with cJun and increase AP-1 transcriptional activity. RBPMS1 inhibited c-Fos or Smad3-mediated AP-1 transactivation and the expression of AP-1 target genes known to be the key regulators of cancer growth and progression, including vascular endothelial growth factor (VEGF) and cyclin D1. Mechanistically, RBPMS1 blocks the formation of the cFos/cJun or Smad3/cJun complex as well as the recruitment of cFos or Smad3 to the promoters of AP-1 target genes. In cultured cells and a mouse xenograft model, RBPMS1 inhibited the growth and migration of breast cancer cells through c-Fos or Smad3. These data suggest that RBPMS1 is a critical repressor of AP-1 signaling and RBPMS1 activation may be a useful strategy for cancer treatment.

Identification of the RNA recognition element of the RBPMS family of RNA-binding proteins and their transcriptome-wide mRNA targets.

Recent studies implicated the RNA-binding protein with multiple splicing (RBPMS) family of proteins in oocyte, retinal ganglion cell, heart, and gastrointestinal smooth muscle development. These RNA-binding proteins contain a single RNA recognition motif (RRM), and their targets and molecular function have not yet been identified. We defined transcriptome-wide RNA targets using photoactivatable-ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP) in HEK293 cells, revealing exonic mature and intronic pre-mRNA binding sites, in agreement with the nuclear and cytoplasmic localization of the proteins. Computational and biochemical approaches defined the RNA recognition element (RRE) as a tandem CAC trinucleotide motif separated by a variable spacer region. Similar to other mRNA-binding proteins, RBPMS family of proteins relocalized to cytoplasmic stress granules under oxidative stress conditions suggestive of a support function for mRNA localization in large and/or multinucleated cells where it is preferentially expressed.

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.

lncRNA Biomarkers of Glioblastoma Multiforme.

Long noncoding RNAs (lncRNAs) are RNA molecules of 200 nucleotides or more in length that are not translated into proteins. Their expression is tissue-specific, with the vast majority involved in the regulation of cellular processes and functions. Many human diseases, including cancer, have been shown to be associated with deregulated lncRNAs, rendering them potential therapeutic targets and biomarkers for differential diagnosis. The expression of lncRNAs in the nervous system varies in different cell types, implicated in mechanisms of neurons and glia, with effects on the development and functioning of the brain. Reports have also shown a link between changes in lncRNA molecules and the etiopathogenesis of brain neoplasia, including glioblastoma multiforme (GBM). GBM is an aggressive variant of brain cancer with an unfavourable prognosis and a median survival of 14-16 months. It is considered a brain-specific disease with the highly invasive malignant cells spreading throughout the neural tissue, impeding the complete resection, and leading to post-surgery recurrences, which are the prime cause of mortality. The early diagnosis of GBM could improve the treatment and extend survival, with the lncRNA profiling of biological fluids promising the detection of neoplastic changes at their initial stages and more effective therapeutic interventions. This review presents a systematic overview of GBM-associated deregulation of lncRNAs with a focus on lncRNA fingerprints in patients' blood.

Transcriptomic profiling of retinal cells reveals a subpopulation of microglia/macrophages expressing Rbpms marker of retinal ganglion cells (RGCs) that confound identification of RGCs.

Analysis of retinal ganglion cells (RGCs) by scRNA-seq is emerging as a state-of-the-art method for studying RGC biology and subtypes, as well as for studying the mechanisms of neuroprotection and axon regeneration in the central nervous system (CNS). Rbpms has been established as a pan-RGC marker, and Spp1 has been established as an αRGC type and macrophage marker. Here, we analyzed by scRNA-seq retinal microglia and macrophages, and found Rbpms+ subpopulations of retinal microglia/macrophages, which pose a potential pitfall in scRNA-seq studies involving RGCs. We performed comparative analysis of cellular identity of the presumed RGC cells isolated in recent scRNA-seq studies, and found that Rbpms+ microglia/macrophages confounded identification of RGCs. We also showed using immunohistological analysis that, Rbpms protein localizes to stress granules in a subpopulation of retinal microglia after optic nerve injury, which was further supported by bioinformatics analysis identifying stress granule-associated genes enriched in the Rbpms+ microglia/macrophages. Our findings suggest that the identification of Rbpms+ RGCs by immunostaining after optic nerve injury should exclude cells in which Rbpms signal is restricted to a subcellular granule, and include only those cells in which the Rbpms signal is labeling cell soma diffusely. Finally, we provide solutions for circumventing this potential pitfall of Rbpms-expressing microglia/macrophages in scRNA-seq studies, by including in RGC and αRGC selection criteria other pan-RGC and αRGC markers.

Overexpressed RBPMS-AS1 increased cell radiosensitivity by sponging miR-19a-3p in lung cancer cell lines (A549 and SK-MES-1) via regulating PTEN/AKT axis.

PURPOSE: This paper intended to study RBPMS-AS1 in lung cancer (LC) radiosensitivity. MATERIALS AND METHODS: LC cells were transfected with RBPMS-AS1 overexpression plasmid and miR-19a-3p mimic and treated with radiation. PTEN, AKT, p-AKT, RBPMS-AS1, and miR-19a-3p expressions were detected via Western blot and qRT-PCR. The localization of RBPMS-AS1 in cells was determined through fluorescence in situ hybridization assay. The targeting relationships of RBPMS-AS1 and miR-19a-3p/miR-19a-3p and PTEN were determined through RIP and dual luciferase reporter analysis. Cell survival, viability, and apoptosis were assessed through colony formation, CCK-8, and flow-cytometry assays. RESULTS: RBPMS-AS1 was downregulated in LC and mainly distributed in cytoplasm. RBPMS-AS1 targeted miR-19a-3p in LC cells. Radiation suppressed LC cell survival, viability, and induced apoptosis, as overexpressed RBPMS-AS1 performed the similar effects and enhanced those effects induced by radiation. MiR-19a-3p mimic reversed the effect of overexpressed RBPMS-AS1 on enhancing radiation-induced LC cell apoptosis. MiR-19a-3p targeted PTEN and miR-19a-3p mimic reversed the effect of overexpressed RBPMS-AS1 on PTEN and phosphorylation of AKT in LC cells. CONCLUSION: Overexpressed RBPMS-AS1 sponged miR-19a-3p to increase cell radiosensitivity in LC via regulating PTEN/AKT axis.

Pan-cancer tRNA-derived fragment CAT1 coordinates RBPMS to stabilize NOTCH2 mRNA to promote tumorigenesis.

Transfer RNA-derived fragments (tRFs) are a class of small non-coding regulatory RNAs that are involved in the pathophysiology of many diseases. However, the role of tRFs in cancer progression remains largely elusive. Here, we demonstrate that a pan-cancer 3'-tRF, CAT1 (cancer associated tRF 1), is ubiquitously upregulated in tumors and associated with poor prognosis of a variety of cancers, including lung cancer. The upregulated CAT1 in cancer cells binds to RNA-binding protein with multiple splicing (RBPMS) and displaces NOTCH2 association from RBPMS, thereby inhibiting the subsequent CCR4-NOT deadenylation-complex-mediated NOTCH2 mRNA decay. The CAT1-enhanced NOTCH2 expression promotes lung cancer cell proliferation and metastasis in vitro and in vivo. In addition, plasma CAT1 levels are substantially increased in patients with lung cancer compared to non-cancer control subjects. Our findings reveal an intrinsic connection between cancer-specific upregulation of CAT1 and cancer progression, show the regulation of NOTCH signaling in cancer by a 3'-tRF, and highlight its great clinical potential.

Pan-retinal ganglion cell markers in mice, rats, and rhesus macaques.

Univocal identification of retinal ganglion cells (RGCs) is an essential prerequisite for studying their degeneration and neuroprotection. Before the advent of phenotypic markers, RGCs were normally identified using retrograde tracing of retinorecipient areas. This is an invasive technique, and its use is precluded in higher mammals such as monkeys. In the past decade, several RGC markers have been described. Here, we reviewed and analyzed the specificity of nine markers used to identify all or most RGCs, i.e., pan-RGC markers, in rats, mice, and macaques. The best markers in the three species in terms of specificity, proportion of RGCs labeled, and indicators of viability were BRN3A, expressed by vision-forming RGCs, and RBPMS, expressed by vision- and non-vision-forming RGCs. NEUN, often used to identify RGCs, was expressed by non-RGCs in the ganglion cell layer, and therefore was not RGC-specific. γ-SYN, TUJ1, and NF-L labeled the RGC axons, which impaired the detection of their somas in the central retina but would be good for studying RGC morphology. In rats, TUJ1 and NF-L were also expressed by non-RGCs. BM88, ERRß, and PGP9.5 are rarely used as markers, but they identified most RGCs in the rats and macaques and ERRß in mice. However, PGP9.5 was also expressed by non-RGCs in rats and macaques and BM88 and ERRß were not suitable markers of viability.

DNA methylation of RNA-binding protein for multiple splicing 2 functions as diagnosis biomarker in gastric cancer pathogenesis and its potential clinical significance.

Higher methylation levels of RNA-binding protein for multiple splicing 2 (RBPMS2) was reported to be related with unfavorable outcome in gastric cancer (GC). However, molecular function and diagnostic significance of DNA methylation of RBPMS2 remains indistinct. Here we aimed to whether DNA methylation of RBPMS2 acts as a diagnosis biomarker in GC pathogenesis and its potential clinical significance. Western blot and immunochemistry assays were carried out to explore the level of RBPMS2. GC malignancy behaviors were determined by cell counting kit-8, Transwell, flow cytometry analysis and terminal-deoxynucleoitidyl transferase mediated nick end labeling staining. The inflammatory cell infiltration in xenograft model was observed by hematoxylin and eosin staining. CpG Islands was predicted by MethPrimer and the DNA methylation of RBPMS2 was evaluated by methylation-specific polymerase chain reaction. The results showed that RBPMS2 was downregulated in GC specimens. Poor survival rates were associated with low RBPMS2 expression. Overexpression of RBPMS2 inhibited GC growth while facilitated apoptosis in GC cells. In addition, level of DNA methylation of RBPMS2 in GC tissues was increased and DNA methylation of RBPMS2 was strongly associated with tumor invasion, Borrmann classification and TNM stage. We also observed that DNA methylation inhibitors counteracted the role of RBPMS2 in restraining GC development and tumorigenesis. To sum, our data demonstrated that DNA methylation of RBPMS2 was responsible for its downregulation in GC and promoted tumor progression, indicating DNA methylation of RBPMS2 might serve as a valuable potential parameter in GC pathogenesis.