RBM8A Human

What is RBM8A and what are its primary functions in human cells?

RBM8A is an RNA-binding protein involved in several crucial cellular processes including RNA transcription, translation, cell cycle regulation, and apoptosis regulation . It serves as a core component in post-transcriptional regulation networks, participating in ribosome signaling, RNA transport, mRNA surveillance, and spliceosome signaling pathways . The protein's physiological function is closely related to RNA splicing and protein translation, positioning it as a critical factor in gene expression regulation.

Where is RBM8A primarily expressed in human cells?

RBM8A primarily localizes to the nucleus in human cells, as demonstrated through immunohistochemical analysis of glioblastoma specimens . This nuclear localization aligns with its role in RNA processing and transcriptional regulation. The protein forms part of the exon junction complex (EJC) which assembles near exon-exon junctions on mRNAs during splicing, highlighting its importance in RNA maturation processes.

What cellular pathways are most affected by RBM8A alterations?

Enrichment analysis of RBM8A functional networks reveals that the protein significantly impacts several cellular pathways. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis shows that RBM8A alterations primarily affect ribosome signaling, RNA transport, mRNA surveillance, and spliceosome signaling pathways . In cancer contexts such as glioblastoma, RBM8A has been shown to regulate the Notch1/STAT3 signaling network, influencing cellular proliferation and invasion capabilities .

How is RBM8A expression altered across different cancer types?

Analysis of transcriptional sequencing data from clinical samples in GEO and TCGA databases has revealed that RBM8A is significantly dysregulated in multiple cancer types. In hepatocellular carcinoma (HCC), RBM8A mRNA levels and copy number variations (CNVs) are significantly higher than in normal liver tissue . Similarly, in glioblastoma (GBM), RBM8A expression is substantially elevated compared to control samples . The fold change in expression is consistent across various geographic regions and ethnic HCC studies, ranking RBM8A among the top 3-4% of all genes upregulated in HCC based on CNVs .

What types of RBM8A alterations are observed in hepatocellular carcinoma?

RBM8A alterations in HCC show distinct patterns and frequencies. Analysis of sequencing data from the TCGA database revealed that RBM8A was altered in 22% (80/370) of LIHC patients, with various types of alterations observed :

This data indicates that amplification is the most common type of RBM8A copy number variation in HCC, suggesting that altered RBM8A expression and dysfunction may result from chromosomal structure changes .

How does RBM8A expression correlate with cancer prognosis?

What are effective methods for knocking down RBM8A expression in experimental models?

Researchers have successfully employed lentiviral-based RNA interference approaches to knock down RBM8A expression. Specifically:

The methodology involves infecting target cells with these shRNA-encoding lentiviruses at a multiplicity of infection of 1 using Polybrene (8 μg/ml) . GFP expression should be confirmed approximately 72 hours after viral infection, followed by selection in puromycin-containing medium (4 μg/ml) for a minimum of 14 days. Puromycin-resistant cells should then be amplified in medium containing reduced puromycin (2 μg/ml) for 7-9 days before culture in puromycin-free medium . Knockdown efficiency should be validated using qRT-PCR and western blot analyses.

How can RBM8A overexpression be achieved in cell culture models?

For RBM8A overexpression studies, researchers have utilized retroviral expression systems. The methodology involves engineering an expression plasmid encoding FLAG-tagged RBM8A using the pMSCV-IRES-GFP vector . These plasmids are then transfected into packaging cells (such as 293T cells) to produce recombinant retroviruses. Target cells are subsequently infected with these retroviruses at a multiplicity of infection of 1, generating stable RBM8A-overexpressing cell lines . This approach provides a controlled system for studying the effects of elevated RBM8A expression on cellular phenotypes and molecular pathways.

What in vivo models are suitable for studying RBM8A function in cancer?

Orthotopic xenograft models have proven effective for studying RBM8A function in cancer. For GBM research, RBM8A-KD or control GBM cells are intracranially implanted into the corpus striatum of anesthetized athymic nude mice (typically 6-week-old) using a stereotactic frame for small animals . Tumor development and growth are monitored using magnetic resonance imaging (MRI). This approach allows for assessment of how RBM8A affects tumor growth in a physiologically relevant microenvironment. Studies have shown that mice inoculated with RBM8A knockdown cells developed significantly smaller tumors compared to control cells, with average tumor diameters of 0.25 ± 0.15 mm³ versus 11.92 ± 4.98 mm³, respectively (p=0.001) .

How does RBM8A interact with the Notch signaling pathway?

RBM8A demonstrates significant interaction with the Notch signaling pathway through multiple mechanisms. Molecular docking studies have revealed that RBM8A can bind to genes whose transcription is regulated by C promoter-binding factor 1 (CBF1) . CBF1 is a known interactor with the Notch1 receptor and plays a critical role in activating the Notch signaling pathway. Functional studies have confirmed that RBM8A overexpression significantly enhances activation of the CBF1 luciferase reporter in GBM cells, while RBM8A knockdown attenuates this activation .

These findings suggest that RBM8A is sufficient to activate the Notch signaling pathway, potentially through direct or indirect regulation of CBF1 activity. The molecular mechanism appears to involve RBM8A promoting the proliferation and migration of GBM cells by activating the Notch/STAT3 pathway, establishing a functional link between RBM8A and this critical cancer-associated signaling network .

What are the downstream effects of RBM8A modulation on cellular signaling networks?

RBM8A modulation induces substantial changes in multiple signaling cascades. Western blot analyses have revealed that RBM8A knockdown decreases levels of Notch, phospho-STAT3, and phospho-H3 proteins in GBM cells . Conversely, RBM8A overexpression increases the levels of these signaling proteins.

The γ-secretase inhibitor DAPT, which blocks Notch signaling, significantly reverses RBM8A-enhanced GBM cell proliferation and invasion capabilities . This reversal is associated with down-regulation of p-STAT3 and Notch1 protein levels, confirming that the Notch/STAT3 axis is a critical downstream mediator of RBM8A function. These findings establish RBM8A as an upstream regulator of multiple cancer-associated signaling pathways, with particularly strong effects on the Notch/STAT3 network.

How do genomic alterations in RBM8A affect broader gene expression networks?

Genomic alterations in RBM8A significantly impact broader gene expression networks across multiple cellular pathways. Gene ontology (GO) analysis of RBM8A-associated genes indicates that these genes primarily encode proteins localized to the cytosol, ribosome, and ribosomal subunits . Functionally, these proteins are primarily involved in viral gene expression and RNA catabolism, while also serving as structural constituents of ribosomes and participating in mRNA binding .

The biological interaction network of RBM8A alterations appears to be involved in the core node of post-transcriptional regulation, closely related to RNA splicing and protein translation . This aligns with the physiological function of RBM8A and suggests that alterations in this gene can trigger widespread changes across multiple RNA processing and translational networks, potentially contributing to disease states when dysregulated.

What is the potential of RBM8A as a diagnostic biomarker in cancer?

RBM8A shows considerable promise as a diagnostic biomarker for multiple cancer types. In hepatocellular carcinoma, RBM8A overexpression has been documented across diverse geographic regions and ethnic populations, suggesting it may be a universally applicable marker . Early detection of HCC remains challenging, with traditional markers like Alpha-fetoprotein (AFP) being positive in only approximately 70% of HCC patients. RBM8A could potentially complement existing biomarkers to improve early diagnosis accuracy .

Additionally, the gene set variance analysis score of genes involved in regulation of the Notch1/STAT3 network by RBM8A has shown good diagnostic value for glioblastoma . The consistent upregulation of RBM8A across multiple cancer types positions it as a potentially valuable addition to diagnostic panels, particularly for malignancies where early detection remains challenging.

How might targeting RBM8A improve cancer therapeutic strategies?

Targeting RBM8A offers several promising therapeutic avenues for cancer treatment. Experimental evidence shows that knockdown of RBM8A inhibits GBM progression and invasion ability both in vitro and in vivo . The γ-secretase inhibitor DAPT, which blocks Notch signaling, significantly reverses RBM8A-enhanced GBM cell proliferation and invasion .

This suggests that combination therapies targeting both RBM8A and the Notch pathway might provide synergistic benefits. Additionally, since RBM8A appears to function upstream of multiple cancer-associated signaling networks, including Notch/STAT3, developing specific inhibitors against RBM8A could potentially disrupt multiple oncogenic pathways simultaneously, offering a more comprehensive therapeutic approach than targeting individual downstream pathways .

What is the relationship between RBM8A expression and tumor grade/progression?

A clear relationship exists between RBM8A expression levels and tumor aggressiveness across multiple cancer types. In glioblastoma patients, RBM8A expression strongly correlates with tumor grade:

This data demonstrates that a large proportion of patients with low-grade tumors (78.57%) exhibit low RBM8A expression, while those with high-grade tumors (66.25%) predominantly show high RBM8A expression . The correlation between elevated RBM8A expression and advanced tumor grade suggests that RBM8A may play an important role in tumor progression and could potentially serve as a marker for assessing malignancy potential and disease progression.