RBM45 is an RNA-binding protein with roles in neural development by regulating RNA splicing. Its dysfunction and aggregation are associated with neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS) and frontotemporal lobar dementia (FTLD). RBM45 harbors three RRM domains that potentially bind RNA. While the recognitions of RNA by its N-terminal tandem RRM domains (RRM1 and RRM2) have been well understood, the RNA-binding property of its C-terminal RRM (RRM3) remains unclear. In this work, we identified that the RRM3 of RBM45 sequence-specifically binds RNA with a GACG sequence, similar but not identical to those recognized by the RRM1 and RRM2. Further, we determined the crystal structure of RBM45RRM3 in complex with a GACG sequence-containing single-stranded DNA. Our structural results, together with the RNA-binding assays of mutants at key amino acid residues, revealed the molecular mechanism by which RBM45RRM3 recognizes an RNA sequence. Our finding on the RNA-binding property of the individual RRM module of RBM45 provides the foundation for unraveling the RNA-binding characteristics of full-length RBM45 and for understanding the biological functions of RBM45.

Alternative splicing is a complex gene regulatory process that distinguishes itself from canonical splicing by rearranging the introns and exons of an immature pre-mRNA transcript. This process plays a vital role in enhancing transcriptomic and proteomic diversity from the genome. Alternative splicing has emerged as a pivotal mechanism governing complex biological processes during both heart development and the development of cardiovascular diseases. Multiple alternative splicing factors are involved in a synergistic or antagonistic manner in the regulation of important genes in relevant physiological processes. Notably, circular RNAs have only recently garnered attention for their tissue-specific expression patterns and regulatory functions. This resurgence of interest has prompted a reevaluation of the topic. Here, we provide an overview of our current understanding of alternative splicing mechanisms and the regulatory roles of alternative splicing factors in cardiovascular development and pathological process of different cardiovascular diseases, including cardiomyopathy, myocardial infarction, heart failure and atherosclerosis.

Ataxin-2 (ATXN2) was originally discovered in the context of spinocerebellar ataxia type 2 (SCA2), but it has become a key player in various neurodegenerative diseases. This review delves into the multifaceted roles of ATXN2 in human diseases, revealing its diverse molecular and cellular pathways. The impact of ATXN2 on diseases extends beyond functional outcomes; it mainly interacts with various RNA-binding proteins (RBPs) to regulate different stages of post-transcriptional gene expression in diseases. With the progress of research, ATXN2 has also been found to play an important role in the development of various cancers, including breast cancer, gastric cancer, pancreatic cancer, colon cancer, and esophageal cancer. This comprehensive exploration underscores the crucial role of ATXN2 in the pathogenesis of diseases and warrants further investigation by the scientific community. By reviewing the latest discoveries on the regulatory functions of ATXN2 in diseases, this article helps us understand the complex molecular mechanisms of a series of human diseases related to this intriguing protein.

BACKGROUND: LINC-PINT was downregulated in nasopharyngeal carcinoma (NPC) and correlated with treatment efficiency of NPC. However, the underlying mechanism of LINC-PINT in NPC has not yet been fully explored.
METHODS: We used CellTiter luminescent assay, clone formation assay, Hoechst staining, and SYTO-9/PI staining to examine cell viability and cell apoptosis regulated by LINC-PINT in NPC cells. Xenograft tumor model, HE staining, Ki67 staining, and TUNEL assay were conducted to assess the role of LINC-PINT in vivo. Bioinformatics and RNA immunoprecipitation assay was performed to identify the binding protein of LINC-PINT. Fluorescence in situ hybridization and immunofluorescence were utilized to measure the colocalization of XRCC6 with LINC-PINT and DNA-PKcs. Mito-Tracker red CMXRos staining was used to label mitochondria in cells specifically.
RESULTS: We found LINC-PINT was downregulated in many tumors (including NPC) and associated with poor prognosis. The cell viability was significantly inhibited and cell apoptosis was remarkably promoted in LINC-PINT overexpressed cells in contrast to control cells. The growth of tumor xenografts was significantly suppressed and the tumor weight was significantly decreased in LINC-PINT overexpression group compared to the control group. Correspondingly, the positive Ki67 foci was decreased while TUNEL foci was increased in LINC-PINT overexpression group. Mechanically, we verified XRCC6 as a new binding protein of LINC-PINT through RNA binding domains prediction, RIP and colocalization of LINC-PINT and XRCC6. By binding to XRCC6, LINC-PINT interfered the formation of DNA-PK complex, regulated mitochondria accumulation status and affected the modification of apoptosis proteins, leading to more cell apoptosis.
CONCLUSIONS: Our study provided the first evidence that LINC-PINT promotes cell apoptosis in NPC by binding to XRCC6 and affecting its function.

The RNA-binding PUF proteins are post-transcriptional regulators found throughout the eukaryotic domain. In Trypanosoma cruzi, ten Puf genes termed Puf1 to Puf10 have been identified. Considering that the control of gene expression in this parasite is mainly at the post-transcriptional level, we characterized the PUF3 protein by knocking out and overexpressing the gene in T. cruzi epimastigotes and studied different genetic and biological features. The RNA-seq analyses in both genotypes showed significant changes in the number of regulated transcripts compared with wild-type parasites. Thus, the number of differentially expressed genes in the knockout (ΔTcPuf3) and the overexpressor (pTEXTcPuf3) were 238 and 187, respectively. In the knockout, a more significant proportion of genes was negatively regulated (166 out of 238). In contrast, in the overexpressor, positively regulated genes were predominant (149 out of 170). Additionally, when we predicted the subcellular location of the differentially expressed genes, the results revealed an important representation of nuclear genes encoding mitochondrial proteins. Therefore, we determined whether overexpression or knockout of TcPuf3 could lead to changes in both mitochondrial structure and cellular respiration. When mitochondria from ΔTcPuf3 and pTEXTcPuf3 parasites were analyzed by transmission electron microscopy (TEM), it was observed that the overexpressor had an abnormal mitochondrial morphology, evidenced by swelling. The results associated with cellular respiration showed that both the ΔTcPuf3 and pTEXTcPuf3 had a lower efficiency in routine respiration and the electron transport system capacity. Likewise, the mitochondria from overexpressing parasites showed a slight hyperpolarization. Additionally, several biological features, depending on the function of the mitochondria, were altered, such as growth, cell division, metacyclogenesis, ROS production, and response to benznidazole. In conclusion, our results suggest that although PUF3 is not an essential protein in T. cruzi, it influences mitochondrial transcripts, affecting mitochondrial morphology and function.

Loss of functional fragile X mental retardation protein (FMRP) causes fragile X syndrome and is the leading monogenic cause of autism spectrum disorders and intellectual disability. FMRP is most notably a translational repressor and is thought to inhibit translation elongation by stalling ribosomes as FMRP-bound polyribosomes from brain tissue are resistant to puromycin and nuclease treatment. Here, we present data showing that the C-terminal noncanonical RNA-binding domain of FMRP is essential and sufficient to induce puromycin-resistant mRNA•ribosome complexes. Given that stalled ribosomes can stimulate ribosome collisions and no-go mRNA decay (NGD), we tested the ability of FMRP to drive NGD of its target transcripts in neuroblastoma cells. Indeed, FMRP and ribosomal proteins, but not poly(A)-binding protein, were enriched in isolated nuclease-resistant disomes compared to controls. Using siRNA knockdown and RNA-seq, we identified 16 putative FMRP-mediated NGD substrates, many of which encode proteins involved in neuronal development and function. Increased mRNA stability of four putative substrates was also observed when either FMRP was depleted or NGD was prevented via RNAi. Taken together, these data support that FMRP stalls ribosomes but only stimulates NGD of a small select set of transcripts, revealing a minor role of FMRP that would be misregulated in fragile X syndrome.

BACKGROUND: Chronic Obstructive Pulmonary Disease (COPD) is characterized by high morbidity, disability, and mortality rates worldwide. RNA-binding proteins (RBPs) might regulate genes involved in oxidative stress and inflammation in COPD patients. Single-cell transcriptome sequencing (scRNA-seq) offers an accurate tool for identifying intercellular heterogeneity and the diversity of immune cells. However, the role of RBPs in the regulation of various cells, especially AT2 cells, remains elusive.
METHODS: A scRNA-seq dataset (GSE173896) and a bulk RNA-seq dataset acquired from airway tissues (GSE124180) were employed for data mining. Next, RNA-seq analysis was performed in both COPD and control patients. Differentially expressed genes (DEGs) were identified using criteria of fold change (FC ≥ 1.5 or ≤ 1.5) and P value ≤ 0.05. Lastly, Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and alternative splicing identification analyses were carried out.
RESULTS: RBP genes exhibited specific expression patterns across different cell groups and participated in cell proliferation and mitochondrial dysfunction in AT2 cells. As an RBP, AZGP1 expression was upregulated in both the scRNA-seq and RNA-seq datasets. It might potentially be a candidate immune biomarker that regulates COPD progression by modulating AT2 cell proliferation and adhesion by regulating the expression of SAMD5, DNER, DPYSL3, GBP5, GBP3, and KCNJ2. Moreover, AZGP1 regulated alternative splicing events in COPD, particularly DDAH1 and SFRP1, holding significant implications in COPD.
CONCLUSIONS: RBP gene AZGP1 inhibits epithelial cell proliferation by regulating genes participating in alternative splicing in COPD.

Transfer RNAs (tRNAs) are the most highly modified cellular RNAs, both with respect to the proportion of nucleotides that are modified within the tRNA sequence and with respect to the extraordinary diversity in tRNA modification chemistry. However, the functions of many different tRNA modifications are only beginning to emerge. tRNAs have two general clusters of modifications. The first cluster is within the anticodon stem-loop including several modifications essential for protein translation. The second cluster of modifications is within the tRNA elbow, and roles for these modifications are less clear. In general, tRNA elbow modifications are typically not essential for cell growth, but nonetheless several tRNA elbow modifications have been highly conserved throughout all domains of life. In addition to forming modifications, many tRNA modifying enzymes have been demonstrated or hypothesized to also play an important role in folding tRNA acting as tRNA chaperones. In this review, we summarize the known functions of tRNA modifying enzymes throughout the lifecycle of a tRNA molecule, from transcription to degradation. Thereby, we describe how tRNA modification and folding by tRNA modifying enzymes enhance tRNA maturation, tRNA aminoacylation, and tRNA function during protein synthesis, ultimately impacting cellular phenotypes and disease.

The alteration of gene expression is not restricted to transcriptional regulation but includes a variety of post-transcriptional mechanisms, however, the role of the latter underlying many diseases remains relatively unknown. By utilizing an RNA-Seq dataset of 1510 brain samples from individuals with autism spectrum disorder (ASD), bipolar disorder (BD), schizophrenia (SCZ), and controls, we assessed the contribution of post-transcriptional dysregulation and identified top perturbators accountable for transcriptomic changes of expression in neuropsychiatric disorders. Around 30% of the variability in expression can be attributed to post-transcriptional dysregulation. Interestingly, RNA stability tended to decrease in SCZ and BD, leading to the inhibition of neurogenesis and neural differentiation, while the increase in ASD, resulted in enhanced activity of apoptosis. This finding implicated contrasting pathologies involving RNA stability among neuropsychiatric disorders. An RNA binding protein (RBP)-ELAVL3 - is predicted to be significantly involved in the disruption of RNA stability in all three disorders. To validate, we knocked down its expression in cerebral organoids. Not only differentially expressed genes in ELAVL3-knockdown covered a considerable proportion of predicted targets in three disorders, we also found neurogenesis was significantly affected, given the diminished proliferation and consequently the reduced size of the organoids. Our study extends the current understanding of the link between post-transcriptional regulation and neuropsychiatric disorders and provides new therapeutic targets for early intervention.

Post-transcriptional regulation by RNA binding proteins can determine gene expression levels and drive changes in cancer cell proteomes. Identifying mechanisms of protein-RNA binding, including preferred sequence motifs bound in vivo, provides insights into protein-RNA networks and how they impact mRNA structure, function, and stability. In this review, we will focus on proteins that bind to AU-rich elements (AREs) in nascent or mature mRNA where they play roles in response to stresses encountered by cancer cells. ARE-binding proteins (ARE-BPs) specifically impact alternative splicing, stability, decay and translation, and formation of RNA-rich biomolecular condensates like cytoplasmic stress granules (SGs). For example, recent findings highlight the role of ARE-BPs - like TIAR and HUR - in chemotherapy resistance and in translational regulation of mRNAs encoding pro-inflammatory cytokines. We will discuss emerging evidence that different modes of ARE-BP activity impact leukaemia and lymphoma development, progression, adaptation to microenvironment and chemotherapy resistance.