Identification of genes related to ribosomal proteins in colorectal cancer: exploring their potential as biomarkers, prognostic indicators, and therapeutic targets.

BACKGROUND: Colorectal cancer (CRC) ranks as the third most commonly diagnosed cancer in both females and males, underscoring the need for the identification of effective biomarkers. METHODS AND RESULTS: We assessed the expression levels of ribosomal proteins (RPs) at both mRNA and protein levels. Subsequently, leveraging the STRING database, we constructed a protein-protein interaction network and identified hub genes. The co-expression network of differentially expressed genes associated with CRC and their target hub RPs was constructed using the weighted gene co-expression network analysis algorithm. Gene ontology and molecular signatures database were conducted to gain insights into the biological roles of genes associated with the identified module. To confirm the results, the expression level of the candidate genes in the CRC samples compared to the adjacent healthy was evaluated by the RT-qPCR method. Our findings indicated that the genes related to RPs were predominantly enriched in biological processes associated with Myc Targets, Oxidative Phosphorylation, and cell proliferation. Also, results demonstrated that elevated levels of GRWD1, MCM5, IMP4, and RABEPK that related to RPs were associated with poor prognostic outcomes for CRC patients. Notably, IMP4 and RABEPK exhibited higher diagnostic value. Moreover, the expression of IMP4 and RABEPK showed a significant association with drug resistance using cancer cell line encyclopedia and genomics of drug sensitivity in cancer databases. Also, the results showed that the expression level of IMP4 and RABEPK in cancerous samples was significantly higher compared to the adjacent healthy ones. CONCLUSION: The general results of this study have shown that many genes related to RPs are increased in cancer and could be associated with the death rate of patients. We also highlighted the therapeutic and prognostic potentials of RPs genes in CRC.

Knockdown of ribosome RNA processing protein 15 suppresses migration of hepatocellular carcinoma through inhibiting PATZ1-associated LAMC2/FAK pathway.

BACKGROUND: Ribosomal RNA processing protein 15 (RRP15) has been found to regulate the progression of hepatocellular carcinoma (HCC). Nevertheless, the extent to which it contributes to the spread of HCC cells remains uncertain. Thus, the objective of this research was to assess the biological function of RRP15 in the migration of HCC. METHODS: The expression of RRP15 in HCC tissue microarray (TMA), tumor tissues and cell lines were determined. In vitro, the effects of RRP15 knockdown on the migration, invasion and adhesion ability of HCC cells were assessed by wound healing assay, transwell and adhesion assay, respectively. The effect of RRP15 knockdown on HCC migration was also evaluated in vivo in a mouse model. RESULTS: Bioinformatics analysis showed that high expression of RRP15 was significantly associated with low survival rate of HCC. The expression level of RRP15 was strikingly upregulated in HCC tissues and cell lines compared with the corresponding controls, and TMA data also indicated that RRP15 was a pivotal prognostic factor for HCC. RRP15 knockdown in HCC cells reduced epithelial-to-mesenchymal transition (EMT) and inhibited migration in vitro and in vivo, independent of P53 expression. Mechanistically, blockade of RRP15 reduced the protein level of the transcription factor POZ/BTB and AT hook containing zinc finger 1 (PATZ1), resulting in decreased expression of the downstream genes encoding laminin 5 subunits, LAMC2 and LAMB3, eventually suppressing the integrin ß4 (ITGB4)/focal adhesion kinase (FAK)/nuclear factor κB kappa-light-chain-enhancer of activated B cells (NF-κB) signaling pathway. CONCLUSIONS: RRP15 promotes HCC migration by activating the LAMC2/ITGB4/FAK pathway, providing a new target for future HCC treatment.

RPL35A drives ovarian cancer progression by promoting the binding of YY1 to CTCF promoter.

Ovarian cancer is one of the most common gynaecological malignancies with poor prognosis and lack of effective treatment. The improvement of the situation of ovarian cancer urgently requires the exploration of its molecular mechanism to develop more effective molecular targeted drugs. In this study, the role of human ribosomal protein l35a (RPL35A) in ovarian cancer was explored in vitro and in vivo. Our data identified that RPL35A expression was abnormally elevated in ovarian cancer. Clinically, high expression of RPL35A predicted short survival and poor TNM staging in patients with ovarian cancer. Functionally, RPL35A knock down inhibited ovarian cancer cell proliferation and migration, enhanced apoptosis, while overexpression had the opposite effect. Mechanically, RPL35A promoted the direct binding of transcription factor YY1 to CTCF in ovarian cancer cells. Consistently, RPL35A regulated ovarian cancer progression depending on CTCF in vitro and in vivo. Furthermore, RPL35A affected the proliferation and apoptosis of ovarian cancer cells through PPAR signalling pathway. In conclusion, RPL35A drove ovarian cancer progression by promoting the binding of YY1 and CTCF promoter, and inhibiting this process may be an effective strategy for targeted therapy of this disease.

Ribosomal collision is not a prerequisite for ZNF598-mediated ribosome ubiquitination and disassembly of ribosomal complexes by ASCC.

Ribosomal stalling induces the ribosome-associated quality control (RQC) pathway targeting aberrant polypeptides. RQC is initiated by K63-polyubiquitination of ribosomal protein uS10 located at the mRNA entrance of stalled ribosomes by the E3 ubiquitin ligase ZNF598 (Hel2 in yeast). Ubiquitinated ribosomes are dissociated by the ASC-1 complex (ASCC) (RQC-Trigger (RQT) complex in yeast). A cryo-EM structure of the ribosome-bound RQT complex suggested the dissociation mechanism, in which the RNA helicase Slh1 subunit of RQT (ASCC3 in mammals) applies a pulling force on the mRNA, inducing destabilizing conformational changes in the 40S subunit, whereas the collided ribosome acts as a wedge, promoting subunit dissociation. Here, using an in vitro reconstitution approach, we found that ribosomal collision is not a strict prerequisite for ribosomal ubiquitination by ZNF598 or for ASCC-mediated ribosome release. Following ubiquitination by ZNF598, ASCC efficiently dissociated all polysomal ribosomes in a stalled queue, monosomes assembled in RRL, in vitro reconstituted 80S elongation complexes in pre- and post-translocated states, and 48S initiation complexes, as long as such complexes contained ≥ 30-35 3'-terminal mRNA nt. downstream from the P site and sufficiently long ubiquitin chains. Dissociation of polysomes and monosomes both involved ribosomal splitting, enabling Listerin-mediated ubiquitination of 60S-associated nascent chains.

The small nucleolar RNA SNORA51 enhances breast cancer stem cell-like properties via the RPL3/NPM1/c-MYC pathway.

Breast cancer stem cells (BCSCs) are key players in carcinogenesis and development. Small nucleolar RNAs (snoRNAs) seem to have a crucial influence on regulating stem cell-like properties in various cancers, but the underlying mechanism in breast cancer has not been determined. In this study, we first found that the expression of SNORA51 might be strongly and positively related to BCSCs-like properties. SNORA51 expression was assessed in breast cancer tissues (n = 158 patients) by in situ hybridization. Colony formation, cell counting kit-8, and sphere formation assays were used to detect cell proliferation and self-renewal, respectively. Wound healing and transwell assays were used to detect cell migration. Coimmunoprecipitation and molecular docking were used to determine the underlying mechanism through which SNORA51 regulates BCSCs-like properties. High SNORA51 expression was associated with a worse prognosis, overall survival, and disease-free survival, in 158 breast cancer patients and was also closely related to lymph node status, ER status, the Ki-67 index, histological grade, and TNM stage. Further analysis proved that SNORA51 could enhance and maintain stem cell-like properties, including cell proliferation, self-renewal, and migration, in breast cancer. Moreover, high SNORA51 expression could reduce nucleolar RPL3 expression, induce changes in the expression of NPM1 in the nucleolus and nucleoplasm, and ultimately increase c-MYC expression. Taken together, our findings demonstrated that SNORA51 could enhance BCSCs-like properties via the RPL3/NPM1/c-MYC pathway both in vitro and in vivo. Therefore, SNORA51 might be a significant biomarker and potential therapeutic target and might even provide a new viewpoint on the regulatory mechanism of snoRNAs in breast cancer or other malignant tumors.

RPL35A promotes the progression of cholangiocarcinoma by mediating HSPA8 ubiquitination.

BACKGROUND: Cholangiocarcinoma (CCA) is a biliary epithelial malignant tumor with an increasing incidence worldwide. Therefore, further understanding of the molecular mechanisms of CCA progression is required to identify new therapeutic targets. METHODS: The expression of RPL35A in CCA and para-carcinoma tissues was detected by immunohistochemical staining. IP-MS combined with Co-IP identified downstream proteins regulated by RPL35A. Western blot and Co-IP of CHX or MG-132 treated CCA cells were used to verify the regulation of HSPA8 protein by RPL35A. Cell experiments and subcutaneous tumorigenesis experiments in nude mice were performed to evaluate the effects of RPL35A and HSPA8 on the proliferation, apoptosis, cell cycle, migration of CCA cells and tumor growth in vivo. RESULTS: RPL35A was significantly upregulated in CCA tissues and cells. RPL35A knockdown inhibited the proliferation and migration of HCCC-9810 and HUCCT1 cells, induced apoptosis, and arrested the cell cycle in G1 phase. HSPA8 was a downstream protein of RPL35A and overexpressed in CCA. RPL35A knockdown impaired HSPA8 protein stability and increased HSPA8 protein ubiquitination levels. RPL35A overexpression promoted CCA cell proliferation and migration. HSPA8 knockdown inhibited CCA cell proliferation and migration, and reversed the promoting effect of RPL35A. Furthermore, RPL35A promoted tumor growth in vivo. In contrast, HSPA8 knockdown suppressed tumor growth, while was able to restore the effects of RPL35A overexpression. CONCLUSION: RPL35A was upregulated in CCA tissues and promoted the progression of CCA by mediating HSPA8 ubiquitination.

Androgen deprivation induces double-null prostate cancer via aberrant nuclear export and ribosomal biogenesis through HGF and Wnt activation.

Androgen deprivation therapy (ADT) targeting androgen/androgen receptor (AR)- signaling pathways is the main therapy for advanced prostate cancer (PCa). However, ADT eventually fails in most patients who consequently develop castration-resistant prostate cancer (CRPC). While more potent AR antagonists and blockers for androgen synthesis were developed to improve clinical outcomes, they also show to induce more diverse CRPC phenotypes. Specifically, the AR- and neuroendocrine-null PCa, DNPC, occurs in abiraterone and enzalutamide-treated patients. Here, we uncover that current ADT induces aberrant HGF/MET signaling activation that further elevates Wnt/ß-catenin signaling in human DNPC samples. Co-activation of HGF/MET and Wnt/ß-catenin axes in mouse prostates induces DNPC-like lesions. Single-cell RNA sequencing analyses identify increased expression and activity of XPO1 and ribosomal proteins in mouse DNPC-like cells. Elevated expression of XPO1 and ribosomal proteins is also identified in clinical DNPC specimens. Inhibition of XPO1 and ribosomal pathways represses DNPC growth in both in vivo and ex vivo conditions, evidencing future therapeutic targets.

UFM1 E3 ligase promotes recycling of 60S ribosomal subunits from the ER.

Reversible modification of target proteins by ubiquitin and ubiquitin-like proteins (UBLs) is widely used by eukaryotic cells to control protein fate and cell behaviour1. UFM1 is a UBL that predominantly modifies a single lysine residue on a single ribosomal protein, uL24 (also called RPL26), on ribosomes at the cytoplasmic surface of the endoplasmic reticulum (ER)2,3. UFM1 conjugation (UFMylation) facilitates the rescue of 60S ribosomal subunits (60S) that are released after ribosome-associated quality-control-mediated splitting of ribosomes that stall during co-translational translocation of secretory proteins into the ER3,4. Neither the molecular mechanism by which the UFMylation machinery achieves such precise target selection nor how this ribosomal modification promotes 60S rescue is known. Here we show that ribosome UFMylation in vivo occurs on free 60S and we present sequential cryo-electron microscopy snapshots of the heterotrimeric UFM1 E3 ligase (E3(UFM1)) engaging its substrate uL24. E3(UFM1) binds the L1 stalk, empty transfer RNA-binding sites and the peptidyl transferase centre through carboxy-terminal domains of UFL1, which results in uL24 modification more than 150 Å away. After catalysing UFM1 transfer, E3(UFM1) remains stably bound to its product, UFMylated 60S, forming a C-shaped clamp that extends all the way around the 60S from the transfer RNA-binding sites to the polypeptide tunnel exit. Our structural and biochemical analyses suggest a role for E3(UFM1) in post-termination release and recycling of the large ribosomal subunit from the ER membrane.

The UFM1 E3 ligase recognizes and releases 60S ribosomes from ER translocons.

Stalled ribosomes at the endoplasmic reticulum (ER) are covalently modified with the ubiquitin-like protein UFM1 on the 60S ribosomal subunit protein RPL26 (also known as uL24)1,2. This modification, which is known as UFMylation, is orchestrated by the UFM1 ribosome E3 ligase (UREL) complex, comprising UFL1, UFBP1 and CDK5RAP3 (ref. 3). However, the catalytic mechanism of UREL and the functional consequences of UFMylation are unclear. Here we present cryo-electron microscopy structures of UREL bound to 60S ribosomes, revealing the basis of its substrate specificity. UREL wraps around the 60S subunit to form a C-shaped clamp architecture that blocks the tRNA-binding sites at one end, and the peptide exit tunnel at the other. A UFL1 loop inserts into and remodels the peptidyl transferase centre. These features of UREL suggest a crucial function for UFMylation in the release and recycling of stalled or terminated ribosomes from the ER membrane. In the absence of functional UREL, 60S-SEC61 translocon complexes accumulate at the ER membrane, demonstrating that UFMylation is necessary for releasing SEC61 from 60S subunits. Notably, this release is facilitated by a functional switch of UREL from a 'writer' to a 'reader' module that recognizes its product-UFMylated 60S ribosomes. Collectively, we identify a fundamental role for UREL in dissociating 60S subunits from the SEC61 translocon and the basis for UFMylation in regulating protein homeostasis at the ER.

RNA-binding protein RPS7 promotes hepatocellular carcinoma progression via LOXL2-dependent activation of ITGB1/FAK/SRC signaling.

BACKGROUND: Metastasis is one of the leading cause contributes to treatment failure and poor prognosis of hepatocellular carcinoma (HCC) patients. The underlying mechanism of HCC metastasis remains to be determined. Although several RNA binding proteins (RBPs) have been found to participate in tumorigenesis and progression of liver cancer, the role of RBPs in HCC patients with extrahepatic metastases is poorly understood. METHODS: By performing RNA-seq of primary HCC tissues (including HCC with extrahepatic metastasis and those did not develop metastasis), we identified a set of HCC metastasis-associated RBPs candidates. Among which, ribosomal protein S7 (RPS7) was found to be remarkably increased in HCC tissues and be strongly related to HCC poor survival. Overexpression or CRISPR-Cas9-mediated knockout were applied to investigate the role of RPS7 on the metastasis-associated phenotypes of HCC cells. RNA sequencing, RIP, RNA-pull down, dual luciferase reporter assay, nascent RNA capture assay, and RNA decay and so on, were applied to reveal the underlying mechanism of RPS7 induced HCC metastasis. RESULTS: Gain- and loss- of function analyses revealed that RPS7 promoted HCC cells adhesion, migration and invasion capabilities, as well as lung metastasis. Mechanistically, we uncovered that lysyl oxidase-like 2 (LOXL2) was a critical downstream target of RPS7. RPS7 could stabilize LOXL2 mRNA by binding to AUUUA motifs in the 3155-3375 region of the 3'UTR of LOXL2 mRNA, thus increased LOXL2 expression via elevating LOXL2 mRNA abundance. Further research revealed that LOXL2 could accelerate focal adhesion formation through maintaining the protein stability of ITGB1 and activating ITGB1-mediated FAK/SRC signaling pathway, and thereby contribute to the pro-metastasis effect of RPS7. CONCLUSIONS: Taken together, our data reveal a novel function of RPS7 in HCC metastasis, also reveal the critical roles of the RPS7/LOXL2/ITGB1 axis in HCC metastasis and shed new light on the exploration of molecular drugs against HCC.

Deficiency of the ribosomal protein uS10 (RPS20) reorganizes human cells translatome according to the abundance, CDS length and GC content of mRNAs.

Ribosomal protein uS10, a product of the RPS20 gene, is an essential constituent of the small (40S) subunit of the human ribosome. Disruptive mutations in its gene are associated with a predisposition to hereditary colorectal carcinoma. Here, using HEK293T cells, we show that a deficiency of this protein leads to a decrease in the level of ribosomes (ribosomal shortage). RNA sequencing of the total and polysome-associated mRNA samples reveals hundreds of genes differentially expressed in the transcriptome (t)DEGs and translatome (p)DEGs under conditions of uS10 deficiency. We demonstrate that the (t)DEG and (p)DEG sets partially overlap, determine genes with altered translational efficiency (TE) and identify cellular processes affected by uS10 deficiency-induced ribosomal shortage. We reveal that translated mRNAs of upregulated (p)DEGs and genes with altered TE in uS10-deficient cells are generally more abundant and that their GC contents are significantly lower than those of the respective downregulated sets. We also observed that upregulated (p)DEGs have longer coding sequences. Based on our findings, we propose a combinatorial model describing the process of reorganization of mRNA translation under conditions of ribosomal shortage. Our results reveal rules according to which ribosomal shortage reorganizes the transcriptome and translatome repertoires of actively proliferating cells.

Suppressor analysis links trans-translation and ribosomal protein uS7 to RluD function in Escherichia coli.

The pseudouridine (ψ) synthase, RluD is responsible for three ψ modifications in the helix 69 (H69) of bacterial 23S rRNA. While normally dispensable, rluD becomes critical for rapid cell growth in bacteria that are defective in translation-termination. In slow-growing rluD- bacteria, suppressors affecting termination factors RF2 and RF3 arise frequently and restore normal termination and rapid cell growth. Here we describe two weaker suppressors, affecting rpsG, encoding ribosomal protein uS7 and ssrA, encoding tmRNA. In K-12 strains of E. coli, rpsG terminates at a TGA codon. In the suppressor strain, alteration of an upstream CAG to a TAG stop codon results in a shortened uS7 and partial alleviation of slow growth, likely by replacing an inefficient TGA stop codon with the more efficient TAG. Inefficient termination events, such as occurs in some rluD- strains, are targeted by trans-translation. Inactivation of the ssrA gene in slow-growing, termination-defective mutants lacking RluD and RF3, also partially restores robust growth, most probably by preventing destruction of completed polypeptides on ribosomes at slow-terminating stop codons. Finally, an additional role for RluD has been proposed, independent of its pseudouridine synthase activity. This is based on the observation that plasmids expressing catalytically dead (D139N or D139T) RluD proteins could nonetheless restore robust growth to an E. coli K-12 rluD- mutant. However, newly constructed D139N and D139T rluD plasmids do not have any growth-restoring activity and the original observations were likely due to the appearance of suppressors.

DNA methylation-mediated epigenetic regulation of oncogenic RPS2 as a novel therapeutic target and biomarker in hepatocellular carcinoma.

Ribosomal Protein S2 (RPS2) has emerged as a potential prognostic biomarker due to its involvement in key cellular processes and its altered expression pattern in certain types of cancer. However, its role in hepatocellular carcinoma (HCC) has yet to be investigated. Herein, we analyzed RPS2 mRNA expression and promoter methylation in HCC patient samples and HepG2 cells. Subsequently, loss-of-function experiments were conducted to determine the function of RPS2 in HCC cells in vitro. Our results revealed that RPS2 mRNA expression is significantly elevated, and its promoter is hypomethylated in HCC patient samples compared to controls. In addition, 5-Azacytidine treatment in HepG2 cells decreased RPS2 promoter methylation level and increased its mRNA expression. RPS2 knockdown in HepG2 cells suppressed cell proliferation and promoted apoptosis. Functional pathway analysis of genes positively and negatively associated with RPS2 expression in HCC showed enrichment in ribosomal biogenesis, translation machinery, cell cycle regulation, and DNA processing. Furthermore, utilizing drug-protein 3D docking, we found that doxorubicin, sorafenib, and 5-Fluorouracil, showed high affinity to the active sites of RPS2, and in vitro treatment with these drugs reduced RPS2 expression. For the first time, we report on DNA methylation-mediated epigenetic regulation of RPS2 and its oncogenic role in HCC. Our findings suggest that RPS2 plays a significant role in the development and progression of HCC, hence its potential prognostic and therapeutic utility. Moreover, as epigenetic changes happen early in cancer development, RPS2 may serve as a potential biomarker for tumor progression.

Autophagy-dependent alternative splicing of ribosomal protein S24 produces a more stable isoform that aids in hypoxic cell survival.

Cells remodel splicing and translation machineries to mount specialized gene expression responses to stress. Here, we show that hypoxic human cells in 2D and 3D culture models increase the relative abundance of a longer mRNA variant of ribosomal protein S24 (RPS24L) compared to a shorter mRNA variant (RPS24S) by favoring the inclusion of a 22 bp cassette exon. Mechanistically, RPS24L and RPS24S are induced and repressed, respectively, by distinct pathways in hypoxia: RPS24L is induced in an autophagy-dependent manner, while RPS24S is reduced by mTORC1 repression in a hypoxia-inducible factor-dependent manner. RPS24L produces a more stable protein isoform that aids in hypoxic cell survival and growth, which could be exploited by cancer cells in the tumor microenvironment.

Potential prognostic biomarkers of hepatocellular carcinoma based on 4D label-free quantitative proteomics analysis pilot investigation.

BACKGROUND: Hepatocellular carcinoma carries a poor prognosis and poses a serious threat to global health. Currently, there are few potential prognostic biomarkers available for the prognosis of hepatocellular carcinoma. METHODS: This pilot study used 4D label-free quantitative proteomics to compare the proteomes of hepatocellular carcinoma and adjacent non-tumor tissue. A total of 66,075 peptides, 6363 identified proteins, and 772 differentially expressed proteins were identified in specimens from three hepatocellular carcinoma patients. Through functional enrichment analysis of differentially expressed proteins by Gene Ontology, KEGG pathway, and protein domain, we identified proteins with similar functions. RESULTS: Twelve differentially expressed proteins (RPL17, RPL27, RPL27A, RPS5, RPS16, RSL1D1, DDX18, RRP12, TARS2, YARS2, MARS2, and NARS1) were selected for identification and validation by parallel reaction monitoring. Subsequent Western blotting confirmed overexpression of RPL27, RPS16, and TARS2 in hepatocellular carcinoma compared to non-tumor tissue in 16 pairs of clinical samples. Analysis of The Cancer Genome Atlas datasets associated the increased expression of these proteins with poor prognosis. Tissue microarray revealed a negative association between high expression of RPL27 and TARS2 and the prognosis of hepatocellular carcinoma patients, although RPS16 was not significant. CONCLUSIONS: These data suggest that RPL27 and TARS2 play an important role in hepatocellular carcinoma progression and may be potential prognostic biomarkers of overall survival in hepatocellular carcinoma patients.

Perspectives of current understanding and therapeutics of Diamond-Blackfan anemia.

ABSTACT: Diamond-Blackfan anemia (DBA) is a rare congenital bone marrow failure disorder characterized by erythroid hypoplasia. It primarily affects infants and is often caused by heterozygous allelic variations in ribosomal protein (RP) genes. Recent studies also indicated that non-RP genes like GATA1, TSR2, are associated with DBA. P53 activation, translational dysfunction, inflammation, imbalanced globin/heme synthesis, and autophagy dysregulation were shown to contribute to disrupted erythropoiesis and impaired red blood cell production. The main therapeutic option for DBA patients is corticosteroids. However, half of these patients become non-responsive to corticosteroid therapy over prolonged treatment and have to be given blood transfusions. Hematopoietic stem cell transplantation is currently the sole curative option, however, the treatment is limited by the availability of suitable donors and the potential for serious immunological complications. Recent advances in gene therapy using lentiviral vectors have shown promise in treating RPS19-deficient DBA by promoting normal hematopoiesis. With deepening insights into the molecular framework of DBA, emerging therapies like gene therapy hold promise for providing curative solutions and advancing comprehension of the underlying disease mechanisms.

Ribosomal protein SA is a common component of neuronal intranuclear inclusions in polyglutamine diseases and Marinesco bodies.

Neuronal intranuclear inclusions (NIIs) are common key structures in polyglutamine (polyQ) diseases such as Huntington disease (HD), spinocerebellar ataxia type 1 (SCA1), and SCA3. Marinesco bodies (MBs) of dopaminergic neurons in the substantia nigra are also intranuclear structures and are frequently seen in normal elderly people. Ribosomal dysfunction is closely related to two differential processes; therefore, we aimed to identify the pathological characteristics of ribosomal protein SA (RPSA), a ribosomal protein, in both states. To this end, we evaluated the autopsy findings in four patients with HD, two SCA3, and five normal elderly cases (NCs). Immunohistochemical studies demonstrated that both NIIs and MBs contain RPSA. In polyQ diseases, RPSA was co-localized with polyQ aggregations, and 3D-reconstructed images revealed their mosaic-like distribution. Assessments of the organization of RPSA and p62 in NIIs showed that RPSA was more localized toward the center than p62 and that this unique organization was more evident in the MBs. Immunoblotting of the temporal cortices revealed that the nuclear fraction of HD patients contained more RPSA than that of NCs. In conclusion, our study revealed that RPSA is a common component of both NIIs and MBs, indicating that a similar mechanism contributes to the formation of polyQ NIIs and MBs.

The m6A Reader YTHDC2 Suppresses Lung Adenocarcinoma Tumorigenesis by Destabilizing MRPL12.

N6-methyladenosine (m6A) is the most common posttranscriptional RNA modification and plays significant roles in physiological and pathological progression. Here, we probed the functions and mechanism of the m6A reader YTH domain containing 2 (YTHDC2) in Lung Adenocarcinoma (LUAD) tumorigenesis. Levels of genes and proteins of YTHDC2 and Mitochondrial ribosomal protein L7/L12 (MRPL12) were assayed by quantitative real-time polymerase chain reaction, western blotting and Immunohistochemistry (IHC) analyses. In vitro analysis was conducted using 5-ethynyl-2'-deoxyuridine (EdU), colony formation, flow cytometry, and transwell assays, respectively. In vivo assay was performed by using the mouse lung adenocarcinoma model. The methylated RNA immunoprecipitation (MeRIP) assay was used to detect the m6A modification profile of MRPL12 mRNA. YTHDC2 was lowly expressed in lung adenocarcinoma tissues and cells. Overexpression of YTHDC2 suppressed the proliferation, invasion and migration of lung adenocarcinoma cells, but induced cell apoptosis. As expected, forced expression of YTHDC2 hindered lung adenocarcinoma tumor growth in vivo. Mechanistically, YTHDC2 preferentially bound to m6A-modified MRPL12 mRNA and destabilized its expression. MRPL12 was highly expressed in lung adenocarcinoma tissues and cells, and MRPL12 silencing repressed the growth and mobility of lung adenocarcinoma cells. Moreover, MRPL12 upregulation attenuated the anticancer activity of YTHDC2 in lung adenocarcinoma cells. In vivo assay also showed YTHDC2 suppressed tumor growth in the lung adenocarcinoma mouse model via downregulating MRPL12. The m6A reader YTHDC2 repressed lung adenocarcinoma tumorigenesis by destabilizing MRPL12 in an m6A-dependent manner.

Characterization of nucleolar SUMO isopeptidases unveils a general p53-independent checkpoint of impaired ribosome biogenesis.

Ribosome biogenesis is a multi-step process, in which a network of trans-acting factors ensures the coordinated assembly of pre-ribosomal particles in order to generate functional ribosomes. Ribosome biogenesis is tightly coordinated with cell proliferation and its perturbation activates a p53-dependent cell-cycle checkpoint. How p53-independent signalling networks connect impaired ribosome biogenesis to the cell-cycle machinery has remained largely enigmatic. We demonstrate that inactivation of the nucleolar SUMO isopeptidases SENP3 and SENP5 disturbs distinct steps of 40S and 60S ribosomal subunit assembly pathways, thereby triggering the canonical p53-dependent impaired ribosome biogenesis checkpoint. However, inactivation of SENP3 or SENP5 also induces a p53-independent checkpoint that converges on the specific downregulation of the key cell-cycle regulator CDK6. We further reveal that impaired ribosome biogenesis generally triggers the downregulation of CDK6, independent of the cellular p53 status. Altogether, these data define the role of SUMO signalling in ribosome biogenesis and unveil a p53-independent checkpoint of impaired ribosome biogenesis.

RNA-binding motif protein 10 inactivates c-Myc by partnering with ribosomal proteins uL18 and uL5.

RNA-binding motif protein 10 (RBM10) is a frequently mutated tumor suppressor in lung adenocarcinoma (LUAD). Yet, it remains unknown whether cancer-derived mutant RBM10 compromises its tumor suppression function and, if so, the molecular insight of the underlying mechanisms. Here, we show that wild-type RBM10 suppresses lung cancer cell growth and proliferation by inactivating c-Myc that is essential for cancer cell survival. RBM10 directly binds to c-Myc and promotes c-Myc's ubiquitin-dependent degradation, while RBM10 knockdown leads to the induction of c-Myc level and activity. This negative action on c-Myc is further boosted by ribosomal proteins (RPs) uL18 (RPL5) and uL5 (RPL11) via their direct binding to RBM10. Cancer-derived mutant RBM10-I316F fails to bind to uL18 and uL5 and to inactivate c-Myc, thus incapable of suppressing tumorigenesis. Our findings uncover RBM10 as a pivotal c-Myc repressor by cooperating with uL18 and uL5 in lung cancer cells, as its failure to do so upon mutation favors tumorigenesis.