Like protein and DNA, different types of RNA molecules undergo various modifications. Accumulating evidence suggests that these RNA modifications serve as sophisticated codes to mediate RNA behaviors and many important biological functions. N-methyladenosine (mA) is the most abundant internal RNA modification found in a variety of eukaryotic RNAs, including but not limited to mRNAs, tRNAs, rRNAs, and long non-coding RNAs (lncRNAs). In mammalian cells, mA can be incorporated by a methyltransferase complex and removed by demethylases, which ensures that the mA modification is reversible and dynamic. Moreover, mA is recognized by the YT521-B homology (YTH) domain-containing proteins, which subsequently direct different complexes to regulate RNA signaling pathways, such as RNA metabolism, RNA splicing, RNA folding, and protein translation. Herein, we summarize the recent progresses made in understanding the molecular mechanisms underlying the mA recognition by YTH domain-containing proteins, which would shed new light on mA-specific recognition and provide clues to the future identification of reader proteins of many other RNA modifications.
Adenosine ; analogs & derivatives ; chemistry ; metabolism ; Animals ; Humans ; Protein Binding ; Protein Domains ; RNA ; chemistry ; metabolism ; RNA-Binding Proteins ; chemistry ; metabolism

The genomic instability may lead to an initiation of cancer in many organisms. Homologous recombination repair (HRR) is vital in maintaining cellular genomic stability. RAD51 associated protein 1 (RAD51AP1), which plays a crucial role in HRR and primarily participates in forming D-loop, was reported as an essential protein for maintaining cellular genomic stability. However, recent studies showed that RAD51AP1 was significantly overexpressed in various cancer types and correlated with poor prognosis. These results suggested that RAD51AP1 may play a significant pro-cancer effect in multiple cancers. The underlying mechanism is still unclear. Cancer stemness-maintaining effects of RAD51AP1 might be considered as the most reliable mechanism. Meanwhile, RAD51AP1 also promoted resistance to radiation therapy and chemotherapy in many cancers. Thus, researches focused on RAD51AP1, and its regulatory molecules may provide new targets for overcoming cancer progression and treatment resistance. Here, we reviewed the latest research on RAD51AP1 in cancers and summarized its differential expression and prognostic implications. In this review, we also outlined the potential mechanisms of its pro-cancer and drug resistance-promoting effects to provide several potential directions for further research.
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Humans ; DNA-Binding Proteins/metabolism* ; RNA-Binding Proteins/metabolism* ; Lung Neoplasms ; DNA Repair ; Genomic Instability ; Rad51 Recombinase/metabolism*

OBJECTIVETo study the mechanism of the cellular proteins involved in the process of replication of hepatitis C virus (HCV) negative-strand RNA.

METHODSUltraviolet (UV) cross-linking was used to identify the cellular proteins that would bind to the 3'-end of HCV negative-strand RNA. Competition experiment was used to confirm the specificity of this binding, in which excess nonhomologous protein and RNA transcripts were used as competitors. The required binding sequence was determined by mapping, then the binding site was predicted through secondary structure analysis.

RESULTSA cellular protein of 45 kD (p45) was found to bind specifically to the 3'-end of HCV negative-strand RNA by UV cross-linking. Nonhomologous proteins and RNA transcripts could not compete out this binding, whereas the unlabeled 3'-end of HCV negative-strand RNA could. Mapping of the protein-binding site suggested that the 3'-end 131-278nt of HCV negative-strand RNA was the possible protein-binding region. Analysis of RNA secondary structure presumed that the potential binding site was located at 194-GAAAGAAC-201.

CONCLUSIONThe cellular protein p45 could specifically bind to the secondary structure of the 3'-end of HCV intermediate negative-strand RNA, and may play an important role in HCV RNA replication.

Binding Sites ; Hepacivirus ; genetics ; Nucleic Acid Conformation ; RNA, Viral ; chemistry ; metabolism ; RNA-Binding Proteins ; analysis ; metabolism ; Virus Replication

RNA binding protein (RBP) plays a key role in gene regulation and participate in RNA translation, modification, splicing, transport and other important biological processes. Studies have shown that abnormal expression of RBP is associated with a variety of diseases. The Musashi (Msi) family of mammals is an evolutionarily conserved and powerful RBP, whose members Msi1 and Msi2 play important roles in the regulation of stem cell activity and tumor development. The Msi family members regulate a variety of biological processes by binding and regulating mRNA translation, stability and downstream cell signaling pathways, and among them, Msi2 is closely related to embryonic growth and development, maintenance of tumor stem cells and development of hematological tumors. Accumulating evidence has shown that Msi2 also plays a crucial role in the development of solid tumors, mainly by affecting the proliferation, invasion, metastasis and drug resistance of tumors, involving Wnt/β-catenin, TGF-β/SMAD3, Akt/mTOR, JAK/STAT, Numb and their related signaling pathways (Notch, p53, and Hedgehog pathway). Preclinical studies of Msi2 gene as a therapeutic target for tumor have achieved preliminary results. This review summarizes the molecular structure, physiological function, role of Msi2 in the development and progression of various solid tumors and the signaling pathways involved.
Animals ; Hedgehog Proteins ; Mammals/metabolism* ; Neoplasms/genetics* ; Neoplastic Stem Cells ; RNA-Binding Proteins/metabolism* ; Signal Transduction

OBJECTIVETo investigate the role of Sam68 (Src-associated substrate during mitosis 68 kD) in the occurrence and development of colorectal cancer.

METHODSColorectal cancer cell lines with stable Sam68 over-expressing and low Sam68 expression were established to test the effect of Sam68 in the proliferation, invasion, and migration of the cancer cells using colony formation, MTT and Transwell assays.

RESULTSSW480 and Ls174t colorectal cell lines over-expressing Sam68 showed significantly enhanced cell proliferation, invasion and migration (P<0.05). Conversely, the low Sam68 expression in SW620 and HCT116 colorectal cell lines significantly suppressed the cell proliferation, invasion and migration (P<0.05).

CONCLUSIONThe expression of Sam68 can promote the proliferation, invasion and migration of colorectal cancer cells lines in vitro.

Adaptor Proteins, Signal Transducing ; metabolism ; Cell Cycle Proteins ; Cell Line, Tumor ; Cell Movement ; Cell Proliferation ; Colorectal Neoplasms ; metabolism ; pathology ; DNA-Binding Proteins ; metabolism ; Humans ; RNA-Binding Proteins ; metabolism

Influenza C virus is an important respiratory pathogen not only infecting people, but also pigs, dogs, and other animals. Polymerase is central to the replication of influenza C virus and is an important target for studying the mechanism of viral replication. However, there is no commercial monoclonal antibody (MAb) targeting influenza C virus polymerase, which hampers the development of relevant research to some extent. In order to prepare MAb targeting the polymerase basic protein 2 (PB2) of influenza C virus, influenza C virus RNA-dependent RNA polymerase (RdRp, consists of PB1, PB2 and P3) was co-immunoprecipitated with Flag-tagged human acidic nuclear phosphoprotein 32A (huANP32A-Flag) from 293T cells based on the interaction between huANP32A and influenza virus RdRp. The purified RdRp was used as antigen to immunize BALB/c mice. Six positive hybridoma cell lines (7B11-5, 8A4-5, 13D9-6, 8D4-1, 8D4-3, 9F9-4) that stably secrete and recognize PB2 MAb were screened by indirect ELISA and Western blotting. The subtypes of MAb 7B11-5, 8A4-5, 8D4-1 and 8D4-3 antibody were identified as IgG1, the subtypes of MAb 13D9-6 and 9F9-4 were IgG2a and IgG3, respectively. All the light chains of the MAbs were κ chain. A hybridoma cell line 8D4-1 with high titer was further selected to prepare ascites. The titer of mouse ascites antibody was determined to be 1:64 000. Western blotting results showed that the MAb 8D4-1 had a specific immune response with ICV PB2; laser confocal assay showed that the prepared MAb 8D4-1 accurately detected the subcellular localization of PB2 subunits. Moreover, ICV RdRp was highly enriched by ANP32A. The high specific of the prepared PB2 MAb 8D4-1 may facilitate the polymerase detection, structural analysis and mechanism study of influenza C virus.
Animals ; Antibodies, Monoclonal/metabolism* ; Ascites ; Humans ; Influenzavirus C/metabolism* ; Mice ; Nuclear Proteins/metabolism* ; RNA-Binding Proteins ; RNA-Dependent RNA Polymerase/genetics* ; Viral Proteins/metabolism* ; Virus Replication

The zinc-finger antiviral protein (ZAP) is a host factor that specifically inhibits the replication of certain viruses by eliminating viral mRNAs in the cytoplasm. In previous studies, we demonstrated that ZAP directly binds to the viral mRNAs and recruits the RNA exosome to degrade the target RNA. In this article, we provide evidence that a DEXH box RNA helicase, DHX30, is required for optimal antiviral activity of ZAP. Pull-down and co-immunoprecipitation assays demonstrated that DHX30 and ZAP interacted with each other via their N terminal domains. Downregulation of DHX30 with shRNAs reduced ZAP's antiviral activity. These data implicate that DHX30 is a cellular factor involved in the antiviral function of ZAP.
Cytoplasm ; metabolism ; physiology ; DEAD-box RNA Helicases ; metabolism ; Humans ; Immunoprecipitation ; Protein Binding ; physiology ; RNA ; metabolism ; physiology ; RNA Helicases ; metabolism ; physiology ; RNA, Messenger ; metabolism ; physiology ; RNA, Viral ; metabolism ; RNA-Binding Proteins ; metabolism

OBJECTIVE: To detect the expression of nucleolin in cardiac hypertrophy rats induced by pressure overload. METHODS: A total of 40 SD rats with body weight 180 g and 220 g were recruited and randomly divided into 2 groups: a transverse aortic constriction (TAC) group and a sham surgery group. Cardiac hypertrophy model was employed by transverse aortic constriction surgery. Then 2 weeks and 4 weeks after the experiment, the heart mass index (HMI), left ventricle mass index (LVMI) were measured. β-MHC mRNA in the heart tissue was detected with RT-PCR. Nucleolin in the heart, brain and kidney was respectively detected with Western blot. RESULTS: Compared with the sham surgery group, HMI, LVMI in the TAC group increased significantly (P<0.01) 4 weeks after the surgery; the expression of β-MHC mRNA in the heart tissue increased (P<0.05) in the TAC group 4 weeks after the surgery; and the expression of nucleolin protein in the heart tissue of the TAC group was remarkably upregulated (P<0.05) 2 weeks after the surgery, with no change in the brain and kidney tissue between the 2 groups. CONCLUSION Expression of nucleolin protein has been upregulated in response to pressure overload, which may suggest that nucleolin plays a role in cardiac hypertrophy induced by pressure overload.
Animals ; Blood Pressure ; Cardiomegaly ; metabolism ; Myocardium ; metabolism ; Phosphoproteins ; metabolism ; RNA, Messenger ; RNA-Binding Proteins ; metabolism ; Rats ; Rats, Sprague-Dawley

Caveolae are specialized lipid rafts that form flask-shaped invaginations of the plasma membrane. Many researches show that caveolae are involved in cell signaling and transport. Caveolin-1 is the major coat protein essential for the formation of caveolae. Recently, several reports indicated that the other caveolae-associated proteins, Cavins, are required for caveola formation and organization. It's worth noting that Cavin-1 could cooperate with Caveolin-1 to accommodate the structural integrity and function of caveolae. Here, we reviewed that the relationship between Cavins and Caveolins and explore the role of them in regulating caveolae.
Animals ; Caveolae ; physiology ; Caveolin 1 ; metabolism ; physiology ; Caveolins ; metabolism ; physiology ; Humans ; Membrane Proteins ; metabolism ; physiology ; RNA-Binding Proteins ; metabolism ; physiology

OBJECTIVETo identify the interaction partners of a new splicing product of LMO2 gene (LMO2-C), and study its function in K562 cells.

METHODSMaltose binding protein (MBP) pull down and mammalian two-hybrid assay (MTHA) were used to identify the interaction partners of LMO2-C in K562 cells. Semiquantitative RT-PCR was used to detect the expression of hematopoietic specific gene glycoprotein (GPA) in K562 cells.

RESULTSMBP-LMO2-C fusion protein was expressed and purified in soluble form successfully. Endogenous GATA1 and LDB1 proteins were confirmed to bind to LMO2-C by MBP pull down analysis. The MTHA also showed that LMO2-C had comparable binding affinities to LDB1 with LMO2-L, and over expression of LMO2-C prevented LMO2-L from binding to LDB1, the inhibition rate being (81.13 +/- 0.68)%. RT-PCR results showed that the expression level of GPA was reduced [(51.00 +/- 1.58)%] in K562 cells while LMO2-C overexpressed.

CONCLUSIONLMO2-C can bind endogenous GATA1 and LDB1 protein in K562 cells and down regulates the expression of GPA.

Adaptor Proteins, Signal Transducing ; DNA-Binding Proteins ; genetics ; metabolism ; GATA1 Transcription Factor ; metabolism ; Humans ; K562 Cells ; LIM Domain Proteins ; Maltose-Binding Proteins ; Metalloproteins ; genetics ; metabolism ; Periplasmic Binding Proteins ; Proto-Oncogene Proteins ; RNA Splicing ; Transcription Factors ; metabolism ; Two-Hybrid System Techniques