SummaryGene expression is dynamically regulated in a variety of mammalian physiologies. During mammalian aging, there are changes that occur in protein expression that are highly controlled by the regulatory steps in transcription, post‐transcription, and post‐translation. Although there are global profiles of human transcripts during the aging processes available, the mechanism(s) by which transcripts are differentially expressed between young and old cohorts remains unclear. Here, we report on N6‐methyladenosine (m6A) RNA modification profiles of human peripheral blood mononuclear cells (PBMCs) from young and old cohorts. An m6A RNA profile identified a decrease in overall RNA methylation during the aging process as well as the predominant modification on proteincoding mRNAs. The m6A‐modified transcripts tend to be more highly expressed than nonmodified ones. Among the many methylated mRNAs, those of DROSHA and AGO2 were heavily methylated in young PBMCs which coincided with a decreased steady‐state level of AGO2 mRNA in the old PBMC cohort. Similarly, downregulation of AGO2 in proliferating human diploid fibroblasts (HDFs) also correlated with a decrease in AGO2 mRNA modifications and steady‐state levels. In addition, the overexpression of RNA methyltransferases stabilized AGO2 mRNA but not DROSHA and DICER1 mRNA in HDFs. Moreover, the abundance of miRNAs also changed in the young and old PBMCs which are possibly due to a correlation with AGO2 expression as observed in AGO2‐depleted HDFs. Taken together, we uncovered the role of mRNA methylation on the abundance of AGO2 mRNA resulting in the repression of miRNA expression during the process of human aging.
Long non-coding RNAs (lncRNAs) regulate vital biological processes, including cell proliferation, differentiation and development. A subclass of lncRNAs is synthesized from microRNA (miRNA) host genes (MIRHGs) due to pre-miRNA processing, and are categorized as miRNA-host gene lncRNAs (lnc-miRHGs). Presently, the cellular function of most lnc-miRHGs is not well understood. We demonstrate a miRNA-independent role for a nuclear-enriched lnc-miRHG in cell cycle progression. MIR100HG produces spliced and stable lncRNAs that display elevated levels during the G1 phase of the cell cycle. Depletion of MIR100HG-encoded lncRNAs in human cells results in aberrant cell cycle progression without altering the levels of miRNA encoded within MIR100HG. Notably, MIR100HG interacts with HuR/ELAVL1 as well as with several HuR-target mRNAs. Further, MIR100HG-depleted cells show reduced interaction between HuR and three of its target mRNAs, indicating that MIR100HG facilitates interaction between HuR and target mRNAs. Our studies have unearthed novel roles played by a MIRHG-encoded lncRNA in regulating RNA binding protein activity, thereby underscoring the importance of determining the function of several hundreds of lnc-miRHGs that are present in human genome.
Protein dynamics, modifications, and trafficking are all processes that can modulate protein activity. Accumulating evidence strongly suggests that many proteins play distinctive roles dependent on cellular location. Nonsteroidal anti-inflammatory drug activated gene-1 (NAG-1) is a TGF-β superfamily protein that plays a role in cancer, obesity, and inflammation. NAG-1 is synthesized and cleaved into a mature peptide, which is ultimately secreted into the extracellular matrix (ECM). In this study, we have found that full-length NAG-1 is expressed in not only the cytoplasm and ECM, but also in the nucleus. NAG-1 is dynamically moved to the nucleus, exported into cytoplasm, and further transported into the ECM. We have also found that nuclear NAG-1 contributes to inhibition of the Smad pathway by interrupting the Smad complex. Overall, our study indicates that NAG-1 is localized in the nucleus and provides new evidence that NAG-1 controls transcriptional regulation in the Smad pathway.
microRNA (miRNA) and RNA-binding proteins (RBPs) have been studied widely in post-transcriptional gene regulation. Previous work has focused on defining how miRNA and RBPs modulate target mRNA decay and translation as well as investigating how they interplay each other. Emerging studies indicate that certain RBPs other than the AGO-family proteins directly interact with mature miRNAs. These findings implicate competitive binding of RBPs to target miRNAs, sequestration of miRNAs from AGO, promotion of AGO binding to miRNAs, and transfer of miRNAs from RBPs to AGO. Recent work also indicates that AGO-free cytoplasmic miRNAs establish complexes with novel miRNA-binding proteins (miRBPs). This review covers the recent discovery of novel miRBPs, offering a new perspective on the miRNA-mediated gene silencing mechanism. WIREs RNA 2017, 8:e1414. doi: 10.1002/wrna.1414 For further resources related to this article, please visit the WIREs website.
Since the 1980s, growing evidence suggested that the cellular localization of proteins determined their activity and biological functions. In a classical view, a protein is characterized by the single cellular compartment where it primarily resides and functions. It is now believed that when proteins appear in different subcellular locations, the cells surpass the expected activity of proteins given the same genomic information to fulfill complex biological behavior. Many proteins are recognized for having the potential to exist in multiple locations in cells. Dysregulation of translocation may cause cancer or contribute to poorer cancer prognosis. Thus, quantitative and comprehensive assessment of dynamic proteins and associated protein movements could be a promising indicator in determining cancer prognosis and efficiency of cancer treatment and therapy. This review will summarize these so-called moonlighting proteins, in terms of a coupled intracellular cancer signaling pathway. Determination of the detailed biological intracellular and extracellular transit and regulatory activity of moonlighting proteins permits a better understanding of cancer and identification of potential means of molecular intervention.
Purpose The public has paid attention to green tea due to its health benefits. Epigallocatechin-3-gallate (EGCG), the major component of green tea, is well documented to induce apoptosis and cell cycle arrest in cancer cells by targeting multiple signal transduction pathways. However, the detailed mechanism(s) of action needs to be determined. Methods Cell growth was evaluated by MTT assay, cell cycle analysis, and caspase 3/7 activity. Protein expression was analyzed through Western blotting. Reverse transcription polymerase chain reaction was used for examining mRNA expression of p21 and cyclin D1. The promoter activity of p21 was assessed by the luciferase reporter system. Results We identified cyclin D1 and p21 as molecular targets of EGCG in human colorectal cancer cells. We observed that cyclin D1 was down-regulated, while p21 expression was up-regulated by EGCG in dose- and time-dependent manners. Furthermore, we found EGCG decreased cyclin D1 protein stability, therefore triggering ubiquitin-dependent proteasomal degradation. Meanwhile, EGCG increased p21 promoter activity, resulting in up-regulation of p21 mRNA and protein, which was likely dependent on extracellular-signal-regulated kinase (ERK), inhibitor of nuclear factor kappa-B kinase (IKK) and phosphoinositide 3-kinase (PI3 K). Conclusion The data presented here details a novel mechanism by which EGCG inhibits cell growth of colorectal cancer cells. Namely, EGCG-induced cyclin D1 degradation and p21 transcriptional activation partially contribute to growth suppression in these cells.
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