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3′ end processing is required for the maturation of all eukaryotic RNAs. Current model suggests that canonical mRNA 3′ processing is carried out exclusively within a protein complex termed mRNA 3′ processing complex. In a recent study, by using RNA-biotin based pull-down assay and high-throughput sequencing, we reported that a subset of small nucleolar RNAs (snoRNAs) were physically associated with this macromolecular machinery. Through detailed characterization of one of these snoRNAs, SNORD50A, we revealed that non-coding RNA, such as snoRNA, may play a regulatory role in mRNA 3′ processing. Our results provided novel insight into both the regulatory mechanism of mRNA 3′ processing and the non-canonical functions of snoRNAs.

mRNA 3′ end processing is an essential step in gene expression. It is well established that canonical eukaryotic pre-mRNA 3′ processing is carried out within a macromolecular machinery consisting of dozens of trans-acting proteins. However, it is unknown whether RNAs play any role in this process. Unexpectedly, we found that a subset of small nucleolar RNAs (snoRNAs) are associated with the mammalian mRNA 3′ processing complex. These snoRNAs primarily interact with Fip1, a component of cleavage and polyadenylation specificity factor (CPSF). We have functionally characterized one of these snoRNAs and our results demonstrated that the U/A-rich SNORD50A inhibits mRNA 3′ processing by blocking the Fip1-poly(A) site (PAS) interaction. Consistently, SNORD50A depletion altered the Fip1–RNA interaction landscape and changed the alternative polyadenylation (APA) profiles and/or transcript levels of a subset of genes. Taken together, our data revealed a novel function for snoRNAs and provided the first evidence that non-coding RNAs may play an important role in regulating mRNA 3′ processing.

// Yongpeng Tong 1 , Chunliu Huang 2 and Junfang Zhang 3 1 College of Physics and Energy, Shenzhen University, Shenzhen, 518060, China 2 School of Medicine, Sun Yat-Sen University, Guangzhou, 510080, China 3 School of Medicine, Shenzhen University, Shenzhen Second People's Hospital, Shenzhen, 518060, China Correspondence to: Junfang Zhang, email: junfang.zhang@163.com Keywords: cAMP-H 3 BO 3 complex, EGFR-TKI, thermal therapy, NSCLC Received: February 01, 2017 Accepted: April 21, 2017 Published: May 05, 2017 ABSTRACT Purpose: Although EGFR-TKIs (epidermal growth factor receptor tyrosine kinase inhibitors) induce favorable responses as first-line non-small cell lung cancer treatments, drug resistance remains a serious problem. Meanwhile, thermal therapy also shows promise as a cancer therapy strategy. Here we combine a novel EGFR-TKI treatment with thermal therapy to improve lung cancer treatment outcomes. Results: The results suggest that the cAMP-H 3 BO 3 complex effectively inhibits EGFR auto-phosphorylation, while inducing apoptosis and cell cycle arrest in vitro . Compared to the negative control, tumor growth was significantly suppressed in mice treated with oxidative phosphorylation uncoupler thyroxine sodium and either cAMP-H 3 BO 3 complex or cAMP-H 3 BO 3 complex ( P < 0.05). Moreover, the body temperature increase induced by treatment with thyroxine sodium inhibited tumor growth. Immunohistochemical analyses showed that A549 cell apoptosis was significantly higher in the cAMP-H 3 BO 3 complex plus thyroxine sodium treatment group than in the other groups. Moreover,Ca 2+ content analysis showed that the Ca 2+ content of tumor tissue was significantly higher in the cAMP-H 3 BO 3 complex plus thyroxine sodium treatment group than in other groups. Materials and Methods: Inhibition of EGFR auto-phosphorylation by cAMP and cAMP-H 3 BO 3 complex was studied using autoradiography and western blot. The antitumor activity of the novel EGFR inhibitor (cAMP-H 3 BO 3 complex) with or without an oxidative phosphorylation uncoupler (thyroxine sodium) was investigated in vitro and in a nude mouse xenograft lung cancer model incorporating human A549 cells. Conclusions: cAMP-H 3 BO 3 complex is a novel EGFR-TKI. Combination therapy using cAMP-H 3 BO 3 with thyroxine sodium-induced thermal therapy may improve lung cancer treatment outcomes.

Ahead of Print article withdrawn by publisher.

It is increasingly appreciated that U1 snRNP transcriptomically suppresses the usage of intronic polyadenylation site (PAS) of mRNAs, an outstanding question is why frequently used PASs are not suppressed. Here we found that U1 snRNP could be transiently associated with sequences upstream of actionable PASs in human cells, and RNA-RNA interaction might contribute to the association. By focusing on individual PAS, we showed that the stable assembly of U1 snRNP near PAS might be generally required for U1 inhibition of mRNA 3' processing. Therefore, actionable PASs that often lack optimal U1 snRNP docking site nearby is free from U1 inhibitory effect. Consistently, natural 5' splicing site (5'-SS) is moderately enriched ~250 nt upstream of intronic PASs whose usage is sensitive to functional knockdown of U1 snRNA. Collectively, our results provided an insight into how U1 snRNP selectively inhibits the usage of PASs in a cellular context, and supported a prevailing model that U1 snRNP scans pre-mRNA through RNA-RNA interaction to find a stable interaction site to exercise its function in pre-mRNA processing, including repressing the usage of cryptic PASs.