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DOE OSTI2024

Long- and short-read sequencing methods discover distinct circular RNA pools in Lotus japonicus

Circular RNAs (circRNAs) are covalently closed single-stranded RNAs, generated through a back-splicing process that links a downstream 5′ site to an upstream 3′ end. The only distinction in the sequence between circRNA and their linear cognate RNA is the back splice junction. Their low abundance and sequence similarity with their linear origin RNA have made the discovery and identification of circRNA challenging. We have identified almost 6000 novel circRNAs from Lotus japonicus leaf tissue using different enrichment, amplification, and sequencing methods as well as alternative bioinformatics pipelines. The different methodologies identified different pools of circRNA with little overlap. We validated circRNA identified by the different methods using reverse transcription polymerase chain reaction and characterized sequence variations using nanopore sequencing. We compared validated circRNA identified in L. japonicus to other plant species and showed conservation of high-confidence circRNA-expressing genes. This is the first identification of L. japonicus circRNA and provides a resource for further characterization of their function in gene regulation. CircRNAs identified in this study originated from genes involved in all biological functions of eukaryotic cells. The comparison of methodologies and technologies to sequence, identify, analyze, and validate circRNA from plant tissues will enable further research to characterize the function and biogenesis of circRNA in L. japonicus.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2023

Probing lepton flavor violation at Circular Electron-Positron Colliders

Lepton flavor violation is one of the cleanest probes of physics beyond the standard model. In this work, we explore the sensitivity of the process e + e – → τμ to new physics above the TeV scale at the proposed circular electron-positron colliders FCC-ee and CEPC. We compute the e + e – → τμ cross-section in the Standard Model Effective Field Theory and assess the relevant backgrounds. We compare our sensitivity projections to existing and expected constraints from tau decays and Z decays and find that the future electron-positron colliders provide competitive probes of new physics. We highlight the complementarity of searches for resonant e + e – → Z → τμ production on the Z pole and searches for non-resonant e + e – → τμ at higher center-of-mass energies.

72 PHYSICS OF ELEMENTARY PARTICLES AND FIELDS↗

DOE OSTI2018

Microwave Heating of a Liquid Stably Flowing in a Circular Channel Under the Conditions of Nonstationary Radiative-Convective Heat Transfer

A class of nonlinear problems of nonstationary radiative-convective heat transfer under the microwave action with a small penetration depth is considered in a stabilized coolant flow in a circular channel. The solutions to these problems are obtained, using asymptotic procedures at the stages of nonstationary and stationary convective heat transfer on the heat-radiating channel surface. The nonstationary and stationary stages of the solution are matched, using the 'longitudinal coordinate–time' characteristic. The approximate solutions constructed on such principles correlate reliably with the exact ones at the limiting values of the operation parameters, as well as with numerical and experimental data of other researchers. An important advantage of these solutions is that they allow the determination of the main regularities of the microwave and thermal radiation influence on convective heat transfer in a channel even before performing cumbersome calculations. It is shown that, irrespective of the heat exchange regime (nonstationary or stationary), the Nusselt number decreases and the rate of the surface temperature change increases with increase in the intensity of thermal action.

42 ENGINEERING↗

DOE OSTI2018

Series Solution for Magnetic Flux Density of Circular Current Loop and a Relevant Application

The series solution to the magnetic flux density of a circular current loop is presented in the paper. The convergences of these series are assessed. Comparing the result with elliptic integral method, the validity and feasibility of series solution were verified. A computer program has been developed by the method to calculate the magnetic flux density of a multi-coil magnet and its uniformity successfully.

70 PLASMA PHYSICS AND FUSION TECHNOLOGY↗

DOE OSTI2018

Eccentric Form of the Loss of Stability of a Rotating Elastoplastic Radially Inhomogeneous Stepped Circular Disk

We propose a procedure for the investigation of the possible loss of stability of a rotating elastoplastic radially inhomogeneous stepped circular disk by the method of small parameter. The characteristic equation for the critical radius of the plastic zone is obtained in the first approximation. The values of the critical angular rotational velocity are numerically found for various parameters of the disk.

97 MATHEMATICS AND COMPUTING↗

DOE OSTI2019

Polarization Amplitude-Based Method of Determining the Bearing and Roll Angles of an Object in Motion from Orthogonal Circularly Polarized Signals of a Radio Beacon

The possibility of determining the bearing and roll angles of an object in motion is investigated based on orthogonal left- and right-hand circularly polarized signals simultaneously radiated by a radio beacon from two points with the known coordinates spatially spaced in the horizontal plane. The bearing and roll angles are determined onboard the object in motion using a two-channel receiving system based on polarization amplitude-based processing of resultant vector signals received in a linear polarization basis.

36 MATERIALS SCIENCE↗

DOE OSTI2019

Circular RNA expression and circPTPRM promotes proliferation and migration in hepatocellular carcinoma

Circular RNAs (circRNAs) play a critical role during hepatocellular carcinoma (HCC) development. CircRNA PTPRM (circPTPRM) has not been reported to cause disease and its role in HCC is unclear. This study explored circRNA expression and the function of circPTPRM in HCC. RNA sequencing (RNA-seq) was performed on 3 randomly selected pairs of HCC tissues and their corresponding adjacent non-tumor tissues. Three differentially expressed circRNAs, circPTPRM, circSMAD2 and circPTBP3 were selected and verified by real-time quantitative reverse transcription-polymerase chain reactions in 30 pairs of tissue samples, In vitro cultured hepatoma cells, and normal liver cells. Clinical data analysis was performed to select target circRNAs. Anti-target circRNA siRNAs were transfected into hepatoma cell lines, and the biological behavior of hepatoma cells following silencing of the target circRNA were detected by cell proliferation, plate cloning, and transwell assays. There were 86 differentially expressed circRNAs from RNA-seq, of which 53 were significantly upregulated and 33 were significantly downregulated in HCC. CircPTPRM expression was significantly upregulated in HCC tissue (p = 0.023) based on the analysis of 30 paired samples. CircPTPRM expression positively correlated with HCC recurrence and metastasis (p = 0.039). CircPTPRM silencing reduced HCC cell proliferation, migration and invasion. CircRNAs were differentially expressed in HCC samples. CircPTPRM was significantly upregulated in HCC and may function during the tumorigenesis and metastasis of HCC.

62 RADIOLOGY AND NUCLEAR MEDICINE↗

DOE OSTI2017

Circular RNA profiling reveals that circCOL3A1-859267 regulate type I collagen expression in photoaged human dermal fibroblasts

Production of type I collagen declines is a main characteristic during photoaging, but the mechanism is still not fully understood. Circular RNAs (circRNAs) are a class of newly identified non-coding RNAs with regulatory potency by sequestering miRNAs like a sponge. It's more stable than linear RNAs, and would be a useful tool for regulation of gene expression. However, the role of circRNAs in collagen expression during photoaging is still unclear. Here we performed deep sequencing of RNA generated from UVA irradiated and no irradiated human dermal fibroblasts (HDFs) and identified 29 significantly differentially expressed circRNAs (fold change ≥ 1.5, P < 0.05), 12 circRNAs were up-regulated and 17 circRNAs were down-regulated.3 most differentially expressed circRNAs were verified by qRT-PCR and the down-regulated circCOL3A1-859267 exhibited the most significantly altered in photoaged HDFs. Overexpression of circCOL3A1-859267 inhibited UVA-induced decrease of type I collagen expression and silencing of it reduced type I collagen intensity. Via a bioinformatic method, 44 miRNAs were predicted to binding with circCOL3A1-859267, 5 of them have been confirmed or predicted to interact with type I collagen. This study show that circCOL3A1-859267 regulate type I collagen expression in photoaged HDFs, suggesting it may be a novel target for interfering photoaging.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

Circular RNA expression profiles in placental villi from women with gestational diabetes mellitus

Highlights: • More than 40,000 circRNAs are sequenced in the placental villi. • The expression profile of circRNAs in GDM is different from that in controls. • The potential effects of many differential circRNAs are involve in the development of GDM. • Some differential circRNAs have binding sites of GDM-related miRNAs. Circular RNAs (circRNAs) have recently been shown to exert their effects on multiple pathological processes by acting as microRNA (miRNA) sponges. However, the roles of circRNAs in gestational diabetes mellitus (GDM) are largely unknown. This study aimed to identify the circRNAs involved in GDM and predict their potential biological functions. We first performed next-generation sequencing (NGS) to generate unbiased placental villi circRNA expression profiles of GDM and normal controls. In total, 48,270 circRNAs from the placental villi of the two groups were sequenced. Of these, 227 circRNAs were significantly up-regulated and 255 circRNAs were significantly down-regulated. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) biological pathway analyses demonstrated that glycometabolism and lipometabolism processes, which are important in GDM development, were significantly enriched. Further analysis showed that most of the circRNAs harbored miRNA binding sites, and some were associated with GDM. These results showed that circRNAs are aberrantly expressed in the placental villi of GDM patients and play potential roles in the development of GDM.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

Circular RNA circMAN2B2 facilitates lung cancer cell proliferation and invasion via miR-1275/FOXK1 axis

Highlights: • CircMAN2B2 knockdown suppresses lung cancer cell proliferation and invasion. • CircMAN2B2 sponges and represses miR-1275 expression in lung cancer. • miR-1275 exerts its role by regulation of FOXK1 expression in lung cancer. • CircMAN2B2 promotes lung cancer cell proliferation an invasion via circMAN2B2/miR-1275/FOXK1 signaling. Lung cancer remains a leading cause of cancer-related deaths worldwide. In the past years, increasing reports indicate that circular RNAs (circRNAs) exert a great important role in human cancers, including lung cancer. However, the knowledge about circRNA in lung cancer remains very little so far. In the present study, we screened out a highly expressed novel circRNA named circMAN2B2 in lung cancer tissues. We investigated the function of circMAN2B2 and found that circMAN2B2 knockdown significantly inhibited cell proliferation and invasion in both H1299 and A549 lung cancer cells. Mechanistically, we found that circMAN2B2 could sponge miR-1275 to inhibit its level. Through a series of functional experiments, we dissected the role of miR-1275 in lung cancer and proved the anti-tumor role of miR-1275. Furthermore, we found that miR-1275 exerted its role in lung cancer by regulating FOXK1 expression. In addition, we demonstrated that restoration of FOXK1 could rescue circMAN2B2 knockdown-induced repression of cell proliferation and invasion. Taken together, our study demonstrated that circMAN2B2 acts as an oncogenic role in lung cancer through promoting FOXK1 expression by sponging miR-1275.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

Circular RNA circC3P1 suppresses hepatocellular carcinoma growth and metastasis through miR-4641/PCK1 pathway

Highlights: • CircC3P1 was downregulated in HCC tissues. • CircC3P1 overexpression suppressed HCC growth and migration in vitro and in vivo. • CircC3P1 promoted PCK1 expression by sponging miR-4641 in HCC. • CircC3P1 regulated HCC cell proliferation, migration and invasion via modulating PCK1 expression. In the recent years, increasing evidences identify circular RNAs (circRNAs) as a class of important regulators in various human cancers. Nevertheless, the functions and mechanisms of most circRNAs in cancer cells remain largely unknown. In this study, we found out a significantly downregulated circRNA circC3P1 in hepatocellular carcinoma (HCC) tissues compared to adjacent normal tissues. And our results indicated that circC3P1 expression level was negatively correlated with TNM stage, tumor size and vascular invasion in HCC. Moreover, we found that circC3P1 overexpression dramatically inhibited the proliferation, migration and invasion of HCC cells in vitro and in vivo. In terms of mechanism, we found that circC3P1 could promote PCK1 expression through sponging miR-4641 in HCC cells. We showed that miR-4641 expression was negatively correlated with that of either circC3P1 or PCK1 in HCC tissues. Finally, by functional experiments, we demonstrated that knockdown of PCK1 significantly attenuated the effects of circC3P1 overexpression on HCC cell proliferation, migration and invasion. In summary, our findings illustrated that circC3P1 acts as a tumor suppressor via enhancing PCK1 expression by sponging miR-4641 in HCC.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

Circular RNA hsa{sub c}irc{sub 0}000673 promotes hepatocellular carcinoma malignance by decreasing miR-767-3p targeting SET

Highlights: • Hsa{sub c}irc{sub 0}000673 was highly expressed in HCC tissues. • Hsa{sub c}irc{sub 0}000673 knockdown suppressed HCC cell proliferation and invasion. • Hsa{sub c}irc{sub 0}000673 promoted HCC proliferation and invasion by sponging miR-767-3p. • Hsa{sub c}irc{sub 0}000673 facilitated HCC cell proliferation and invasion via regulating miR-767-3p/SET pathway. The importance of circular RNAs (circRNAs) in human cancers has gradually been acknowledged. In hepatocellular carcinoma (HCC), several circRNAs have been reported to regulate tumor growth and metastasis. However, the role of hsa{sub c}irc{sub 0}000673 in HCC remains largely unknown. In this study, we found that hsa{sub c}irc{sub 0}000673 was significantly upregulated in HCC tissues compared to adjacent non-tumor tissues. Moreover, we found that hsa{sub c}irc{sub 0}000673 knockdown markedly inhibited the proliferation and invasion of HCC cells in vitro. Besides, hsa{sub c}irc{sub 0}000673 silence led to delayed tumor growth in vivo. In terms of mechanism, we showed that hsa{sub c}irc{sub 0}000673 directly associated with miR-767-3p in HCC cells. Via inhibiting miR-767-3p, hsa{sub c}irc{sub 0}000673 promoted HCC cell proliferation and invasion. Furthermore, we demonstrated that SET was a downstream effector of hsa{sub c}irc{sub 0}000673/miR-767-3p signaling. We showed that miR-767-3p could significantly promote SET expression by sponging miR-767-3p in HCC cells. Finally, rescue assays indicated that SET expression was essential for the effects of hsa{sub c}irc{sub 0}000673/miR-767-3p signaling on HCC cell proliferation and invasion. Taken together, our findings demonstrated that hsa{sub c}irc{sub 0}000673 promoted HCC malignant behaviors via regulating miR-767-3p/SET pathway.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

Circular RNA circNASP modulates the malignant behaviors in osteosarcoma via miR-1253/FOXF1 pathway

Highlights: • circNASP knockdown suppressed OS cell proliferation and invasion. • circNASP acted as a sponge of miR-1253 targeting FOXF1. • miR-1253 inhibitor reversed the effects of circNASP knockdown in OS cells. • circNASP modulated OS cell proliferation and invasion by miR-1253/FOXF1 pathway. Increasing evidences demonstrate that circular RNAs (circRNAs) serve as essential regulators in various human diseases, including cancer. However, the study on circRNA function in osteosarcoma (OS) is just emerging. In the present study, we screened out a novel circRNA termed circNASP which was significantly upregulated in OS tissues compared to adjacent normal tissues. We found that circNASP knockdown dramatically inhibited the proliferation, cell cycle progression and invasion of OS cells. Moreover, we showed that circNASP expression was positively correlated with tumor size and metastasis in OS patients. In terms of mechanism, we found that circNASP acts as a sponge of miR-1253 targeting FOXF1 in OS cells. By inhibiting miR-1253 availability, circNASP promoted FOXF1 expression. Rescue assays indicated that inhibition of miR-1253 could significantly reverse the effects of circNASP knockdown on OS cell proliferation and invasion while restoration of FOXF1 rescued the proliferation and invasion of OS cells transfected with miR-1253 mimics. Taken together, our findings demonstrated that circNASP contributes to malignant behaviors of OS cells by miR-1253/FOXF1 pathway, which suggested circNASP/miR-1253/FOXF1axis might be a potential therapeutic target.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

Circular RNA expression profiles in hippocampus from mice with perinatal glyphosate exposure

Glyphosate is the active ingredient in numerous herbicide formulations. The roles of glyphosate in embryo-toxicity and neurotoxicity have been reported in human and animal models. Recently, several studies have reported evidence linking neurodevelopmental disorders (NDDs) with gestational glyphosate exposure. However, the role of glyphosate in neuronal development is still not fully understood. Our previous study found that perinatal glyphosate exposure resulted in differential microRNA expression in the prefrontal cortex of mouse offspring. However, the mechanism of glyphosate-induced neurotoxicity in the developing brain is still not fully understood. Considering the pivotal role of Circular RNAs (circRNAs) in the regulation of gene expression, a circRNA microarray method was used in this study to investigate circRNA expression changes in the hippocampus of mice with perinatal glyphosate exposure. The circRNA microarrays revealed that 663 circRNAs were significantly altered in the perinatal glyphosate exposure group compared with the control group. Among them, 330 were significantly upregulated, and the other 333 were downregulated. Furthermore, the relative expression levels of mmu-circRNA-014015, mmu-circRNA-28128 and mmu-circRNA-29837 were verified using quantitative real-time polymerase chain reaction (qRT-PCR). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses demonstrated that stress-associated steroid metabolism pathways, such as aldosterone synthesis and secretion pathways, may be involved in the neurotoxicity of glyphosate. These results showed that circRNAs are aberrantly expressed in the hippocampus of mice with perinatal glyphosate exposure and play potential roles in glyphosate-induced neurotoxicity.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

Circular RNA hsa{sub c}irc{sub 0}023404 exerts an oncogenic role in cervical cancer through regulating miR-136/TFCP2/YAP pathway

Cervical cancer (CC) is one of the most prevalent malignances among women. However, the mechanism underlying CC development remains elusive. Recently, circular RNAs (circRNAs) have been known as important regulators in tumorigenesis. Whether circRNAs are involved in CC requires to be determined. In the present study, we found that circRNA hsa{sub c}irc{sub 0}023404 was significantly upregulated in CC tissues compared to adjacent normal tissues. And its overexpression was correlated with poor prognosis in CC patients. Functionally, we showed that knockdown of hsa{sub c}irc{sub 0}023404 significantly suppressed the proliferation, arrested the cell-cycle progression and inhibited cell migration and invasion in CC. In terms of mechanism, we found that hsa{sub c}irc{sub 0}023404 acted as a sponge of miR-136 and miR-136 targeted TFCP2, which is an activator of YAP signaling pathway. We showed that hsa{sub c}irc{sub 0}023404 activated YAP pathway in CC via promoting TFCP2 expression by sponging miR-136, leading to CC development and progression. Taken together, our study for the first time demonstrated the pivot role of hsa{sub c}irc{sub 0}023404 and revealed a novel regulatory loop of hsa{sub c}irc{sub 0}023404/miR-136/TFCP2/YAP axis in CC progression.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

Identification and characterization of circular RNAs in rapid atrial pacing dog atrial tissue

Highlights: • Revealed the expression characterization of circRNAs in dog atrial tissue. • Compared differentially expressed circRNAs between atrial fibrillation and control dog. • Found extensive interaction among circRNAs and AF related miRNAs and mRNAs. Circular RNAs (circRNAs) have emerged as novel molecules of interest in gene regulation as other noncoding RNAs, and participating in the process of many diseases. However, the expression and functions of circRNAs in Rapid atrial pacing (RAP) dog atrial tissue still unknown. 12 canines were randomly assigned to control and pacing group. RAP at 500 beats per minute was maintained 14 days in the pacing group. The expression characterization of circRNAs were revealed by high-throughput sequencing. We totally predicted 15,990 circRNAs in dog atrial tissues. Moreover, we found 146 differentially expressed circRNAs between control and RAP dogs. Five circRNAs were selected for subsequent RT-PCR validation, and four circRNAs confirmed with the high throughput sequencing analysis. GO analysis showed that the differentially expressed circRNAs might involve in the process of “structural constituent of cytoskeleton, ion channel activity”. We explored the circRNA-miRNA interaction network, and found extensive interaction among differentially expressed circRNAs and AF related miRNAs and mRNAs. Our work firstly identified the characterization of circRNAs in the dog atrial, and revealed the differentially expressed circRNAs in the RAP dog, this might lay a solid foundation on the function of circRNA in the mechanisms of AF.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

Circular RNA hsa{sub c}irc{sub 0}008039 promotes breast cancer cell proliferation and migration by regulating miR-432-5p/E2F3 axis

As the development of sequencing technology, more and more circular RNAs (circRNAs) are identified in human cancer tissues. Increasing evidences imply circRNAs are important regulators in tumor progression. Nevertheless, how circRNAs participate in breast cancer development and progression is not well understood. In the present study, we identified a novel circRNA hsa{sub c}irc{sub 0}008039 with upregulated expression level in breast cancer tissues. By functional experiments, we found that hsa{sub c}irc{sub 0}008039 depletion significantly suppressed the proliferation, arrested cell-cycle progression and reduced migration in breast cancer. Mechanistic investigations suggested that hsa{sub c}irc{sub 0}008039 served as a competing endogenous RNA (ceRNA) of miR-432-5p. Subsequently, E2F3 was identified as the functional target of miR-432-5p and overexpression of hsa{sub c}irc{sub 0}008039 elevated E2F3 expression in breast cancer. On the whole, our study indicated that hsa{sub c}irc{sub 0}008039 exerted oncogenic roles in breast cancer and suggested the hsa{sub c}irc{sub 0}008039/miR-432-5p/E2F3 axis might be a potential therapeutic target.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

Circular RNA circZFR contributes to papillary thyroid cancer cell proliferation and invasion by sponging miR-1261 and facilitating C8orf4 expression

In recent years, more and more circular RNAs (circRNAs) have been identified in multiple tissues and cells. Increasing evidences show circRNAs play important roles in human cancers. However, the role of circRNAs in papillary thyroid carcinoma (PTC) remains largely unknown. In this study, we identified a new circRNA circZFR that was significantly upregulated in PTC tissues compared to adjacent normal tissues. Furthermore, circZFR expression level was negatively correlated with clinical severity. We found that circZFR knockdown dramatically inhibited the proliferation, migration and invasion of PTC cells in vitro. Mechanistically, we found circZFR could promote C8orf4 expression via serving as a competing endogenous RNA (ceRNA) of miR-1261 in PTC cells. Rescue assays indicated that restoration of C8orf4 significantly attenuated the inhibitory effects of circZFR knockdown on PTC cell proliferation, migration and invasion. In summary, our findings demonstrated that circRNA circZFR exerted oncogenic roles via regulating miR-1261/C8orf4 axis in PTC, which suggested circZFR might be a potential therapeutic target.

60 APPLIED LIFE SCIENCES↗