Relationship between porcine miR-20a and its putative target low-density lipoprotein receptor based on dual luciferase reporter gene assays

Yueyun Ding,Shujiao Zhu,Chaodong Wu,Li Qian,DengTao Li,Li Wang,Yuanlang Wang,Wei Zhang,Min Yang,Jian Ding,Xudong Wu,Xiao-Dong Zhang,Yafei Gao,Zongjun Yin 아세아·태평양축산학회 2019 Animal Bioscience Vol.32 No.7

Objective: Mutations in low-density lipoprotein receptor (LDLR), which encodes a critical protein for cholesterol homeostasis and lipid metabolism in mammals, are involved in cardiometabolic diseases, such as familial hypercholesterolemia in pigs. Whereas microRNAs (miRNAs) can control LDLR regulation, their involvement in circulating cholesterol and lipid levels with respect to cardiometabolic diseases in pigs is unclear. We aimed to identify and analyze LDLR as a potential target gene of SSC-miR-20a. Methods: Bioinformatic analysis predicted that porcine LDLR is a target of SSC-miR-20a. Wild-type and mutant LDLR 3′-untranslated region (UTR) fragments were generated by polymerase chain reaction (PCR) and cloned into the pGL3-Control vector to construct pGL3 Control LDLR wild-3′-UTR and pGL3 Control LDLR mutant-3′-UTR recombinant plasmids, respectively. An miR-20a expression plasmid was constructed by inserting the porcine pre-miR-20a-coding sequence between the HindIII and BamHI sites in pMR-mCherry, and constructs were confirmed by sequencing. HEK293T cells were co-transfected with the miR-20a expression or pMR-mCherry control plasmids and constructs harboring the corresponding 3′-UTR, and relative luciferase activity was determined. The relative expression levels of miR-20a and LDLR mRNA and their correlation in terms of expression levels in porcine liver tissue were analyzed using reverse-transcription quantitative PCR. Results: Gel electrophoresis and sequencing showed that target gene fragments were successfully cloned, and the three recombinant vectors were successfully constructed. Compared to pMR-mCherry, the miR-20a expression vector significantly inhibited wild-type LDLR-3′-UTR-driven (p<0.01), but not mutant LDLR-3′-UTR-driven (p>0.05), luciferase reporter activity. Further, miR-20a and LDLR were expressed at relatively high levels in porcine liver tissues. Pearson correlation analysis revealed that porcine liver miR-20a and LDLR levels were significantly negatively correlated (r = –0.656, p<0.05). Conclusion: LDLR is a potential target of miR-20a, which might directly bind the LDLR 3′-UTR to post-transcriptionally inhibit expression. These results have implications in understanding the pathogenesis and progression of porcine cardiovascular diseases.

Relationship among porcine lncRNA TCONS_00010987, miR-323, and leptin receptor based on dual luciferase reporter gene assays and expression patterns

Ding, Yueyun,Qian, Li,Wang, Li,Wu, Chaodong,Li, DengTao,Zhang, Xiaodong,Yin, Zongjun,Wang, Yuanlang,Zhang, Wei,Wu, Xudong,Ding, Jian,Yang, Min,Zhang, Liang,Shang, Jinnan,Wang, Chonglong,Gao, Yafei Asian Australasian Association of Animal Productio 2020 Animal Bioscience Vol.33 No.2

Objective: Considering the physiological and clinical importance of leptin receptor (LEPR) in regulating obesity and the fact that porcine LEPR expression is not known to be controlled by lncRNAs and miRNAs, we aim to characterize this gene as a potential target of SSC-miR-323 and the lncRNA TCONS_00010987. Methods: Bioinformatics analyses revealed that lncRNA TCONS_00010987 and LEPR have SSC-miR-323-binding sites and that LEPR might be a target of lncRNA TCONS_00010987 based on cis prediction. Wild-type and mutant TCONS_00010987-target sequence fragments and wild-type and mutant LEPR 3'-UTR fragments were generated and cloned into pmiRRB-REPORT^TM-Control vectors to construct respective recombinant plasmids. HEK293T cells were co-transfected with the SSC-miR-323 mimics or a negative control with constructs harboring the corresponding binding sites and relative luciferase activities were determined. Tissue expression patterns of lncRNA TCONS_00010987, SSC-miR-323, and LEPR in Anqing six-end-white (AQ, the obese breed) and Large White (LW, the lean breed) pigs were detected by real-time quantitative polymerase chain reaction; backfat expression of LEPR protein was detected by western blotting. Results: Target gene fragments were successfully cloned, and the four recombinant vectors were constructed. Compared to the negative control, SSC-miR-323 mimics significantly inhibited luciferase activity from the wild-type TCONS_00010987-target sequence and wild-type LEPR-3'-UTR (p<0.01 for both) but not from the mutant TCONS_00010987-target sequence and mutant LEPR-3'-UTR (p>0.05 for both). Backfat expression levels of TCONS_00010987 and LEPR in AQ pigs were significantly higher than those in LW pigs (p<0.01), whereas levels of SSC-miR-323 in AQ pigs were significantly lower than those in LW pigs (p<0.05). LEPR protein levels in the backfat tissues of AQ pigs were markedly higher than those in LW pigs (p<0.01). Conclusion: LEPR is a potential target of SSC-miR-323, and TCONS_00010987 might act as a sponge for SSC-miR-323 to regulate LEPR expression.

Inequalities in Awareness and Attitude towards HPV and Its Vaccine between Local and Migrant Residents Who Participated in Cervical Cancer Screening in Shenzhen, China

Wei Lin,Yueyun Wang,Zhihua Liu,Bin Chen,Shixin Yuan,Bo Wu,Lin Gong 대한암학회 2020 Cancer Research and Treatment Vol.52 No.1

Purpose A cross-sectional survey was conducted to evaluate the differences on awareness and attitude towards human papillomavirus (HPV) and its vaccine between local and migrant residents who participated in cervical cancer screening in Shenzhen, China. Materials and Methods A total of 9,855 females sampled from healthcare institutions in 20 street blocks through the Cervical Cancer Prevention Network were surveyed in this study by a self-administered questionnaire. Multivariate logistic regression was conducted to explore the role of the hukou and resident status in the willingness to receive HPV vaccination. Results Local residents had a relatively higher awareness of HPV (62.0% vs. 35.6% vs. 29.9%, p < 0.001) and its vaccine (35.3% vs. 15.4% vs. 14.8%, p < 0.001), as well as a higher willingness to receive HPV vaccination (68.5% vs. 62.5% vs. 56.2%, p < 0.001) than non-permanent residents and floating population. Except for age, education level, marital status, monthly income, having daughter(s), and heard of HPV and its vaccine, the hukou and resident status significantly associated with the willingness to receive HPV vaccination (local residents vs. floating population: odds ratio, 1.216; 95% confidence interval, 1.057 to 1.398). None significant difference on the associated factors was found between local residents and internal migrants (p for interactions > 0.05). Conclusion Inequalities in awareness and attitude towards HPV and its vaccine existed between local and migrant residents in Shenzhen. The hukou and resident status did impact on the willingness to receive HPV vaccination, therefore, it is critical to implement effective health education campaigns on HPV and its vaccine among internal migrants.

Cyanide- and Phenoxo-Bridged Heterobimetallic Fe(III)-Mn(III) Coordination Polymer: Synthesis, Crystal Structures and Magnetic Properties

Daopeng Zhang,Lingqian Kong,Yueyun Li,Ping Wang,Xia Chen 대한화학회 2014 Bulletin of the Korean Chemical Society Vol.35 No.9

Two two-dimensional cyanide- and phenoxo-bridged heterometallic M(II)-Mn(III) (M = Ni, Pd) coordination polymers {[Mn(saltmen)]4[Ni(CN)4]}(ClO4)2·CH3OH·H2O (1) and {[Mn(saltmen)]4[Pd(CN)4]}(ClO4)2· CH3CN·H2O (2) (saltmen2– = N,N'-(1,1,2,2-tetramethylethylene)bis(salicylideneaminato)dianion) have been obtained by using K2[M(CN)4] as building blocks and a salen-tpye Schiff-base manganese(III) compound as assembling segment. Single X-ray analysis reveals their isostrutural cyanide-bridged MMn4 pentanuclear cationic structure. The four Schiff base manganese units of the pentanuclear entity are self-complementary through the phenoxo oxygen atoms from the neighboring complex, therefore forming cyanide- and phenoxobridged 2D sheet-like structure. Investigation over magnetic susceptibilities reveals the overall ferromagnetic coupling between the adjacent Mn(III) ions bridged by the phenoxo oxygen atoms with J = 2.13 and 2.21 cm−1 for complexes 1 and 2, respectively.

Genomic analysis reveals selection signatures of the Wannan Black pig during domestication and breeding

Zhang, Wei,Yang, Min,Wang, Yuanlang,Wu, Xudong,Zhang, Xiaodong,Ding, Yueyun,Yin, Zongjun Asian Australasian Association of Animal Productio 2020 Animal Bioscience Vol.33 No.5

Objective: The Wannan Black pig is a typical Chinese indigenous, disease-resistant pig breed with high fertility, and a crude-feed tolerance that has been bred by artificial selection in the south of Anhui province for a long time. However, genome variation, genetic relationships with other pig breeds, and domestication, remain poorly understood. Here, we focus on elucidating the genetic characteristics of the Wannan Black pig and identifying selection signatures during domestication and breeding. Methods: We identified the whole-genome variation in the Wannan Black pig and performed population admixture analyses to determine genetic relationships with other domesticated pig breeds and wild boars. Then, we identified the selection signatures between the Wannan Black pig and Asian wild boars in 100-kb windows sliding in 10 kb steps by using two approaches: the fixation index (F_ST) and π ratios. Results: Resequencing the Wannan Black pig genome yielded 501.52 G of raw data. After calling single-nucleotide variants (SNVs) and insertions/deletions (InDels), we identified 21,316,754 SNVs and 5,067,206 InDels (2,898,582 inserts and 2,168,624 deletions). Additionally, we found genes associated with growth, immunity, and digestive functions. Conclusion: Our findings help in explaining the unique genetic and phenotypic characteristics of Wannan Black pigs, which in turn can be informative for future breeding programs of Wannan Black pigs.

Cyanide- and Phenoxo-Bridged Heterobimetallic Fe(III)-Mn(III) Coordination Polymer: Synthesis, Crystal Structures and Magnetic Properties

Zhang, Daopeng,Kong, Lingqian,Li, Yueyun,Wang, Ping,Chen, Xia Korean Chemical Society 2014 Bulletin of the Korean Chemical Society Vol.35 No.9

Two two-dimensional cyanide- and phenoxo-bridged heterometallic M(II)-Mn(III) (M = Ni, Pd) coordination polymers $\{[Mn(saltmen)]_4[Ni(CN)_4]\}(ClO_4)_2{\cdot}CH_3OH{\cdot}H_2O$ (1) and $\{[Mn(saltmen)]_4[Pd(CN)_4]\}(ClO_4)_2{\cdot}CH_3CN{\cdot}H_2O$ (2) ($saltmen^{2-}$ = N,N'-(1,1,2,2-tetramethylethylene)bis(salicylideneaminato)dianion) have been obtained by using $K_2[M(CN)_4]$ as building blocks and a salen-tpye Schiff-base manganese(III) compound as assembling segment. Single X-ray analysis reveals their isostrutural cyanide-bridged $MMn_4$ pentanuclear cationic structure. The four Schiff base manganese units of the pentanuclear entity are self-complementary through the phenoxo oxygen atoms from the neighboring complex, therefore forming cyanide- and phenoxo-bridged 2D sheet-like structure. Investigation over magnetic susceptibilities reveals the overall ferromagnetic coupling between the adjacent Mn(III) ions bridged by the phenoxo oxygen atoms with J = 2.13 and $2.21cm^{-1}$ for complexes 1 and 2, respectively.

Genome-wide scan for runs of homozygosity identifies candidate genes in Wannan Black pigs

Wu Xudong,Zhou Ren,Zhang Wei,Cao Bangji,Xia Jingjing,Wang Caiyun,Zhang Xiaodong,ChuMing-Xing,Yin Zongjun,Ding Yueyun 아세아·태평양축산학회 2021 Animal Bioscience Vol.34 No.12

Objective: Runs of homozygosity (ROH) are contiguous lengths of homozygous genotypes that can reveal inbreeding levels, selection pressure, and mating schemes. In this study, ROHs were evaluated in Wannan Black pigs to assess the inbreeding levels and the genome regions with high ROH frequency.Methods: In a previous study, we obtained 501.52 GB of raw data from resequencing (10×) of the genome and identified 21,316,754 single-nucleotide variants in 20 Wannan Black pig samples. We investigated the number, length, and frequency of ROH using resequencing data to characterize the homozygosity in Wannan Black pigs and identified genomic regions with high ROH frequencies.Results: In this work, 1,813 ROHs (837 ROHs in 100 to 500 kb, 449 ROHs in 500 to 1,000 kb, 527 ROHs in >1,000 kb) were identified in all samples, and the average genomic inbreeding coefficient (FROH) in Wannan Black pigs was 0.5234. Sixty-one regions on chromosomes 2, 3, 7, 8, 13, 15, and 16 harbored ROH islands. In total, 105 genes were identified in 42 ROH islands, among which some genes were related to production traits.Conclusion: This is the first study to identify ROH across the genome of Wannan Black pigs, the Chinese native breed of the Anhui province. Overall, Wannan Black pigs have high levels of inbreeding due to the influence of ancient and recent inbreeding due to the genome. These findings are a reliable resource for future studies and contribute to save and use the germplasm resources of Wannan Black pigs. Objective: Runs of homozygosity (ROH) are contiguous lengths of homozygous genotypes that can reveal inbreeding levels, selection pressure, and mating schemes. In this study, ROHs were evaluated in Wannan Black pigs to assess the inbreeding levels and the genome regions with high ROH frequency. Methods: In a previous study, we obtained 501.52 GB of raw data from resequencing (10×) of the genome and identified 21,316,754 single-nucleotide variants in 20 Wannan Black pig samples. We investigated the number, length, and frequency of ROH using resequencing data to characterize the homozygosity in Wannan Black pigs and identified genomic regions with high ROH frequencies. Results: In this work, 1,813 ROHs (837 ROHs in 100 to 500 kb, 449 ROHs in 500 to 1,000 kb, 527 ROHs in >1,000 kb) were identified in all samples, and the average genomic inbreeding coefficient (FROH) in Wannan Black pigs was 0.5234. Sixty-one regions on chromosomes 2, 3, 7, 8, 13, 15, and 16 harbored ROH islands. In total, 105 genes were identified in 42 ROH islands, among which some genes were related to production traits. Conclusion: This is the first study to identify ROH across the genome of Wannan Black pigs, the Chinese native breed of the Anhui province. Overall, Wannan Black pigs have high levels of inbreeding due to the influence of ancient and recent inbreeding due to the genome. These findings are a reliable resource for future studies and contribute to save and use the germplasm resources of Wannan Black pigs.

Cervical Cancer Screening Rate and Willingness among Female Migrants in Shenzhen, China: Three-Year Changes in Citywide Surveys

Wei Lin,Bin Chen,Bo Wu,Shixin Yuan,Chuyan Zhong,Weikang Huang,Haiyan Hu,Zhihua Liu,Yueyun Wang 대한암학회 2021 Cancer Research and Treatment Vol.53 No.1

Purpose This study attempted to detect the changes of cervical cancer screening rate and willingness among female migrants, and the associated socio-demographic factors in Shenzhen city. Materials and Methods Two citywide surveys were conducted using a multistage random cluster sampling method in 2011 and 2014, respectively. Data on demographic characteristics, screening participation, and willingness to screen were collected. Logistic regression models were applied to detect possible associated socio-demographic characteristics, and their variations with survey years. Results In total, 12,017 female migrants were enrolled, with a mean age (standard deviation) of 36.73 (6.55) years. From 2011 to 2014, the screening rate increased (25.8% vs. 35.1%, p < 0.001), while the willingness to screen remained stable (82.2% vs. 82.8%, p=0.46). Overall, socio-demographic characteristics of female migrants, including age, marital status, education, monthly income, employment, and medical insurance, were found to be positively associated with screening participation. Similar impacts in relation to willingness were observed except for age. However, these associations varied with survey years, mainly in the contributions of education and monthly income to screening participation, as well as age, monthly income, and medical insurance to willingness of being screened. Conclusion Identifying changes of associated socio-demographic factors precisely is warranted of necessity, which provides novel clues to adjust targeted actions regularly in promoting cervical cancer screening participation among female migrants in Shenzhen.