克隆山羊成纤维细胞IGF2-H19基因座甲基化分析

doi:10.11843/j.issn.0366-6964.2017.12.007

Abstract

Abstract:

The aim of this experiment was to study the influence of somatic cell nuclear transfer on IGF2-H19 locus methylation in cloned goat fibroblast cells. Goat ear fibroblast cells (GFC, control group) and cloned goat ear fibroblast cells (CFC, experimental group) were used as experimental materials. When cells in the 2 groups were cultured to passage 5, using cell counting method for cell growth curves, flow cytometer for cell apoptosis, real-time PCR for gene relative expression level and bisulfite sequencing PCR for the methylation status of differentially methylated region (DMR). The results showed that cell growth curves in the 2 groups were typical "S"-shaped, but apoptosis rate in CFC group was increased significantly(P<0.01) compared with that in GFC group. The expression level of Dnmt1 (P<0.01), Dnmt3b (P<0.01), Tet1 (P<0.05), Tet2 (P<0.05), H19 (P<0.05) and IGF2 (P<0.01) in CFC group were decreased significantly compared with that in GFC group. However, no statistical differences were found at the expression levels of Tet3、Dnmt3a and IGF2R between the 2 groups. Compared with GFC group, the methylation level of IGF2 DMR1 (74.1% vs. 57.8%, P<0.01) and DMR2 (76.8% vs. 40.0%, P<0.01) were decreased significantly, while the methylation level of IGF2-H19 imprinting control region (ICR) was increased significantly (68.8% vs. 84.0%, P<0.01) in CFC group. Somatic cell nuclear transfer resulted in aberrant methylation of IGF2-H19 locus by influencing Dnmts and Tets genes expression, which could affect genomic imprinting, and then influence the efficiency of serial recloning.

CLC Number:

S827.2

LIU Zi-fei, DENG Ming-tian, REN Cai-fang, WAN Yong-jie, WANG Feng. IGF2-H19 Locus Methylation Status in Cloned Goat Fibroblast Cells[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2017, 48(12): 2277-2285.

0
/ / Recommend

Add to citation manager EndNote|Reference Manager|ProCite|BibTeX|RefWorks

URL: https://www.xmsyxb.com/EN/10.11843/j.issn.0366-6964.2017.12.007

https://www.xmsyxb.com/EN/Y2017/V48/I12/2277

References

[1] GE H T, CUI C, LIU J, et al. The growth and reproduction performance of TALEN-mediated β-lactoglobulin-knockout bucks[J]. Transgenic Res, 2016, 25(5):721-729.
[2] KUROME M, HISATOMI H, MATSUMOTO S, et al. Production efficiency and telomere length of the cloned pigs following serial somatic cell nuclear transfer[J]. J Reprod Dev, 2008, 54(4):254-258.
[3] YIN X J, LEE H S, YU X F, et al. Production of second-generation cloned cats by somatic cell nuclear transfer[J]. Theriogenology, 2008, 69(8):1001-1006.
[4] SAINI M, SELOKAR N L, AGRAWAL H, et al. Treatment of donor cells and reconstructed embryos with a combination of trichostatin-A and 5-aza-2'-deoxycytidine improves the developmental competence and quality of buffalo embryos produced by handmade cloning and alters their epigenetic status and gene expression[J]. Cell Reprogram, 2017, 19(3):208-215.
[5] NIE X W, LIU Q, WANG R G, et al. DNA demethylation pattern of in-vitro fertilized and cloned porcine pronuclear stage embryos[J]. Clin Chim Acta, 2017, 473:45-50.
[6] ZHANG J, QU P, ZHOU C, et al. microRNA-125b is a key epigenetic regulatory factor that promotes nuclear transfer reprogramming[J]. J Biol Chem, 2017, doi:10.1074/jbc.M117.796771.
[7] SMITH Z D, MEISSNER A. DNA methylation:roles in mammalian development[J]. Nat Rev Genet, 2013, 14(3):204-220.
[8] ZHANG X Y, HU M, LYU X, et al. DNA methylation regulated gene expression in organ fibrosis[J]. Biochim Biophys Acta, 2017, 1863(9):2389-2397.
[9] EDWARDS J R, YARYCHKIVSKA O, BOULARD M, et al. DNA methylation and DNA methyltransferases[J]. Epigenetics Chromatin, 2017, 10:23.
[10] KAUT O, SHARMA A, SCHMITT I, et al. DNA methylation of imprinted loci of autosomal chromosomes and IGF2 is not affected in Parkinson's disease patients' peripheral blood mononuclear cells[J]. Neurol Res, 2017, 39(3):281-284.
[11] BARROCA V, LEWANDOWSKI D, JARACZ-ROS A, et al. Paternal insulin-like growth factor 2 (Igf2) regulates stem cell activity during adulthood[J]. EBioMedicine, 2017, 15:150-162.
[12] 孙晓丽, 李树峰, 佟慧丽, 等. 不同肌肉特异性启动子IGF2表达载体构建及对牛骨骼肌卫星细胞增殖的影响[J]. 畜牧兽医学报, 2015, 46(4):555-560.
SUN X L, LI S F, TONG H L, et al. The construction of different muscle-specific promoter IGF2 expression vector and its impact on bovine skeletal muscle satellite cell proliferation[J]. Acta Veterinaria et Zootechnica Sinica, 2015, 46(4):555-560. (in Chinese)
[13] 刘艳利, 申静, 支丽慧, 等. 叶酸调控鸡脾和胸腺IGF2表达的表观遗传机制探究[J]. 畜牧兽医学报, 2016, 47(2):296-304.
LIU Y L, SHEN J, ZHI L H, et al. The study on epigenetic mechanism of IGF2 expression in spleen and thymus regulated by folic acid in broilers[J]. Acta Veterinaria et Zootechnica Sinica, 2016, 47(2):296-304. (in Chinese)
[14] PAULSEN M, TAKADA S, YOUNGSON N A, et al. Comparative sequence analysis of the imprinted Dlk1-Gtl2 locus in three mammalian species reveals highly conserved genomic elements and refines comparison with the Igf2-H19 region[J]. Genome Res, 2001, 11(12):2085-2094.
[15] WU K F, LIANG W C, FENG L, et al. H19 mediates methotrexate resistance in colorectal cancer through activating Wnt/β-catenin pathway[J]. Exp Cell Res, 2017, 350(2):312-317.
[16] LI X P, HAO C L, WANG Q, et al. H19 gene methylation status is associated with male infertility[J]. Exp Ther Med, 2016, 12(1):451-456.
[17] YANG W W, NING N, JIN X M. The lncRNA H19 promotes cell proliferation by competitively binding to miR-200a and derepressing β-catenin expression in colorectal cancer[J]. Biomed Res Int, 2017, 2017:2767484.
[18] GIABICANI E, BRIOUDE F, LE BOUC Y, et al. Imprinted disorders and growth[J]. Ann Endocrinol (Paris), 2017, 78(2):112-113.
[19] BARROW T M, BARAULT L, ELLSWORTH R E, et al. Aberrant methylation of imprinted genes is associated with negative hormone receptor status in invasive breast cancer[J]. Int J Cancer, 2015, 137(3):537-547.
[20] LEWIS A, LEE J Y, DONALDSON A V, et al. Increased expression of H19/miR-675 is associated with a low fat-free mass index in patients with COPD[J]. J Cachexia Sarcopenia Muscle, 2016, 7(3):330-344.
[21] ZHOU J D, LIN J, ZHANG T J, et al. Hypomethylation-mediated H19 overexpression increases the risk of disease evolution through the association with BCR-ABL transcript in chronic myeloid leukemia[J]. J Cell Physiol, 2017, doi:10.1002/jcp.26119.
[22] MENG L, WAN Y J, SUN Y Y, et al. Generation of five human lactoferrin transgenic cloned goats using fibroblast cells and their methylation status of putative differential methylation regions of IGF2R and H19 imprinted genes[J]. PLoS One, 2013, 8(10):e77798.
[23] LUCIFERO D, SUZUKI J, BORDIGNON V, et al. Bovine SNRPN methylation imprint in oocytes and day 17in vitro-produced and somatic cell nuclear transfer embryos[J]. Biol Reprod, 2006, 75(4):531-538.
[24] CONSTÂNCIA M, DEAN W, LOPES S, et al. Deletion of a silencer element in Igf2 results in loss of imprinting independent of H19[J]. Nat Genet, 2000, 26(2):203-206.
[25] MURATA A, BABA Y, WATANABE M, et al. IGF2 DMR0 methylation, loss of imprinting, and patient prognosis in esophageal squamous cell carcinoma[J]. Ann Surg Oncol, 2014, 21(4):1166-1174.
[26] MURRELL A, HEESON S, BOWDEN L, et al. An intragenic methylated region in the imprinted Igf2 gene augments transcription[J]. EMBO Rep, 2001, 2(12):1101-1106.
[27] IDERAABDULLAH F Y, VIGNEAU S, BARTOLOMEI M S. Genomic imprinting mechanisms in mammals[J]. Mutat Res, 2008, 647(1-2):77-85.
[28] WOLFFE A P. Transcriptional control:imprinting insulation[J]. Curr Biol, 2000, 10(12):R463-R465.
[29] HUMPHERYS D, EGGAN K, AKUTSU H, et al. Epigenetic instability in ES cells and cloned mice[J]. Science, 2001, 293(5527):95-97.
[30] SU J M, YANG B, WANG Y S, et al. Expression and methylation status of imprinted genes in placentas of deceased and live cloned transgenic calves[J]. Theriogenology, 2011, 75(7):1346-1359.
[31] WU X, ZHANG Y. TET-mediated active DNA demethylation:mechanism, function and beyond[J]. Nat Rev Genet, 2017, 18(9):517-534.
[32] PIYASENA C, CARTIER J, PROVENCAL N, et al. Dynamic changes in DNA methylation occur during the first year of life in preterm infants[J]. Front Endocrinol (Lausanne), 2016, 7:158.
[33] JACKSON-GRUSBY L, BEARD C, POSSEMATO R, et al. Loss of genomic methylation causes p53-dependent apoptosis and epigenetic deregulation[J]. Nat Genet, 2001, 27(1):31-39.
[34] LIU L Z, MAO S Q, RAY C, et al. Differential regulation of genomic imprinting by TET proteins in embryonic stem cells[J]. Stem Cell Res, 2015, 15(2):435-443.
[35] BERMEJO-ÁLVAREZ P, RAMOS-IBEAS P, PARK K E, et al. Tet-mediated imprinting erasure in H19 locus following reprogramming of spermatogonial stem cells to induced pluripotent stem cells[J]. Sci Rep, 2015, 5:13691.
[36] WAKAYAMA S, KOHDA T, OBOKATA H, et al. Successful serial recloning in the mouse over multiple generations[J]. Cell Stem Cell, 2013, 12(3):293-297.

IGF2-H19 Locus Methylation Status in Cloned Goat Fibroblast Cells

PDF (PC)

Knowledge

Abstract

Cite this article

share this article

References

Related Articles 15

Recommended Articles

Metrics

Comments

[1]	YAN Xiaochun, XI Haijiao, LI Jinquan, WANG Zhiying, SU Rui. Study on Estimates of Genomic Breeding Value of Fleece Traits in Inner Mongolia Cashmere Goats [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(1): 120-128.
[2]	ZHANG Chenjian, LI Yinxia, DING Qiang, LIU Weijia, WANG Huili, HE Nan, WU Jiashun, CAO Shaoxian. Efficient Preparation of CRISPR/Cas9-mediated Goat SOCS2 Gene Edited Embryos [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(1): 129-141.
[3]	MENG Qiuchi, LU Guangyu, CHEN Dingshuang, LIN Yaqiu, WANG Ruilong, ZHONG Chaosong, WANG Yong, LIU Wei, WANG Youli, LI Yanyan, LI Zhixiong. Goats GPR35 Gene Expression Characteristics Analysis and the Effect on Subcutaneous Fat Cell Differentiation [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(12): 4993-5007.
[4]	HE Zhicheng, QIN Xiaochen, Lü Yongqiang, LI Xiujin, BIAN Huilong, LUO Jun, LI Cong. Multiple Stepwise Regression Analysis and Growth Curve Fitting of Body Measurement Traits in Saanen Dairy Goats [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(12): 5301-5311.
[5]	ZHANG Renbao, ZHOU Donghui, ZHOU Lisheng, GAO Xiaoxiao, LIU Nan, HE Jianning. Analysis of Genetic Structure of Conservation Population in Jining Gray Goats Based on 70 K SNP Chip [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(7): 2836-2847.
[6]	SHAO Yuexin, ZHANG Xinyu, GE Liyan, SHI Huaiping. Cloning and RNA Interference Analysis of ATF4 Gene in Xinong Saanen Dairy Goat [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(6): 2353-2364.
[7]	XU Tiantian, ZHANG Tongtong, WANG Meng, WANG Xin. Transcription Factor Foxq1 Affects the Proliferation of Hair Follicle Stem Cells in Cashmere Goats via WNT/β-catenin Signaling Pathway [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(6): 2653-2661.
[8]	QUBI Wuqie, LI Yanyan, LI Xin, WANG Yong, WANG Youli, LIU Wei, ZHU Jiangjiang, LIN Yaqiu. APOA4 Gene Inhibits Intramuscular Adipocyte Differentiation in Goat [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(5): 1927-1938.
[9]	MA Keyan, HAN Jintao, BAI Yaqin, LI Taotao, MA Youji. Genetic Diversity Analysis of Yongdeng Qishan Sheep Based on Specific-Locus Amplified Fragment Sequencing [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(5): 1939-1950.
[10]	ZHU Junru, WEI Wei, PEI Dangshuai, ZHANG Fenque, DUAN Yu, GUO Yongfeng, JIANG Yue, XIA Shuli, HAN Jing, HOU Jinxing, AN Xiaopeng. Effects of PDCD4 on the Apoptosis of Dairy Goat Mammary Epithelial Cells and the Synthesis of β-casein and TG [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(4): 1429-1440.
[11]	CHEN Cancan, JIANG Jing, SUN Xiaoyan, REN Hangxing, LI Jie. Expression of AGRP Gene in Goat Tissue and Its Action Mechanism on Melanin Production [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(4): 1441-1451.
[12]	YE Junning, DENG Ming, XUE Huiwen, LIU Guangbin, ZOU Xian, SUN Baoli, GUO Yongqing, LIU Dewu, LI Yaokun. Identification and Analysis of mRNA and lncRNA Affecting Goat Fetal Muscle Development [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(3): 989-1002.
[13]	LI Xiaobo, LIU Zhanfa, LIU Yue, CHEN Qian, MA Yuehui, ZHAO Qianjun, YE Shaohui. Mining Genes Related to Wool Bending of Zhongwei Goat Based on WGCNA and GSEA [J]. Acta Veterinaria et Zootechnica Sinica, 2022, 53(9): 2930-2943.
[14]	SUN Hao, ZHE Xiaoshu, ZHANG Wenqi, HAO Fei, LI Wennan, LIU Jie, LIU Dongjun. Effects of Chaetocin on Histone Methylation Modification of Cashmere Goats ADSCs [J]. Acta Veterinaria et Zootechnica Sinica, 2022, 53(7): 2380-2389.
[15]	LIU Kehan, WANG Yong, LI Yanyan, WANG Chongyang, ZHU Jiangjiang, XIONG Yan, LI An, LIN Yaqiu. Effects of SRSF10 on the Differentiation of Intramuscular Preadipocytes in Goats [J]. Acta Veterinaria et Zootechnica Sinica, 2022, 53(6): 1768-1778.