| 533 | 1 | 80 |
| 下载次数 | 被引频次 | 阅读次数 |
为了研究赖氨酸特异性去甲基化酶4aa(lysine demethylase 4aa, kdm4aa)在斑马鱼(Danio rerio)低温耐受能力中的作用，本研究通过实时定量PCR检测kdm4aa-/-斑马鱼中kdm4其他家族成员的表达水平和低温处理后野生型(wild type, WT)斑马鱼中kdm4aa基因的表达水平；通过斑马鱼行为轨迹跟踪系统检测低温下kdm4aa-/-幼鱼的运动能力；采用逐级降温方法检测kdm4aa-/-斑马鱼低温耐受能力；通过DCFH-DA荧光探针、 ATP检测试剂盒、 Western blot检测低温下kdm4aa-/-斑马鱼中活性氧(reactive oxygen species, ROS)、腺苷三磷酸(adenosine triphosphate, ATP)、磷酸化组蛋白H2A.X(gamma histone H2A.X,γH2A.X)和微管相关蛋白1轻链3(microtubule-associated protein 1 light chain 3, LC3)水平。实时定量PCR结果显示，kdm4aa-/-斑马鱼中kdm4ab、kdm4b基因的mRNA水平补偿性增加，kdm4c基因的mRNA水平与野生型斑马鱼的相比无显著性差异。斑马鱼行为轨迹跟踪系统检测发现，在18℃低温下，kdm4aa基因失活后斑马鱼幼鱼的移动距离显著增加。8℃低温下，野生型斑马鱼中kdm4aa基因的mRNA水平显著升高。与野生型斑马鱼相比，低温处理后kdm4aa-/-斑马鱼的半数致死时间LT50显著降低(P<0.000 1); ROS水平显著升高(P<0.000 1); ATP含量显著降低(P<0.01); γH2A.X蛋白水平上升；LC3蛋白水平下降。结果表明：在低温环境下，kdm4aa-/-斑马鱼的ROS积累导致ATP合成能力降低和DNA损伤增加，同时kdm4aa-/-斑马鱼自噬能力降低，从而使斑马鱼的低温耐受能力降低。
Abstract：In order to study the function of kdm4aa on low temperature tolerance of zebrafish, the levels of kdm4 families in kdm4aa-/- zebrafish and the level of kdm4aa in wild type(WT) zebrafish treated by low temperature were detected by real time quantitative PCR. The locomotor ability of kdm4aa-/- juvenile fish at low temperature was detected by the behavior trajectory tracking system of zebrafish. The low temperature tolerance of kdm4aa-/- zebrafish was measured by a stepwise cooling method. Reactive oxygen species(ROS), adenosine triphosphate(ATP), gamma histone H2A.X(γH2A.X), and microtubule-associated protein 1 light chain 3(LC3) levels in kdm4aa-/- zebrafish were determined by DCFH-DA fluorescent probe, ATP detection kit and western blot, respectively. Real time quantitative PCR results showed that compared with WT zebrafish, the mRNA levels of kdm4ab and kdm4b in kdm4aa-/- zebrafish were significantly increased, while no significantly different of kdm4c was observed. The detection of zebrafish behavior trajectory tracking system showed that the movement distance of juvenile zebrafish increased significantly after kdm4aa knockout at a low temperature of 18 ℃. At a low temperature of 8 ℃ mRNA level of kdm4aa in wild type zebrafish was significantly increased. Compared to WT zebrafish, the median lethal time LT50 of kdm4aa-/- zebrafish was significantly decreased after low temperature treatment(P<0.000 1); ROS level was significantly increased(P<0.000 1); ATP content was significantly decreased(P<0.01); The level of γH2A.X protein increased; LC3 protein levels decreased. These results showed that ROS accumulation of kdm4aa-/- led to decreased ATP synthesis and increased DNA damage, and the autophagy ability of kdm4aa-/- zebrafish was decreased, thus reducing the low temperature tolerance of zebrafish.
季涛，燕晓霞，韩兵社，等，2021.KDM4A的结构、 功能及其在肿瘤中作用的研究进展.生命科学，33(6):742-751.[JI T,YAN X X,HAN B S,et al.,2021.Advances in the structure and functions of KDM4A and its roles in cancer.Chinese Bulletin of Life Sciences,33(6):742-751.]
BASNET R,ZIZIOLI D,TAWEEDET S,et al.,2019.Zebrafish larvae as a behavioral model in neuropharmacology.Biomedicines,7(1):23.
BRUSTEIN E,SAINT-AMANT L,BUSS R R,et al.,2003.Steps during the development of the zebrafish locomotor network.J.Physiol.Paris,97(1):77-86.
CARDAMONE M D,TANASA B,CHAN M,et al.,2014.GPS2/KDM4A pioneering activity regulates promoter-specific recruitment of PPARγ.Cell Rep.,8(1):163-176.
CHANG C C,YEH M S,CHENG W,2009.Cold shock-induced norepinephrine triggers apoptosis of haemocytes via caspase-3 in the white shrimp,Litopenaeus vannamei.Fish Shellfish.Immunol.,27(6):695-700.
CHENG C H,YE C X,GUO Z X,et al.,2017.Immune and physiological responses of pufferfish (Takifugu obscurus) under cold stress.Fish Shellfish.Immunol.,64:137-145.
COLMENARES S U,SWENSON J M,LANGLEY S A,et al.,2017.Drosophila histone demethylase KDM4A has enzyma-tic and non-enzymatic roles in controlling heterochromatin integrity.Dev.Cell,42(2):156-169.
COUTURE J F,COLLAZO E,ORTIZ-TELLO P A,et al.,2007.Specificity and mechanism of JMJD2A,a trimethyllysine-specific histone demethylase.Nat.Struct.Mol.Biol.,14(8):689-695.
DELLA TORRE C,LIBERATORI G,GHILARDI A,et al.,2022.The zebrafish (Danio rerio) embryo-larval contact assay combined with biochemical biomarkers and swimming performance in sewage sludge and hydrochar hazard assessment.Environ.Pollut.,302:119053.
DRAPEAU P,SAINT-AMANT L,BUSS R R,et al.,2002.Development of the locomotor network in zebrafish.Prog.Neurobiol.,68(2):85-111.
GAO Q,2019.Oxidative stress and autophagy.Autophagy:Bio-logy and Diseases.Singapore:Springer Singapore:179-198.
HAN L J,LUO J T,QU S Y,et al.,2023.kdm4aa is required for reproduction and development of zebrafish.Aquac.Fish.,8(6):713-719.
HUANG Y,FANG J A,BEDFORD M T,et al.,2006.Recognition of histone H3 lysine-4 methylation by the double Tudor domain of JMJD2A.Science,312(5774):748-751.
KONDURI P C,WANG T Y,SALAMAT N,et al.,2020.Heme,a metabolic sensor,directly regulates the activity of the KDM4 histone demethylase family and their interactions with partner proteins.Cells,9(3):773.
LAPIDOT M,CASE A E,WEISBERG E,et al.,2021.Essential role of the histone lysine demethylase KDM4A in the biology of malignant pleural mesothelioma (MPM).Br.J.Cancer,125(4):582-592.
LI Y H,WEI L,CAO J R,et al.,2016.Oxidative stress,DNA damage and antioxidant enzyme activities in the Pacific white shrimp (Litopenaeus vannamei) when exposed to hypoxia and reoxygenation.Chemosphere,144:234-240.
LIANG L Q,CHANG Y M,HE X L,et al.,2015.Transcriptome analysis to identify cold-responsive genes in Amur carp (Cyprinus carpio haematopterus).PLoS ONE,10(6):e0130526.
LIANG Q J,DONG W N,OU M F,et al.,2021.miR-151 affects low-temperature tolerance of Penaeus vannamei by modulating autophagy under low-temperature stress.Front.Cell Dev.Biol.,9:595108.
MUKHOPADHYAY S,PRAHARAJ P P,NAIK P P,et al.,2020.Identification of Annexin A2 as a key mTOR target to induce roller coaster pattern of autophagy fluctuation in stress.Biochim.Biophys.Acta Mol.Basis Dis.,1866(12):165952.
SHAHJAHAN M,KITAHASHI T,ANDO H,2017.Temperature affects sexual maturation through the control of kisspeptin,kisspeptin receptor,GnRH and GTH subunit gene expression in the grass puffer during the spawning season.Gen.Comp.Endocrinol.,243:138-145.
SUN S J,WANG B J,JIANG K Y,et al.,2015.Target of rapamycin (TOR) in Fenneropenaeus chinensis:cDNA cloning,characterization,tissue expression and response to amino acids.Aquac.Nutr.,21(1):1-9.
WALTERS Z S,ALADOWICZ E,VILLAREJO-BALCELLS B,et al.,2021.Role for the histone demethylase KDM4B in rhabdomyosarcoma via CDK6 and CCNA2:Compensation by KDM4A and apoptotic response of targeting both KDM4B and KDM4A.Cancers,13(7):1734.
WANG L Y,HUNG C L,CHEN Y R,et al.,2016.KDM4A coactivates E2F1 to regulate the PDK-dependent metabolic switch between mitochondrial oxidation and glycolysis.Cell Rep.,16(11):3016-3027.
WEI S F,LIU X Y,FARIDUL HASAN K M,et al.,2022.Extraction and purification of flavonoids from Buddleja officinalis maxim and their attenuation of H2O2-induced cell injury by modulating oxidative stress and autophagy.Molecules,27(24):8985.
XIA Y,YAO R Q,ZHAO P Y,et al.,2022,Publication trends of research on COVID-19 and host immune response:a bibliometric analysis.Front.Public Health,10:939053.
XING X G,JIANG Z S,TANG X,et al.,2016.Sodium butyrate protects against oxidative stress in HepG2 cells through modulating Nrf2 pathway and mitochondrial function.J.Physiol.Biochem.,73(3):405-414.
XU Z H,REGENSTEIN J M,XIE D D,et al.,2018.The oxidative stress and antioxidant responses of Litopenaeus vannamei to low temperature and air exposure.Fish Shellfish.Immunol.,72:564-571.
YILMAZ S,ERGüN S,?ELIK E S,et al.,2021.The impact of acute cold water stress on blood parameters,mortality rate and stress-related genes in Oreochromis niloticus,Oreochromis mossambicus and their hybrids.J.Therm.Biol.,100:103049.
YOUNG N L,DERE R,2021.Mechanistic insights into KDM4A driven genomic instability.Biochem.Soc.Trans.,49(1):93-105.
ZHOU H,ZHUANG Z X,SUN Y Q,et al.,2019.Changes in DNA methylation during epigenetic-associated sex reversal under low temperature in Takifugu rubripes.PLoS ONE,14(8):e0221641.
ZHUANG Y L,MA Q Y,GUO Y,et al.,2017.Protective effects of rambutan (Nephelium lappaceum) peel phenolics on H2O2-induced oxidative damages in HepG2 cells and d-galactose-induced aging mice.Food Chem.Toxicol.,108:554-562.
基本信息:
DOI：10.13417/j.gab.043.000091
中图分类号:Q953
引用信息:
[1]张瑶,罗军涛,张梦冉,等.斑马鱼中kdm4aa基因对低温耐受能力的影响[J].基因组学与应用生物学,2024,43(01):91-99.DOI:10.13417/j.gab.043.000091.
基金信息:
上海市科技兴农重点攻关项目[沪农科创字(2019)第1-2号]资助
2023-10-18
2023-10-18
2023-10-18