中国激光, 2014, 41 (11): 1115002, 网络出版: 2014-10-08   

拉曼光谱技术研究铝胁迫下的土生隐球酵母细胞凋亡

Cell Apoptosis in Yeast under Aluminum Stress Analyzed by Laser Raman Spectroscopy
作者单位
1 昆明理工大学生物工程技术研究中心, 云南 昆明 650500
2 广西科学院生物物理实验室, 广西 南宁 530007
摘要
运用激光镊子拉曼光谱技术(LTRS)研究铝胁迫下土生隐球酵母细胞和线粒体拉曼光谱变化。细胞和线粒体中与核酸、蛋白质和脂类相关的特征峰强度随着铝处理浓度和处理时间的增加而逐渐降低,表明细胞凋亡过程中细胞和线粒体内的核酸、蛋白质和脂类生物大分子逐渐减少。线粒体内与细胞色素c(Cyt c)相关的特征峰强度随铝处理浓度增大和处理时间的延长而显著降低,说明线粒体中的细胞色素c释放到线粒体外。线粒体的呼吸峰随着铝浓度的增加和处理时间的延长而降低,说明线粒体的活性不断减弱,能量代谢受阻。因此,拉曼光谱实时监测了铝胁迫细胞凋亡的动态过程及其线粒体的生理生化变化过程,并且揭示了铝诱导土生隐球酵母凋亡过程中线粒体内细胞色素c的释放行为,这些结果有助于了解酸性土壤中铝对生物的毒害机理。
Abstract
Laser Raman spectroscopy (LTRS) is used to monitor the Raman spectroscopic changes of Cryptococcus humicolus cells and mitochondria stressed by aluminum (Al). The results show that the characteristic Raman peaks of C. humicolus cells and mitochondria are significantly decreased with the increase of aluminum concentration and treatment time, the intensity of Raman peaks associated to nucleic acid, protein and lipid respectively exhibits significant decreasing tendency with the extension of treatment time, indicating that the content of nucleic acid, protein and lipid in the cells and mitochondria is reduced during the apoptosis process. The intensity of characteristic peaks related to cytochrome c (Cyt c) in mitochondria significantly decreases with the increase of aluminum concentration and treatment time, suggesting that the yeast mitochondrial membrane rupture leads to Cyt c release into outer of mitochondria. With the increase of aluminum concentration and treatment time, the respiratory peak is observed to decrease gradually, indicating that the mitochondrial activity becomes weaker, energy metabolism has been hampered. LTRS is used for real-time monitoring of the dynamic process of cell apoptosis and physiological and biochemical changes of in vitro mitochondria under aluminum stress, and revealing the release behavior of Cyt c during the apoptosis process of C. humicolus cells induced by aluminum. The results help understand the biological mechanism of aluminum toxicity in acidic soils on organisms.
参考文献

[1] Pina R G, Cervantes C. Microbial interactions with aluminium[J]. Biometals, 1996, 9(3): 311-316.

[2] Liu X, Kim C N, Yang J, et al.. Induction of apoptotic program in cell-free extracts: Requirement for dATP and cytochrome c[J]. Cell, 1996, 86(1): 147-157.

[3] Verhagen A M, Ekert P G, Pakusch M, et al.. Identification of DIABLO, a mammalian protein that promotes apoptosis by binding to and antagonizing IAP proteins[J]. Cell, 2000, 102(1): 43-53.

[4] Lorenzo H K, Susin S A, Penninger J, et al.. Apoptosis inducing factor (AIF): A phylogenetically old, caspase-independent effector of cell death[J]. Cell Death Differ, 1999, 6(6): 516-524.

[5] Panda S K, Yamamoto Y, Kondo H, et al.. Mitochondrial alterations related to programmed cell death in tobacco cells under aluminum stress[J]. C R Biol, 2008, 311(8): 597-610.

[6] Keith D R, Eric J S, Yogesh K S, et al.. Aluminum induces oxidative stress genes in Arabidopsis thaliana[J]. Plant Physiol, 1998, 116(1): 409-418.

[7] Yamamoto Y, Kobayashi Y, Rama D S, et al.. Aluminum toxicity is associated with mitochondrial dysfunction and the production of reactive oxygen species in plant cells[J]. Plant Physiol, 2002, 128(1): 63-72.

[8] Boscolo P R, Menossi M, Jorge R A. Aluminum induced oxidative stress in maize[J]. Phytochemistry, 2003, 62(2): 181-189.

[9] Yin L N, Mano J C, Wang S W, et al.. The involvement of lipid peroxide-derived aldehydes in aluminum toxicity of tobacco roots[J]. Plant Physiol, 2010, 152(3): 1406-1417.

[10] Li Z, Xing D. Mechanistic study of mitochondria-dependent programmed cell death induced by aluminum phytotoxicity using fluorescence techniques[J]. J Exp Bot, 2010, 62(1): 331-343.

[11] Nian H, Wang G, Chen L. Physiological and transcriptional analysis of the effects of aluminum stress on Cryptococcus humicola[J]. Microbiol Biotechnol, 2012, 28(6): 2319-2329.

[12] Nian H J, Wang G Q, Zhao L W, et al.. Isolation of Al-tolerant yeasts and identification of their Al-tolerance characteristics[J]. J Biol Res, 2012, 18: 227-234.

[13] 金建玲, 高东, 孙忠东. 一种制备酵母菌线粒体DNA的简便方法[J]. 遗传, 1996, 18(2): 46-48.

    Jin Jianling, Gao Dong, Sun Zhongdong. A simple procedure for preparation mtDNA in yeast[J]. Hereditas, 1996, 18(2): 46-48.

[14] Shetty G, Kendall C, Shepherd N, et al.. Raman spectroscopy: Elucidation of biochemical changes in carcinogenesis of oesophagus[J]. Brit J Cancer, 2006, 94(10): 1460-1464.

[15] Notingher I. Raman spectroscopy cell-based biosensors[J]. Sensors, 2007, 7(8): 1343-1358.

[16] 许以明. 拉曼光谱及其在结构生物学中的应用[M]. 北京: 化学工业出版社, 2005.

    Xu Yiming. Raman Spectroscopy and Its Application in Structural Biology[M]. Beijing: Chemical Industry Press, 2005.

[17] Chikao O, Hamaguchi H O. In vivo resonance Raman detection of ferrous cytochrome c from mitochondria of single living yeast cells[J]. Chem Lett, 2010, 39(3): 270-271.

[18] Tonshin A A, Saprunova V B, Solodovnikova I M. Functional activity and ultrastructure of mitochondrial isolated from myocardial apoptotic tissue[J]. Biochemistry (Moscow), 2003, 68(8): 875-881.

[19] Pully V V, Otto C. The intensity of the 1602 cm-1 band in human cells is related to mitochondrial activity[J]. J Raman Spectrosc, 2009, 40(5): 473-475.

[20] Hamada K, Fujita K, Smith N I, et al.. Raman microscopy for dynamic molecular imaging of living cells[J]. J Biomed Opt, 2008, 13(4): 044027.

[21] 李自达, 赖钧灼, 廖威, 等. 浓醪乙醇发酵的单细胞拉曼光谱表征[J]. 光学学报, 2012, 32(3): 0317001.

    Li Zida, Lai Junzhuo, Liao Wei, et al.. Raman spectroscopic profile of ethanol fermentation in high gravity cassava starch brewing[J]. Acta Optica Sinica, 2012, 32(3): 0317001.

[22] 叶宇煌, 陈阳, 李永增, 等. 基于拉曼光谱的鼻咽癌与正常鼻咽细胞株的分类研究[J]. 中国激光, 2012, 39(5): 0504003.

    Ye Yuhuang, Chen Yang, Li Yongzeng, et al.. Discrimination of nasopharyngeal carcinoma and normal nasopharyngeal cell lines based on confocal Raman microspectroscopy[J]. Chinese J Lasers, 2012, 39(5): 0504003.

[23] 刘书朋, 朱鸿飞, 陈娜, 等. 金颗粒为活性基底的裸鼠血清表面增强拉曼散射光谱分析[J]. 中国激光, 2012, 39(5): 0504004.

    Liu Shupeng, Zhu Hongfei, Chen Na, et al.. Surface enhanced Raman scattering spectrum analysis of nude mouse serum with Au nanoparticles active substrate[J]. Chinese J Lasers, 2012, 39(5): 0504004.

[24] 孙美娟, 蒋玉凌, 来爱华, 等. 激光镊子拉曼光谱技术分析圆红冬孢酵母生成油脂和类胡萝卜素[J]. 激光与光电子学进展, 2013, 50(3): 033001.

    Sun Meijuan, Jiang Yuling, Lai Aihua, et al.. Analysis of lipid and carotenoids in Rhodosporidium toruloides using laser tweezer Raman spectroscopy[J]. Laser & Optoelectronics Progress, 2013, 50(3): 033001.

[25] 牛丽媛, 林漫漫, 李雪, 等. 活体糖尿病小鼠中单个白细胞的拉曼光谱分析[J]. 激光与光电子学进展, 2012, 49(6): 063001.

    Niu Liyuan, Lin Manman, Li Xue, et al.. Raman spectroscopic analysis of single white blood cell of DM mouse in vivo[J]. Laser & Optoelectronics Progress, 2012, 49(6): 063001.

[26] Arends M J, Morris R G, Wyllie A H. Apoptosis. The role of the endonuclease[J]. Am J Pathol, 1990, 136(3): 593-608.

[27] Tang H, Yao H, Wang G, et al.. NIR Raman spectroscopic investigation of single mitochondria trapped by optical tweezers[J]. Opt Express, 2007, 15(20): 12708-12716.

[28] Chiu L D, Ando M, Hamaguchi H O. Study of the ‘Raman spectroscopic signature of life’ in mitochondria isolated from budding yeast[J]. J Raman Spectrosc, 2010, 41(1): 2-3.

[29] Kakita M, Kaliaperumal V, Hamaguchi H O. Resonance Raman quantification of the redox state of cytochromes b and c in-vivo and in-vitro[J]. J Biophoton, 2012, 5(1): 20-24.

[30] Jiang X, Wang X. Cytochrome c-mediated apoptosis [J]. Annu Rev Biochem, 2004, 73: 87-106.

李金金, 卢明倩, 张晶晶, 黄庶识, 陈丽梅, 年洪娟. 拉曼光谱技术研究铝胁迫下的土生隐球酵母细胞凋亡[J]. 中国激光, 2014, 41(11): 1115002. Li Jinjin, Lu Mingqian, Zhang Jingjing, Huang Shushi, Chen Limei, Nian Hongjuan. Cell Apoptosis in Yeast under Aluminum Stress Analyzed by Laser Raman Spectroscopy[J]. Chinese Journal of Lasers, 2014, 41(11): 1115002.

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