蛋白磷酸酶-2Ac在不同倍性鱼6种组织中的分化表达模式

DIFFERENTIAL EXPRESSION PATTERNS OF THE PROTEIN PHOSPHATASE-2Ac IN SIX TISSUES OF DIFFERENT PLOIDY FISH

蛋白磷酸酶-2A是最重要的丝氨酸/苏氨酸蛋白磷酸酶之一,对于调控多细胞的生命活动起着非常重要的作用。以异源四倍体鲫鲤及其二倍体父/母本(湘江野鲤/红鲫)和子代三倍体湘云鲫等为实验材料,运用Westernblot技术及荧光免疫组织化学技术等实验手段,得到了Protein Phosphatase-2A(PP2A)的催化亚基在上述不同倍性鱼体内6种不同组织的表达模式:Protein Phosphatase-2Ac(PP2Ac)在异源四倍体鲫鲤及其二倍体父/母本及子代三倍体湘云鲫不同组织中蛋白水平均有表达,而且出现了明显的种属特异性和组织特异性,如在大脑、肌肉、肝脏三组织中,三倍体湘云鲫中PP2Ac的表达相对最高。而在肾脏组织中,PP2Ac在异源四倍体鲫鲤中的表达水平最高,父本与三倍体湘云鲫中的表达比较相近,且最低;而在性腺组织中则是父本精巢中的表达最高;在心脏组织中,PP2Ac在母本红鲫中的表达相对较高。这种明显的种属之间组织特异性可能说明了子代与父母本之间的变异性。荧光免疫组化实验结果显示,从整体水平来看,4种不同鱼的同一组织中,PP2Ac的相对定位是非常相似的,这可能说明了异源四倍体鲫鲤与其二倍体父/母本及子代三倍体湘云鲫之间的遗传相似性。研究结果为进一步探索PP2Ac在脊椎动物不同组织中的功能提供了实验依据。

Protein Phosphatase-2A（PP2A） is one of the most important protein serine/threonine phosphatases and plays distinct roles in regulating gene expression,signal transduction,cell proliferation,differentiation,senescence,apoptosis,autophagy,morphogenesis and organogenesis.Our previous studies have established the expression patterns of the catalytic subunits and the regulatory subunits for PP-2A in mouse eye[1].To explore the possible functions of PP-2A in various tissues of the lower vertebrates,here,we have analyzed the differential expression pattern,functional status,and the cellular localization of the catalytic subunit for PP-2A using western blot and immunofluorescence cytochemistry on four different ploidy level fish.These include the allotetraploid hybrids and their diploid parents,common carp（♂） and red crucian carp（♀） as well as the triploid crucian carp derived from crossover between the allotetraploids and common carp or between the allotetraploids and the red crucian carp.Our study demonstrated the following results：1） In brain,muscle and liver from the four types of fish,the highest expression level of PP-2Ac was observed in the triploid crucian carp;2） In the kidney of the four types of fish,however,the highest expression level was detected in the allotetraploids,and the lowest level was detected in the diploid common carp and the triploid crucian carp;3） In the gonads of the four types of fish,the highest expression level of PP-2Ac was found in testis of the diploid common carp;4） In the heart of the four kinds of fish,the highest expression level of PP-2Ac was observed in the diploid red crucian carp;5） While both phosphorylated and non-phosphorylated PP-2Ac were found in the heart and muscle tissues from all four types of fish,only phosphorylated PP-2Ac was detected in the brain from these organisms;6） In the kidney,liver and gonads,non-phosphorylated PP-2Ac was only detected in the diploid common carp and red crucian carp;7） PP-2Ac was mainly localized in the cytoplasm of the neurons in the brain tissue,of the cardiomyocytes in the heart,and of the myocytes in the muscle of the four types of fish;certain nuclei from the neurons in both allotetraploid and triploid brain have strong immunofluorescence signal;8）PP-2Ac was found in the cytoplasm but not the nuclei of cuboidal cells of the renal collecting and filtration tubes of the kidney,and in the hepatocytes,sinusoids and the bile canaliculi of the liver of the four types of fish;and finally,PP-2Ac was found in the cytoplasm membrane and nuclei of the growing oocytes from the diploid red crucian carp and triploid crucian carp,in the base membranes covering the seminiferous tubules and the interstitial tissues separating the seminiferous tubules,sertoli cells and some spermatocytes of the diploid common carp.Together,these results lead to the following conclusions：1） PP-2Ac was differentially expressed in various tissues of the four different ploidy level fish;2） PP-2A functions are highly regulated in different tissues;3） Within the same tissue of the four types of fish,PP-2A was localized in the same types of cells;and 4） the presence of strong PP-2Ac immunofluorescence signal in certain nuclei of neurons in both allotetraploid and triploid brains but not in those of the diploid parents indicates that the PP-2Ac may be used as a biochemical marker to distinguish the different types of fish.Overall,our results provide important information for the further exploration of PP-2A functions in lower vertebrates.