Analysis of expression patterns of protein phosphatase-1 and phosphatase-2A in rat and bovine lenses.
Analysis of expression patterns of protein phosphatase-1 and phosphatase-2A in rat and bovine lenses.
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发表时间：
2001-10
影响因子：
4.4
通讯作者：
D. W. Li;H. Xiang;U. Fass;X. Y. Zhang
中科院分区：
文献类型：
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作者：
D. W. Li;H. Xiang;U. Fass;X. Y. Zhang
目的蛋白质上丝氨酸和苏氨酸残基的可逆磷酸化和去磷酸化在调节多种细胞活动中发挥着不同的作用。虽然蛋白丝氨酸-苏氨酸激酶已经在晶状体系统中得到了很好的研究，但对丝氨酸-苏氨酸磷酸酶的表达和功能知之甚少。本文报道了蛋白磷酸酶(PP)-1和-2A在成年大鼠和牛晶状体中的表达模式。方法分别从大鼠和牛晶状体上皮细胞和纤维细胞中提取总rna和蛋白。采用RT-PCR和Northern blot检测两种晶状体上皮细胞和不同部位纤维细胞mRNA的表达水平。Western blot检测不同样品的蛋白表达水平。酶促法检测了这些样品中PP-1和-2A的活性分布。结果PP-1催化亚基（PP-1cs）和PP-2A催化亚基（PP-2Acs） mrna在大鼠和牛晶状体上皮细胞和纤维细胞中均有表达。对这两种mrna在纤维细胞不同部位表达模式的详细检查显示，皮层纤维细胞（F1）在纤维细胞不同部位中含有最高水平的PP-1cs和-2Acs mrna（类似于上皮细胞中的mrna）。这两种mrna的水平在接下来的纤维细胞层（F2和F3）中依次下降，在纤维细胞内层（F4和N）中几乎检测不到。与mRNA的表达模式相反，PP-1cs蛋白主要存在于上皮细胞中。在不同的纤维细胞层中，只有皮质（F1）纤维细胞含有可检测水平的PP-1cs蛋白（牛晶状体在该区域的PP-1cs含量相对高于大鼠晶状体）。在剩余的纤维细胞中，PP-1cs蛋白在大鼠晶状体中几乎检测不到，在牛晶状体中略有检测到。PP-2Acs蛋白仅在晶状体上皮细胞中检测到。酶促分析表明，PP-1和-2A蛋白的活性分布模式与pp -1c和-2Acs相似。此外，在晶状体上皮细胞中，PP-1的活性大约是PP-2A活性的4到5倍。结论本研究表明活性的PP-1和-2A主要分布在晶状体上皮细胞中，其中PP-1是主要的磷酸酶。PP-1cs和-2Acs的mrna和蛋白在大鼠和牛晶状体上皮细胞和纤维细胞中有差异表达。
PURPOSE The reversible phosphorylation and dephosphorylation at the serine and threonine residues on proteins play distinct roles in regulating multiple cellular activities. Whereas the protein serine-threonine kinases have been well studied in the lens system, very little is known about the expression and function of the serine-threonine phosphatases. The present article reports the expression patterns of protein phosphatase (PP)-1 and -2A in adult rat and bovine lenses. METHODS Total RNAs and proteins were extracted from the epithelial and fiber cells of rat and bovine lenses. RT-PCR and Northern blot analysis were used to detect the mRNA expression levels in the epithelial cells and different fractions of fiber cells of these two types of lenses. Western blot was used to examine the protein expression levels in these different samples. An enzymatic assay was used to detect the activity distribution of PP-1 and -2A in these samples. RESULTS The mRNAs for the PP-1 catalytic subunit (PP-1cs) and PP-2A catalytic subunit (PP-2Acs) were expressed in both epithelial and fiber cells of rat and bovine lenses. A detailed examination of the expression patterns of the two mRNAs in different fractions of fiber cells revealed that the cortical fiber cells (F1) contain the highest level of PP-1cs and -2Acs mRNAs (similar to those in the epithelial cells) among different fractions of fiber cells. The levels of the two mRNAs were sequentially decreased in the next layers of fiber cells (F2 and F3) and became barely detectable in the inner layers of fiber cells (F4 and N). In contrast to the mRNA expression patterns, the PP-1cs protein was mainly found in the epithelial cells. Among different layers of fiber cells, only cortical (F1) fiber cells contained detectable level of PP-1cs protein (bovine lenses contained a relatively higher level of PP-1cs than rat lenses in this region). In the remaining fiber cells, the PP-1cs protein was hardly detectable in rat lenses and slightly detectable in bovine lenses. The PP-2Acs protein was detectable only in the lens epithelial cells. Enzymatic assays revealed that the distribution patterns of PP-1 and -2A activities were similar to those of PP-1cs and -2Acs proteins. Furthermore, PP-1 activity was approximately four to five times higher than PP-2A activity in the lens epithelial cells. CONCLUSIONS This study demonstrates that active PP-1 and -2A are mainly distributed in the lens epithelial cells, with PP-1 as a major phosphatase. The mRNAs and proteins for PP-1cs and -2Acs are differentially expressed in the epithelial and fiber cells of rat and bovine lenses.