A. I. Virtanen Institute for Molecular Sciences, Biocenter Kuopio, University of Eastern Finland, Yliopistonranta 1E, 70210, Kuopio, Finland.
Amino Acids. 2012 Feb;42(2-3):485-93. doi: 10.1007/s00726-011-1027-7. Epub 2011 Aug 2.
The polyamines, spermidine and spermine, are abundant organic cations participating in many important cellular processes. We have previously shown that the rate-limiting enzyme of polyamine catabolism, spermidine/spermine N(1)-acetyltransferase (SSAT), has an alternative mRNA splice variant (SSATX) which undergoes degradation via nonsense-mediated mRNA decay (NMD) pathway, and that the intracellular polyamine level regulates the ratio of the SSATX and SSAT splice variants. The aim of this study was to investigate the effect of SSATX level manipulation on SSAT activity in cell culture, and to examine the in vivo expression levels of SSATX and SSAT mRNA. Silencing SSATX expression with small interfering RNA led to increased SSAT activity. Furthermore, transfection of SSAT-deficient cells with mutated SSAT gene (which produced only trace amount of SSATX) yielded higher SSAT activity than transfection with natural SSAT gene (which produced both SSAT and SSATX). Blocking NMD in vivo by protein synthesis inhibitor cycloheximide resulted in accumulation of SSATX mRNA, and like in cell culture, the increase of SSATX mRNA was prevented by administration of polyamine analog N(1),N(11)-diethylnorspermine. Although SSATX/total SSAT mRNA ratio did not correlate with polyamine levels or SSAT activity between different tissues, increasing polyamine levels in a given tissue led to decreased SSATX/total SSAT mRNA ratio and vice versa. Taken together, the regulated unproductive splicing and translation of SSAT has a physiological relevance in modulating SSAT activity. However, in addition to polyamine level there seems to be additional factors regulating tissue-specific alternative splicing of SSAT.
多胺，包括亚精胺和精胺，是参与多种重要细胞过程的丰富有机阳离子。我们之前已经表明，多胺分解代谢的限速酶，亚精胺/精胺 N(1)-乙酰基转移酶（SSAT），具有替代的 mRNA 剪接变体（SSATX），其通过无意义介导的 mRNA 降解（NMD）途径进行降解，并且细胞内多胺水平调节 SSATX 和 SSAT 剪接变体的比例。本研究的目的是研究 SSATX 水平操纵对细胞培养中 SSAT 活性的影响，并检查 SSATX 和 SSAT mRNA 的体内表达水平。用小干扰 RNA 沉默 SSATX 表达导致 SSAT 活性增加。此外，用突变的 SSAT 基因（仅产生痕量 SSATX）转染 SSAT 缺陷细胞比用天然 SSAT 基因（产生 SSAT 和 SSATX）转染产生更高的 SSAT 活性。体内通过蛋白质合成抑制剂环己酰亚胺阻断 NMD 导致 SSATX mRNA 的积累，并且与细胞培养一样，多胺类似物 N(1)，N(11)-二乙基-norspermine 的给药阻止了 SSATX mRNA 的增加。尽管 SSATX/总 SSAT mRNA 比值与不同组织中的多胺水平或 SSAT 活性之间没有相关性，但在给定组织中增加多胺水平会导致 SSATX/总 SSAT mRNA 比值降低，反之亦然。总之，SSAT 的调节性无产物剪接和翻译在调节 SSAT 活性方面具有生理相关性。然而，除了多胺水平之外，似乎还有其他因素调节 SSAT 的组织特异性选择性剪接。
