真核細胞的轉錄作用是高度被調控的過程。在組織蛋白的N端會受到乙醯化作用修飾，使染色質結構鬆散，進而產生基因的轉錄作用。先前我們實驗室已經發現在子宮頸上皮癌細胞中，經由上皮生長因子的處理下，會使c-Jun的表現增加，而此增加的情形對其誘導十二位酯氧酵素基因表現是必須且重要的。在本實驗中，主要是想看組織蛋白去乙醯化酶之抑制劑，trichostatin A (TSA) 及sodium butyrate (NaBT)，對上皮生長因子誘導之十二位酯氧酵素基因活化有何影響。首先我們確認在TSA處理一小時的細胞中，組織蛋白乙醯化的程度有增加的情形。進而發現在TSA及NaBT的處理下，隨著劑量的增加，會對上皮生長因子誘導的十二位酯氧酵素的活性有抑制的作用。在A431細胞中，EGF的處理下主要活化Ras-ERK及Ras-JNK此兩條訊息傳遞路徑，進而誘導十二脂氧酵素基因表現。實驗結果得知，TSA及NaBT並不會影響EGF對ERK及JNK的磷酸化。我們由體外激酶活性分析法得知，抑制劑TSA及NaBT會減少了上皮生長因子活化JNK的活性，但並不會影響ERK活性。又因為JNK的受質為c-Jun，所以我們進一步釐清TSA及NaBT是否會影響c-Jun的表現。實驗後發現TSA及NaBT會抑制由上皮生長因子所誘導之c-Jun N端磷酸化。也由北方及西方點墨法發現，抑制劑也由於減少了c-Jun mRNA的表現，進而減少其蛋白質量。為了進一步釐清TSA及NaBT影響EGF所促進之c-Jun磷酸化情形，我們送入了一段表現c-Jun N端 (1-220) 的質體進入細胞，發現在表現同樣量的c-Jun N端蛋白質下，處理抑制劑對生長因子所誘導c-Jun Ser63及Ser73的磷酸化也呈現劑量相關性的抑制。因此我們知道在A431細胞中，組織蛋白去乙醯化酶之抑制劑 (TSA、NaBT) 處理了細胞後，除了增加細胞中的乙醯化程度，也非專一性的抑制了上皮生長因子所誘導之JNK活性，進而減少c-Jun磷酸化及蛋白質表現，這一結果導致十二位酯氧酵素活性的抑制。

Eukayotic transcription is a highly regulated process, and histone acetylation is now known to play a major role in this regulation. Acetylation of histone N-termini reduces chromatin condensation; thus gene transcription may activate. Previously, we found that c-Jun induction was required in epidermal growth factor (EGF) induced human 12(S)-lipoxygenase (12(S)-LOX) gene expression in human epidermoid carcinoma A431 cells. In this study, we studied the effect of histone deacetylase inhibitors, trichostatin A (TSA) and sodium butyrate (NaBT) on EGF induced expression of 12(S)-LOX. TSA could enhance the acetylation level of histone H4 within one hour. In order to study the effect of TSA and NaBT on EGF-induced expression of 12(S)-LOX, the 12(S)-LOX enzyme activity was determined. TSA and NaBT inhibited EGF-induced 12(S)-LOX enzyme activity in a dose-dependent manner. Both inhibitors did not have any significant effects on phosphorylation of ERK and JNK, induced by EGF. In the in vitro kinase assay, TSA and NaBT decreased JNK activity in cells, but not ERK activity. Both TSA and NaBT reduced EGF-induced N-terminal phosphorylation and expression of c-Jun. Furthermore, in cells overexpression of c-Jun mutant (N 1-220), which contains c-Jun N-terminal 1-220 amino acids, we found that TSA inhibited EGF-induced c-Jun Ser63 and Ser73 phosphorylation of c-Jun mutant (N 1-220). These results suggest that the inhibition of TSA and NaBT on EGF-induced the expression of 12(S)-LOX was due to inhibit JNK activity and then attenuate N-terminal phosphorylation of c-Jun, which led to the inhibition of c-Jun expression.

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