Mammalian Sterile 20-like kinase 3 (Mst3) 為一個近年來新發現未知功能的蛋白激酶，它屬於germinal center kinase-III (GCK-III) 家族蛋白激酶。它包含一個演化保留的NH2端激酶區域，以及一個COOH端調控區域。其它的MST家族成員主要有3個，其中Mst1、2屬於GCK-II，而Mst4與Mst3同屬於GCK-III。在功能上，Mst1以及Mst2已被證實參與在細胞凋亡訊息途徑中。而胺基酸序列與Mst3相似度最高(~88%)的Mst4，主要是調控腫瘤細胞的生長、細胞的轉形調控，以及抑制細胞移動。然而，雖然Mst3也被證實參與在細胞凋亡訊息途徑，但對於Mst3的生理功能目前仍不是很清楚。

　　由於Mst3普遍表現在各組織以及細胞中，因此，我們實驗室想利用小片段干擾核醣核酸(siRNA)技術，抑制掉細胞中的Mst3表現，進一步研究Mst3在細胞中所扮演的生理功能。我們由Mst3訊息核醣核酸(mRNA)，挑選六段不同序列片段來構築Mst3 RNAi質體。藉由同時送入RNAi 質體與HA-Mst3質體短暫表現在細胞中，證實有兩種Mst3 RNAi能有效抑制外送的HA-Mst3表現。之後，藉由轉送(transfection)以及G418藥物篩選，使Mst3 RNAi能穩定表現在人類乳癌細胞(MCF-7)中。藉由西方點墨法分析證實，我們利用藥物篩選所建立的Mst3 RNAi細胞株，細胞內的Mst3蛋白表現明顯被抑制下來。之後經由MTT分析細胞生長結果發現，控制組(vector)及實驗組(RNAi)細胞的增生情形，在比較上並無差異。表示細胞內Mst3表現量降低，並不會影響細胞的生長速率。

　　但在各種細胞移動實驗，如:細胞癒合實驗、及Boyden chamber細胞移動分析中，發現相對於控制組(vector)，實驗組(RNAi)細胞株的細胞移動明顯增加。此外，我們將Mst3過度表現在細胞容易移動的MDCK細胞中，發現細胞移動明顯被抑制。因此，由以上結果我們推斷Mst3在細胞中，可能是扮演抑制細胞移動的角色。而在未來我們將更進一步研究Mst3抑制細胞移動的分子機制。

　　Mammalian Sterile 20-like kinase 3 (Mst3), the physiological functions of which is unknown, is a member of the germinal center kinase-III family. It contains a conserved kinase domain at its NH2 terminus, and a regulatory domain at its COOH terminus. There are another three members of MST family. The Mst1/Mst2 belongs to GCK-II subfamily and the Mst4 belongs to the GCK-III. The Mst1 and Mst2 are involved in inducing apoptosis. The Mst4, which shares 88% sequence similarity to Mst3, mediates cell growth, cell transformation, and cell motility instead of promote apoptosis. Nevertheless, the physiological functions of Mst3 are unknown.

　　The Mst3 is ubiquitously expressed in most tissues and cell lines. In order to study the biological function, we exploited the small interfering RNA (siRNA) technique to suppress Mst3 expression. We constructed Mst3 RNAi plasmid targeting six different locations of Mst3 mRNA. Transient transfection of RNAi and HA-Mst3 indicate that two of Mst3 RNAi could suppress exogenous Mst3 expression. One of the functional Mst3 RNAi plasmid was stably expressed in MCF-7 cell by transfection. Western blot analysis indicated that the Mst3 protein expression was significantly silenced by Mst3 RNAi plasmid comparing with the vector control. We investigated whether cell proliferation and cell motility was altered by down-regulating expression of Mst3. There was no difference of cell proliferation between Mst3 RNAi clone and control cells. In contrast, the cell motility was enhanced in Mst3 RNAi clone in wounding assay, boyden chamber assay, and cell Immunofluorescence. Further more, the cell migration was evidently reduced when we stably overexpressed Mst3 in MDCK cell. Therefore, we propose that Mst3 plays a role of inhibiting cell motility. Further study is required to understand the molecular mechanism how Mst3 regulates cell migration.

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