在水稻種子的萌發過程中，α-澱粉水解酶基因的表現受到糖與及吉貝素(GAs)的調控。TA box在先前研究中已被確認為糖與吉貝素調控α-澱粉水解酶基因表現的重要順式DNA序列。在我們之前的研究中已知OsMYBS1、OsMYBS2、OsMYBS3會與TA box有專一性鍵結，利用暫時性分析得知OsMYBS2會專一性的鍵結在TA box上，並且抑制TA box:: Luciferase的表現。另外，OsMYBS2在大麥中的同源基因HvMCB1根據研究也指出HvMCB1在N端的1~41個胺基酸對其抑制α-澱粉水解酶基因表現為必須的。 在本篇論文中，我們利用暫時性分析得知OsMYBS2的N端(1?53的胺基酸)對抑制轉錄活性並非為必須的，並利用轉殖植物的穩定表現系統進一步確認OsMYBS2(tr)(N端刪除的OsMYBS2)在水稻中是否對於α-澱粉水解酶基因有抑制轉錄調控的功能。與野生型比較之後發現，OsMYBS2(tr)大量表現時會讓水稻細胞的α-澱粉水解酶基因表現對糖抑制作用較敏感，而OsMYBS2基因靜默時會讓水稻細胞的α-澱粉水解酶基因表現受糖抑制的速度減緩。在缺糖的處理下，雖然水稻細胞中OsMYBS2的mRNA含量僅有少量的減少，但是OsMYBS2(tr)蛋白質含量卻有大幅的下降，表示OsMYBS2(tr)有受到轉錄後的調控。在種子萌發與生長的實驗中，OsMYBS2(tr)大量表現的種子的萌發以及生長速度會被延遲，這也表示OsMYBS2參與在水稻種子的萌發以及幼苗生長的調控中。 During rice germination, α-amylase genes are controlled by sugar and gibberellins (GAs). The cis-eletment, TA box (TATCCA), has been identified and showed that it plays an important role both in sugar- and in GA-regulated α-amylase gene expression. In our previous study, TA box binding factors, OsMYBS1, OsMYBS2, and OsMYBS3, were identified. Transient expression assay showed that OsMYBS2 recognized TA box specially and functioned as a repressor. Previous studies have shown that the barley HvMCB1 is the orthologous gene of OsMYBS2 and the N-terminal (residues 1-41) of HvMCB1 is essential for negative regulation of α-amylase. In this study, we showed that the N-terminal (residue 1-53) of OsMYBS2 is not essential for the repression of α-amylase by transient assay. Transgenic rice overexpressing OsMYBS2(tr) (N-terminal truncated version of OsMYBS2) shown that OsMYBS2(tr) is a repressor of α-amylase gene. Compared to wild-type, overexpression of OsMYBS2(tr) caused α-amylase gene hypersensitive to sugar repression, whereas RNAi of OsMYBS2 slowed down the sugar repression response. Although sugar starvation slightly decreased OsMYBS2 mRNA, protein level of OsMYBS2(tr) was much less, implying that OsMYBS2(tr) was regulated post-transcriptionally. Seed germination assay indicated that germination rate was delayed in OsMYBS2(tr) overexpression plants. Taken together, it shows that OsMYBS2 plays a role in regulating seed germination and seeding growth in rice.
