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在枯草桿菌(Bacillus subtilis)孢子生成作用(sporulation)初期，菌體
形態上會有一個極明顯而獨特的改變：不對稱細胞中隔 (asymmetric
septum)的形成。而不對稱細胞中隔的形成則與細胞中隔基因 (septum
forming genes),ftsA & ftsZ,有密切的關係。就枯草桿菌而言 ftsA &
ftsZ (or ftsAZ)，是共同受三個啟動子：P1,P2 and P3 的控制，而為同
時轉錄(co-transcribed)之兩個基因。目前對三個啟動子的瞭解為：在一
般增殖期(vegetative stage)時，ftsAZ受P1及P3的轉錄調控並參與對稱
性細胞中隔(symmetric septum)的形成；而在孢子生成期 (sporulation
phase)，則由P2調控轉錄而參與不對稱細胞中隔的形成。在以前的研究顯
示，P1及P3是受σA(rpoD)的辨認及操控；P2則受σH (spoOH)的控制而進
行轉錄， 但對於後者卻一直沒有直接証據證實。本論文最主要目的即以
對偶性抑制作用(allele-specific suppresion)來證明P2是直接受σH調
控的。我們將P2的-10區域中-14,- 13,-12,等三個鹽基分別加以突變(C→
T,G→A,A→G)，放入σH突變的枯草桿菌(spoOH81)中，結果發現：原本在
野生型枯草桿菌(MB24)中無法發揮功用之變異-13鹽基的P2啟動子，在
spoOH81中卻又恢復部份啟動子功能，表示P2與σH之間有專一對偶性的關
聯，證實P2是受σH的直接調控。另外，我們亦針對P2啟動子區域進行一
連串刪剪突變(deletion mutation)，結果發現：P2必須至少含有第-55鹽
基以下的區域，才能使後方的基因保有部份轉錄作用的進行。
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The asymmetric septum formation is a very signal and special
character at the initiation of Bacillus subtilis sporulation.
The septum formation genes, ftsA and ftsZ (or ftsAZ), are co-
transcribed and controlled by three promoter (P1,P2 and P3) in
Bacillus subtilis. The transcription of ftsAZ from P1 or P3 is
required for the symmetric septum formation during vegetative
stage. In contrast, expression of ftsAZ from P2 is only at
sporulation phase. The σH, a very important transcriptional
factor in sporulation, was suspected to recognize the ftsAZ P2
promoter but didn''t confirm yet. We mutated the -14, -13 and
-12 base in -10 region of the ftsAZ P2 promoter (C→T, G→A, A→
G) by site-specific mutagenesis and transformed these mutated
P2 promoter into wild-type (MB24) and σH mutated(spoOH81)
Bacillus subtilis. We found the -13 base mutation could
completely inactivate the P2 promoter in MB24; but P2 promoter
still had 37% activity in spoOH81. These results suggested
that the P2 promoter is recognized by σH directly. In
addition, we further determined the minimum control region of
P2 promoter for ftsAZ transcription. The deletion mutants of
the ftsAZ P2 promoter were constructured either by exonuclease
III or by PCR methods. Total 8 different fragments of ftsAZ P2
promoter clones were obtained. Subcloned these fragments into
transcriptional fusion vector, pDH32, and measured the level of
lacZ (reporter gene of pDH32) expression byβ-Galactosidase
activity assay. The results suggested that at least 55 bases
upstream of the ftsAZ transcriptional initiation site is the
required region for P2 promoter function.
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