Mannose-6-phosphate isomerase調控靈菌群體表面移行之多細胞行為

Abstract

Serratia marcescens是一種會產生紅色色素，隸屬腸道桿菌科 (Enterobacteriaceae)中靈菌屬 (Serratia)的革蘭氏陰性菌。其為一伺機性致病菌，易造成院內感染，而導致免疫低下的病人致病。Swarming為其細菌群體在固體培養基表面移行的一種特殊型態。S. marcescens 野生株CH-1在30℃時，具有在0.8% 培養基上swarming的現象，但在37℃的環境下，卻無swarming的表現型態。為進一步瞭解其中的機制，利用transposon mutagenesis (mini-Tn5-km1) 的方式，在37℃找到一株precocious-swarming mutant。對其進行研究發現，transposon嵌入的位置，在manA ORF上游約211 nucleotides處，此位置位於預測promoter region的範圍內。推測可能是由於此基因的表現量改變，造成37℃時的表現型與野生株有所差異。而目前已知manA製造出的蛋白質mannose-6-phosphate isomerase，其功能為轉換fructose-6-P (Fru-6-P) 及 mannose-6-P (Man-6-P)，是連接能量代謝及mannose代謝的橋樑。為了進一步研究37℃時precocious-swarming的原因是否導因於manA基因的過量表現或者降低表現，我針對野生株CH-1進行了manA的基因剔除以及overexpression的實驗。取得突變株後，觀察其與CH-1在37℃時swarming的表現，推測manA的過量表現應為造成precocious-swarming的原因，且可能同時活化能量代謝及mannose代謝途徑而達成。

Serratia marcescens is a red-pigmented gram-negative enteric organism. In addition, it is an opportunistic human pathogen and causes many nosocomial infections. Swarming is a specialized form of bacterial surface translocation when bacteria were cultured on surface of solid media. S. marcescens CH-1 swarms on 0.8% LB agar at 30℃, but not at 37℃. In order to understand the underlying mechanism of temperature-dependent regulation of swarming behavior, transposon mutagenesis was performed to screen for the mutants that swarm at 37℃. One precocious-swarming mutant (named WW100) was found and further experiments showed that the insertion site of transposon lay in the predicted promoter region of manA. The manA encodes a protein named mannose-6-phosphate isomerase whose function is interconverting fructose-6-phosphate and mannose-6-phosphate, and linking the glucose and mannose metabolic pathway. In order to investigate whether the phenomenon of WW100 precocious-swarming at 37℃ was due to increase or decrease in expression of manA gene, knockout and over-expression for manA were performed. The results here showed that precocious-swarming phenotype of S. marcescens WW100 was probably because of over-expression of manA, leading to activation of both energy metabolic and mannose metabolic pathways.