Department of Chemical Engineering, Texas A & M University, College Station, TX 77843-3122, USA.
Environ Microbiol Rep. 2010 Jun;2(3):449-55. doi: 10.1111/j.1758-2229.2010.00171.x.
Inactivating the tyrosine phosphatase TpbA of Pseudomonas aeruginosa PA14 induces biofilm formation by 150-fold via increased production of the second messenger cyclic diguanylic acid (c-di-GMP). Here, we show the tpbA mutation reduces extracellular DNA (eDNA) and that increased expression of tpbA increases eDNA; hence, eDNA is inversely proportional to c-di-GMP concentrations. Mutations in diguanylate cyclases PA0169, PA4959 and PA5487 and phosphodiesterase PA4781 corroborate this trend. The tpbA mutation also decreases cell lysis while overexpression of tpbA increases cell lysis. To further link c-di-GMP concentrations and eDNA, the gene encoding phosphodiesterase PA2133 was overexpressed which increased eDNA and decreased biofilm formation by decreasing c-di-GMP. Furthermore, the effect of the tpbB mutation along with the tpbA mutation was examined because loss of TpbB restored the phenotypes controlled by enhanced c-di-GMP in the tpbA mutant. The tpbA tpbB double mutations restored eDNA to that of the PA14 wild-type level. These findings suggest that c-di-GMP, rather than TpbA, controls eDNA. Hence, TpbA acts as a positive regulator of eDNA and cell lysis by reducing c-di-GMP concentrations.
铜绿假单胞菌 PA14 的酪氨酸磷酸酶 TpbA 的失活通过增加第二信使环二鸟苷酸（c-di-GMP）的产生将生物膜形成诱导增加 150 倍。在这里，我们表明 tpbA 突变会减少细胞外 DNA（eDNA），并且 tpbA 的过度表达会增加 eDNA；因此，eDNA 与 c-di-GMP 浓度成反比。双鸟苷酸环化酶 PA0169、PA4959 和 PA5487 以及磷酸二酯酶 PA4781 的突变证实了这一趋势。tpbA 突变还会降低细胞裂解，而 tpbA 的过度表达会增加细胞裂解。为了进一步将 c-di-GMP 浓度与 eDNA 联系起来，过表达了编码磷酸二酯酶 PA2133 的基因，该基因增加了 eDNA，并通过降低 c-di-GMP 减少了生物膜形成。此外，还检查了 tpbB 突变与 tpbA 突变的协同作用，因为 TpbB 的缺失恢复了 tpbA 突变体中增强的 c-di-GMP 控制的表型。tpbA tpbB 双突变将 eDNA 恢复到 PA14 野生型水平。这些发现表明 c-di-GMP 而不是 TpbA 控制 eDNA。因此，TpbA 通过降低 c-di-GMP 浓度，作为 eDNA 和细胞裂解的正调控因子。
