División de Genética, Universidad de Alicante, Apartado 99, E-03080 Alicante, Spain.
Lehrstuhl für Mikrobiologie, Organismische Interaktionen, University Tübingen, Auf der Morgenstelle 28 D-72076 Tübingen, Germany.
Microbiology (Reading). 2010 May;156(Pt 5):1517-1526. doi: 10.1099/mic.0.037309-0. Epub 2010 Jan 28.
In Synechococcus elongatus sp. PCC 7942, PipX forms complexes with P(II), a protein found in all three domains of life as an integrator of signals of the nitrogen and carbon balance, and with the cyanobacterial nitrogen regulator NtcA. We recently showed that previous inactivation of pipX facilitates subsequent inactivation of the glnB gene. Here, we show that the three spontaneous pipX point mutations pipX-92delT, pipX160C>T and pipX194T>A, initially found in different glnB strains, are indeed suppressor mutations. When these mutations were reconstructed in the wild-type background, the glnB gene could be efficiently inactivated. Furthermore, the point mutations have different effects on PipX levels, coactivation of NtcA-dependent genes and protein-protein interactions. Further support for an in vivo role of PipX-P(II) complexes is provided by interaction analysis with the in vivo-generated P(II)(T-loop+7) protein, a P(II) derivative unable to interact with its regulatory target N-acetyl-l-glutamate kinase, but which retains the ability to bind to PipX. The implications of these results are discussed.
在集胞藻 PCC 7942 中，PipX 与 P(II)形成复合物，P(II)是一种存在于所有三个生命领域的整合氮和碳平衡信号的蛋白，也是蓝细菌氮调节因子 NtcA 的整合蛋白。我们最近表明，先前 pipX 的失活会促进随后 glnB 基因的失活。在这里，我们表明，最初在不同的 glnB 菌株中发现的三个自发 pipX 点突变 pipX-92delT、pipX160C>T 和 pipX194T>A 确实是抑制突变。当这些突变在野生型背景中重建时，glnB 基因可以被有效地失活。此外，点突变对 PipX 水平、NtcA 依赖性基因的共激活和蛋白质-蛋白质相互作用有不同的影响。与体内生成的 P(II)(T 环+7)蛋白的相互作用分析进一步支持了 PipX-P(II)复合物的体内作用，P(II)(T 环+7)蛋白是一种不能与其调节靶标 N-乙酰-l-谷氨酸激酶相互作用的 P(II)衍生物，但保留与 PipX 结合的能力。讨论了这些结果的含义。
