Department of Molecular Microbiology, John Innes Centre, Norwich, United Kingdom.
mBio. 2013 Sep 24;4(5):e00684-13. doi: 10.1128/mBio.00684-13.
WhiA is a highly unusual transcriptional regulator related to a family of eukaryotic homing endonucleases. WhiA is required for sporulation in the filamentous bacterium Streptomyces, but WhiA homologues of unknown function are also found throughout the Gram-positive bacteria. To better understand the role of WhiA in Streptomyces development and its function as a transcription factor, we identified the WhiA regulon through a combination of chromatin immunoprecipitation-sequencing (ChIP-seq) and microarray transcriptional profiling, exploiting a new model organism for the genus, Streptomyces venezuelae, which sporulates in liquid culture. The regulon encompasses ~240 transcription units, and WhiA appears to function almost equally as an activator and as a repressor. Bioinformatic analysis of the upstream regions of the complete regulon, combined with DNase I footprinting, identified a short but highly conserved asymmetric sequence, GACAC, associated with the majority of WhiA targets. Construction of a null mutant showed that whiA is required for the initiation of sporulation septation and chromosome segregation in S. venezuelae, and several genes encoding key proteins of the Streptomyces cell division machinery, such as ftsZ, ftsW, and ftsK, were found to be directly activated by WhiA during development. Several other genes encoding proteins with important roles in development were also identified as WhiA targets, including the sporulation-specific sigma factor σ(WhiG) and the diguanylate cyclase CdgB. Cell division is tightly coordinated with the orderly arrest of apical growth in the sporogenic cell, and filP, encoding a key component of the polarisome that directs apical growth, is a direct target for WhiA-mediated repression during sporulation.
Since the initial identification of the genetic loci required for Streptomyces development, all of the bld and whi developmental master regulators have been cloned and characterized, and significant progress has been made toward understanding the cell biological processes that drive morphogenesis. A major challenge now is to connect the cell biological processes and the developmental master regulators by dissecting the regulatory networks that link the two. Studies of these regulatory networks have been greatly facilitated by the recent introduction of Streptomyces venezuelae as a new model system for the genus, a species that sporulates in liquid culture. Taking advantage of S. venezuelae, we have characterized the regulon of genes directly under the control of one of these master regulators, WhiA. Our results implicate WhiA in the direct regulation of key steps in sporulation, including the cessation of aerial growth, the initiation of cell division, and chromosome segregation.
WhiA 是一种高度不寻常的转录调节因子，与真核同源内切酶家族有关。WhiA 是丝状细菌链霉菌孢子形成所必需的，但在革兰氏阳性菌中也发现了功能未知的 WhiA 同源物。为了更好地理解 WhiA 在链霉菌发育中的作用及其作为转录因子的功能，我们通过染色质免疫沉淀测序 (ChIP-seq) 和微阵列转录谱分析相结合，利用一种新型的链霉菌属模式生物委内瑞拉链霉菌（Streptomyces venezuelae）来鉴定 WhiA 调节子，该模式生物在液体培养中进行孢子形成。调节子包含约 240 个转录单位，WhiA 似乎既是激活子又是抑制剂。对完整调节子上游区域的生物信息学分析，结合 DNA 酶 I 足迹分析，确定了一个短但高度保守的不对称序列 GACAC，与大多数 WhiA 靶标相关。构建一个缺失突变体表明，whiA 是委内瑞拉链霉菌孢子形成隔膜和染色体分离的起始所必需的，并且发现编码链霉菌细胞分裂机制关键蛋白的几个基因，如 ftsZ、ftsW 和 ftsK，在发育过程中直接被 WhiA 激活。还确定了几个编码在发育中具有重要作用的蛋白质的其他基因是 WhiA 的靶标，包括孢子特异性 sigma 因子 σ(WhiG)和二鸟苷酸环化酶 CdgB。细胞分裂与产孢细胞中顶端生长的有序停止紧密协调，编码指导顶端生长的极性体的关键成分的 filP 是 WhiA 介导的孢子形成过程中抑制的直接靶标。
自最初鉴定出链霉菌发育所需的遗传基因座以来，所有 bld 和 whi 发育的主要调控因子都已被克隆和表征，并在理解驱动形态发生的细胞生物学过程方面取得了重大进展。现在的主要挑战是通过剖析连接两者的调控网络，将细胞生物学过程和发育的主要调控因子联系起来。这些调控网络的研究由于委内瑞拉链霉菌作为该属的一种新模型系统的引入而得到了极大的促进，这是一种在液体培养中进行孢子形成的物种。利用委内瑞拉链霉菌，我们对受这些主要调控因子之一 WhiA 直接控制的基因的调节子进行了表征。我们的结果表明，WhiA 参与了孢子形成的关键步骤的直接调控，包括停止气生生长、启动细胞分裂和染色体分离。
