Phenotypes and gene expression profiles of Saccharopolyspora erythraea rifampicin-resistant (rif) mutants affected in erythromycin production.
Phenotypes and gene expression profiles of Saccharopolyspora erythraea rifampicin-resistant (rif) mutants affected in erythromycin production.
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DOI：
10.1186/1475-2859-8-18
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发表时间：
2009-03-30
影响因子：
6.4
通讯作者：
Alifano P
中科院分区：
文献类型：
--
作者：
Carata E;Peano C;Tredici SM;Ferrari F;Talà A;Corti G;Bicciato S;De Bellis G;Alifano P
之前的研究有证据表明，RNA 聚合酶 (RNAP) 的基因操作可能会刺激细菌的次生代谢。在这项研究中，我们使用利福平选择作为遗传改良红霉素生产者红糖多孢菌的策略。从亲本菌株 NRRL2338 中分离出自发的利福平耐药 (rif) 突变体，并对 rpoB 内的两个 rif 突变（S444F 和 Q426R）进行了表征。相对于亲本菌株，S444F 突变体表现出更高的呼吸性能和高达四倍的最终红霉素产量；相比之下，Q426R突变体生长缓慢、发育缺陷并且红霉素生产严重受损。 DNA 微阵列分析表明，这些 rif 突变深刻改变了 S. erythraea 的转录谱。编码碳（和能量）和氮中心代谢关键酶的基因表达发生显着改变，进而影响通过红霉素饲养途径的代谢物流量。特别是，在S444F突变体中，为红霉素前体6-脱氧赤酮内酯B的生物合成提供丙酰辅酶A的缬氨酸分解代谢途径被强烈上调，而红霉素生物合成基因簇（ery）的表达并未受到显着影响。相反，ery 簇在 Q426R 突变体中下调（<2 倍）。这些菌株还表现出对氮调节子的令人印象深刻的刺激，这可能导致红霉素产量降低，因为红霉素的产生受到铵的强烈抑制。利福平选择是一种简单而可靠的工具，可用于研究红霉素初级代谢和次级代谢与形态分化之间的新联系，并提高红霉素的产量。同时，使用 DNA 微阵列对表达谱进行全基因组分析，可以获得有关 rif 突变对红霉素产生的刺激/抑制作用的机制的信息。
There is evidence from previous works that bacterial secondary metabolism may be stimulated by genetic manipulation of RNA polymerase (RNAP). In this study we have used rifampicin selection as a strategy to genetically improve the erythromycin producer Saccharopolyspora erythraea. Spontaneous rifampicin-resistant (rif) mutants were isolated from the parental strain NRRL2338 and two rif mutations mapping within rpoB, S444F and Q426R, were characterized. With respect to the parental strain, S444F mutants exhibited higher respiratory performance and up to four-fold higher final erythromycin yields; in contrast, Q426R mutants were slow-growing, developmental-defective and severely impaired in erythromycin production. DNA microarray analysis demonstrated that these rif mutations deeply changed the transcriptional profile of S. erythraea. The expression of genes coding for key enzymes of carbon (and energy) and nitrogen central metabolism was dramatically altered in turn affecting the flux of metabolites through erythromycin feeder pathways. In particular, the valine catabolic pathway that supplies propionyl-CoA for biosynthesis of the erythromycin precursor 6-deoxyerythronolide B was strongly up-regulated in the S444F mutants, while the expression of the biosynthetic gene cluster of erythromycin (ery) was not significantly affected. In contrast, the ery cluster was down-regulated (<2-fold) in the Q426R mutants. These strains also exhibited an impressive stimulation of the nitrogen regulon, which may contribute to lower erythromycin yields as erythromycin production was strongly inhibited by ammonium. Rifampicin selection is a simple and reliable tool to investigate novel links between primary and secondary metabolism and morphological differentiation in S. erythraea and to improve erythromycin production. At the same time genome-wide analysis of expression profiles using DNA microarrays allowed information to be gained about the mechanisms underlying the stimulatory/inhibitory effects of the rif mutations on erythromycin production.
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影响因子：
64.5
作者：
Artsimovitch, I;Patlan, V;Vassylyev, DG
通讯作者：
Vassylyev, DG
影响因子：
4.4
作者：
Brockmann-Gretza O;Kalinowski J
通讯作者：
Kalinowski J
影响因子：
2.1
作者：
Irizarry, RA;Hobbs, B;Speed, TP
通讯作者：
Speed, TP
DOI：
10.1073/pnas.0803622105
发表时间：
2008-08-12
影响因子：
11.1
作者：
Chng, Chinping;Lum, Amy M.;Kao, Camilla M.
通讯作者：
Kao, Camilla M.
DOI：
10.1073/pnas.90.15.7119
发表时间：
1993-08-01
影响因子：
11.1
作者：
DONADIO, S;MCALPINE, JB;KATZ, L
通讯作者：
KATZ, L