Small subunits of RNA polymerase: localization, levels and implications for core enzyme composition
Small subunits of RNA polymerase: localization, levels and implications for core enzyme composition
复制标题
DOI：
10.1099/mic.0.041566-0
复制
发表时间：
2010-12-01
期刊：
影响因子：
2.8
通讯作者：
Lewis, Peter J.
中科院分区：
文献类型：
--
作者：
Doherty, Geoff P.;Fogg, Mark J.;Lewis, Peter J.
细菌RNA聚合酶（RNAP）含有已知与核心α、β和β '亚基共纯化的几个小的辅助亚基。Ω亚基在革兰氏阳性和革兰氏阴性细菌之间是保守的，而δ亚基在革兰氏阳性细菌内是保守的，但限于革兰氏阳性细菌。虽然各种功能已被分配到这些亚基通过体外试验，很少有人知道他们在体内的作用。在这项工作中，我们构建了一对载体来研究枯草芽孢杆菌中相对于核心RNAP的δ和ω亚基的亚细胞定位。我们发现这些亚基与参与转录mRNA和rRNA操纵子的RNAP密切相关。这些亚基的定量揭示了δ以与RNAP等摩尔的水平存在，ω以核心RNAP水平的一半左右存在。为了比较，还研究了大肠杆菌中RNAP β '和ω亚基的定位和定量。类似于B。subtilis、beta'和omega与类核紧密结合并形成高绿色荧光蛋白强度的亚类核区域，但与B中的omega不同。枯草杆菌、大肠杆菌中的欧米茄水平。大肠杆菌与β ′菌株接近。这些结果表明，δ可能是一个完整的RNAP亚基在革兰氏阳性，而欧米茄水平显着不同的革兰氏阳性和阴性之间。欧米茄亚基可能是RNAP组装所需的，随后以不同的速率被翻转，或者它可能在革兰氏阴性菌中发挥作用，而革兰氏阳性菌中的其他因子则发挥作用。
Bacterial RNA polymerases (RNAPs) contain several small auxiliary subunits known to co-purify with the core alpha, beta and beta' subunits. The omega subunit is conserved between Gram-positive and Gram-negative bacteria, while the delta subunit is conserved within, but restricted to, Gram-positive bacteria. Although various functions have been assigned to these subunits via in vitro assays, very little is known about their in vivo roles. In this work we constructed a pair of vectors to investigate the subcellular localization of the delta and omega subunits in Bacillus subtilis with respect to the core RNAP. We found these subunits to be closely associated with RNAP involved in transcribing both mRNA and rRNA operons. Quantification of these subunits revealed delta to be present at equimolar levels with RNAP and omega to be present at around half the level of core RNAP. For comparison, the localization and quantification of RNAP beta' and omega subunits in Escherichia coli was also investigated. Similar to B. subtilis, beta' and omega closely associated with the nucleoid and formed subnucleoid regions of high green fluorescent protein intensity, but, unlike omega in B. subtilis, omega levels in E. coli were close to parity with those of beta'. These results indicate that delta is likely to be an integral RNAP subunit in Gram-positives, whereas omega levels differ substantially between Gram-positives and -negatives. The omega subunit may be required for RNAP assembly and subsequently be turned over at different rates or it may play roles in Gram-negative bacteria that are performed by other factors in Gram-positives.