Department of Integrative Biology, University of South Florida, Tampa, Florida, USA.
J Bacteriol. 2012 Apr;194(8):2074-81. doi: 10.1128/JB.06504-11. Epub 2012 Feb 10.
The hydrothermal vent gammaproteobacterium Thiomicrospira crunogena inhabits an unstable environment and must endure dramatic changes in habitat chemistry. This sulfur chemolithoautotroph responds to changes in dissolved inorganic carbon (DIC) (DIC = CO(2) + HCO(3)(-) + CO(3)(-2)) availability with a carbon-concentrating mechanism (CCM) in which whole-cell affinity for DIC, as well as the intracellular DIC concentration, increases substantially under DIC limitation. To determine whether this CCM is regulated at the level of transcription, we resuspended cells that were cultivated under high-DIC conditions in chemostats in growth medium with low concentrations of DIC and tracked CCM development in the presence and absence of the RNA polymerase inhibitor rifampin. Induction of the CCM, as measured by silicone oil centrifugation, was hindered in the presence of rifampin. Similar results were observed for carboxysome gene transcription and assembly, as assayed by quantitative reverse transcription-PCR (qRT-PCR) and transmission electron microscopy, respectively. Genome-wide transcription patterns for cells grown under DIC limitation and those grown under ammonia limitation were assayed via microarrays and compared. In addition to carboxysome genes, two novel genes (Tcr_1019 and Tcr_1315) present in other organisms, including chemolithoautotrophs, but whose function(s) has not been elucidated in any organism were found to be upregulated under low-DIC conditions. Likewise, under ammonia limitation, in addition to the expected enhancement of ammonia transporter and P(II) gene transcription, the transcription of two novel genes (Tcr_0466 and Tcr_2018) was measurably enhanced. Upregulation of all four genes (Tcr_1019, 4-fold; Tcr_131, ∼7-fold; Tcr_0466, >200-fold; Tcr_2018, 7-fold), which suggests that novel components are part of the response to nutrient limitation by this organism, was verified via qRT-PCR.
嗜热硫还原杆菌栖息于不稳定的环境中，必须耐受栖息地化学物质的剧烈变化。这种硫化学自养生物对溶解无机碳 (DIC) (DIC = CO2 + HCO3(-) + CO3(2-)) 可用性的变化作出反应，当 DIC 受到限制时，会通过全细胞对 DIC 的亲和力以及细胞内 DIC 浓度的大幅增加来实现碳浓缩机制 (CCM)。为了确定这种 CCM 是否在转录水平上受到调节，我们将在高 DIC 条件下培养的细胞悬浮在恒化器中的生长培养基中，其中 DIC 浓度较低，并在有无 RNA 聚合酶抑制剂利福平的情况下跟踪 CCM 的发展。如硅酮油离心所测量的那样，在利福平存在下，CCM 的诱导受到阻碍。通过定量逆转录 PCR (qRT-PCR) 和透射电子显微镜分别检测到羧化体基因转录和组装的相似结果。通过微阵列检测在 DIC 限制和氨限制下生长的细胞的全基因组转录模式，并进行比较。除了羧化体基因外，还发现了其他生物体中存在的两个新基因（Tcr_1019 和 Tcr_1315），包括化学自养生物，但在任何生物体中尚未阐明其功能，在低 DIC 条件下被上调。同样，在氨限制下，除了预期增强氨转运体和 P(II)基因的转录外，两个新基因（Tcr_0466 和 Tcr_2018）的转录也可测量增强。所有四个基因（Tcr_1019，上调 4 倍；Tcr_131，上调约 7 倍；Tcr_0466，上调超过 200 倍；Tcr_2018，上调 7 倍）的上调表明，新的成分是该生物体对营养限制的反应的一部分，通过 qRT-PCR 进行了验证。
