Laboratório de Biotecnologia (Setor de Biologia Integrativa), Universidade Estadual do Norte, Fluminense Darcy Ribeiro (UENF), Av. Alberto Lamego, 2000, Campos dos Goytacazes, Rio de Janeiro, Brazil.
Instituto de Química (Departamento de Bioquímica), Universidade de São Paulo, Av. Prof. Lineu Prestes, 748, São Paulo, SP, Brazil.
Microbiol Res. 2021 Feb;243:126654. doi: 10.1016/j.micres.2020.126654. Epub 2020 Nov 22.
The use of plant growth-promoting bacteria represents an alternative to the massive use of mineral fertilizers in agriculture. However, some abiotic stresses commonly found in the environment, like salinity, can affect the efficiency of this approach. Here, we investigated the key mechanisms involved in the response of the plant growth-promoting bacterium Gluconacetobacter diazotrophicus to salt stress by using morphological and cell viability analyses, comparative proteomics, and reverse genetics. Our results revealed that the bacteria produce filamentous cells in response to salt at 100 mM and 150 mM NaCl. However, such a response was not observed at higher concentrations, where cell viability was severely affected. Proteomic analysis showed that salt stress modulates proteins involved in several pathways, including iron uptake, outer membrane efflux, osmotic adjustment, cell division and elongation, and protein transport and quality control. Proteomic data also revealed the repression of several extracytoplasmic proteins, especially those located at periplasm and outer membrane. The role of such pathways in the tolerance to salt stress was analyzed by the use of mutant defectives for Δtbdr (iron uptake), ΔmtlK and ΔotsA (compatible solutes synthesis), and ΔdegP (quality control of nascent extracytoplasmic proteins). ΔdegP presented the highest sensitivity to salt stress, Δtbdr, andΔmtlK also showed increased sensitivity, but ΔotsA was not affected. This is the first demonstration that DegP protein, a protease with minor chaperone activity, is essential for tolerance to salt stress in G. diazotrophicus. Our data contribute to a better understanding of the molecular bases that control the bacterial response/tolerance to salt stress, shedding light on quality control of nascent extracytoplasmic proteins.
植物促生菌的使用代表了一种替代农业中大量使用矿物肥料的方法。然而，一些常见的环境非生物胁迫，如盐度，会影响这种方法的效率。在这里，我们通过形态和细胞活力分析、比较蛋白质组学和反向遗传学研究了植物促生菌 Gluconacetobacter diazotrophicus 对盐胁迫的反应的关键机制。我们的结果表明，细菌在 100mM 和 150mM NaCl 下会产生丝状细胞以应对盐胁迫。然而，在更高的浓度下没有观察到这种反应，因为细胞活力受到严重影响。蛋白质组学分析表明，盐胁迫调节了参与几个途径的蛋白质，包括铁摄取、外膜外排、渗透调节、细胞分裂和伸长以及蛋白质运输和质量控制。蛋白质组学数据还表明，几个细胞外蛋白质，特别是那些位于周质和外膜的蛋白质被抑制。通过使用 Δtbdr（铁摄取）、ΔmtlK 和 ΔotsA（相容性溶质合成）和 ΔdegP（新生细胞外蛋白质的质量控制）缺陷突变体分析了这些途径在耐受盐胁迫中的作用。ΔdegP 对盐胁迫最敏感，Δtbdr 和 ΔmtlK 也表现出较高的敏感性，但 ΔotsA 不受影响。这是首次证明 DegP 蛋白，一种具有较小伴侣活性的蛋白酶，对于 G. diazotrophicus 耐受盐胁迫是必不可少的。我们的数据有助于更好地理解控制细菌对盐胁迫的反应/耐受的分子基础，阐明了新生细胞外蛋白质的质量控制。
