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LEA proteins are involved in cyst desiccation resistance and other abiotic stresses in Azotobacter vinelandii

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Cell Stress and Chaperones Aims and scope

Abstract

Late embryogenesis abundant (LEA) proteins constitute a large protein family that is closely associated with resistance to abiotic stresses in multiple organisms and protect cells against drought and other stresses. Azotobacter vinelandii is a soil bacterium that forms desiccation-resistant cysts. This bacterium possesses two genes, here named lea1 and lea2, coding for avLEA1 and avLEA2 proteins, both containing 20-mer motifs characteristic of eukaryotic plant LEA proteins. In this study, we found that disruption of the lea1 gene caused a loss of the cysts’ viability after 3 months of desiccation, whereas at 6 months, wild-type or lea2 mutant strain cysts remained viable. Vegetative cells of the lea1 mutant were more sensitive to osmotic stress; cysts developed by this mutant were also more sensitive to high temperatures than cysts or vegetative cells of the wild type or of the lea2 mutant. Expression of lea1 was induced several fold during encystment. In addition, the protective effects of these proteins were assessed in Escherichia coli cells. We found that E. coli cells overexpressing avLEA1 were more tolerant to salt stress than control cells; finally, in vitro analysis showed that avLEA1 protein was able to prevent the freeze thaw-induced inactivation of lactate dehydrogenase. In conclusion, avLEA1 is essential for the survival of A. vinelandii in dry conditions and for protection against hyper-osmolarity, two major stress factors that bacteria must cope with for survival in the environment. This is the first report on the role of bacterial LEA proteins on the resistance of cysts to desiccation.

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Authors and Affiliations

  1. Departamento de Microbiología Molecular, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Av. Universidad 2001, Col. Chamilpa, 62210, Cuernavaca, Morelos, Mexico

    Julieta Rodriguez-Salazar, Soledad Moreno & Guadalupe Espín

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  1. Julieta Rodriguez-Salazar
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  2. Soledad Moreno
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  3. Guadalupe Espín
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Correspondence to Guadalupe Espín.

Electronic supplementary material

Fig. S1

Schematic representation of the plasmids constructed in this study, and the integration of plasmid pLEA11012 into the chromosome of strain lea1-lea2. Small arrows represent oligonucleotides used for the construction of the plasmids, and to confirm the presence of the mutations in the strains constructed (GIF 13 kb).

High Resolution Image (TIFF 439 kb).

Fig. S2

In silico analysis of avLEA1 and avLEA2. (a), Hydrophilic pattern of avLEA1 and avLEA2 using Kyte-Doolittle hydropathy plots; regions above a hydropathy score of zero are hydrophobic. (b), Prediction of protein disorder of avLEA1 and avLEA2 using web server PONDR and default parameters (GIF 120 kb).

High Resolution Image (TIFF 2888 kb).

Fig. S3

SDS-PAGE and Immunoblotting analysis of avLEA1. Protein samples were analysed by SDS-PAGE (a), lane M Molecular mass marker; lane 1, soluble supernatant from the E. coli BL(DE3) /pavLEA1 without IPTG; or induced with 1 mM IPTG, lane 2; lane 3 purified recombinant avLEA1 protein. (b), Detection of avLEA1 protein containing 6xHis-Tag by Immunoblotting. (GIF 26 kb).

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Rodriguez-Salazar, J., Moreno, S. & Espín, G. LEA proteins are involved in cyst desiccation resistance and other abiotic stresses in Azotobacter vinelandii . Cell Stress and Chaperones 22, 397–408 (2017). https://doi.org/10.1007/s12192-017-0781-1

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  • DOI: https://doi.org/10.1007/s12192-017-0781-1

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