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Identification and Molecular Characterization of Salt Stress Tolerance in Frankia Isolates from Casuarina Plants
Rediet OSHONE1, Mariama NGOM2,3,5, Nathalie DIAGNE3,5, Diegane DIOUF3,5, Valérie HOCHER4,5, Mame Oureye SY2,5, Laurent LAPLAZE4,5, Antony CHAMPION3,4,5, and Louis S. TISA1
1. University of New Hampshire, USA 2. Laboratoire Campus de Biotechnologie Végétale, Faculté des Sciences & Techniques, Université Cheikh Anta Diop (UCAD), Dakar, Sénégal 3. Laboratoire Commun de Microbiologie IRD/ISRA/UCAD, Dakar, Sénégal 4.
Equipe Rhizogenèse, UMR DIADE, IRD, Montpellier, France 5. Laboratoire mixte international Adaptation des Plantes et microorganismes associés aux Stress Environnementaux (LAPSE), Dakar, Sénégal
Introduction
Salinization of soils and groundwater is a
serious problem causing drastic reduction in
agricultural production. Actinorhizal plants
form a symbiotic association with the
actinobacteria, Frankia, and are able to
tolerate a variety of abiotic stresses
including salt stress. Among the actinorhizal
plants, some trees of the genus Casuarina
have been shown to grow well under these
conditions. The bacterial partner, Frankia,
of the actinorhizal symbiosis plays a role in
the ability of these plants to survive under
harsh conditions. The aim of this study was
to identify salt-tolerant Frankia strains and
to determine the genes responsible for the
molecular mechanisms of salt stress
tolerance.
Material and Methods
 A 24-well growth assay was
used to determine salt
tolerance levels for Frankia
strains.
A 24 well plate growth assay 2
CcI3 identified as the most salt
sensitive strain, while Allo2,
BMG5.23, CcI6, and CeD are salt
tolerant
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Genomic synteny of CcI3 to CcI6, Thr, and BMG5.23
Results
Membrane composition changes
under salt stress
Salt sensitive strain shows drastic change
in amino acid profile under stress
RNA sequencing based differential gene expression analysis
in CcI3 exposed to salt and osmotic stress
Trehalose biosynthesis is
involved in the early response
to salinity
Functional category comparison
COG category
CcI6 CcI3 BMG5.23 Thr
Amino acid transport and metabolism
219
218
215
218
Carbohydrate transport and metabolism
133
132
130
135
Cell cycle control, cell division, chromosome partitioning
33
30
34
34
Cell motility
1
1
1
1
142
139
143
134
Cell wall/membrane/envelope biogenesis
Chromatin structure and dynamics
Coenzyme transport and metabolism
1
1
1
1
146
150
141
150
Defense mechanisms
41
39
35
39
Energy production and conversion
201
203
197
204
Function unknown
186
177
188
185
General function prediction only
378
369
365
366
Inorganic ion transport and metabolism
116
118
111
117
Intracellular trafficking, secretion, and vesicular transport
27
28
26
30
Lipid transport and metabolism
159
157
153
166
Nucleotide transport and metabolism
82
85
78
83
Posttranslational modification, protein turnover, chaperones
102
94
98
99
Replication, recombination and repair
180
212
183
169
RNA processing and modification
Secondary metabolites biosynthesis, transport and catabolism
1
113
1
1
1
105
112
116
Signal transduction mechanisms
147
145
147
147
Transcription
240
227
236
234
Translation, ribosomal structure and biogenesis
 RNA sequencing was performed in triplicate
for cells under salt and osmotic stress.
 The genomes of salt tolerant strains were
sequenced and compared to the genome of
the salt sensitive strain.
 Quantitative PCR was performed on a sub set
of the genes to confirm results of the RNA
seq analysis.
 Amino acid analysis was used to determine
changes in the amino acid profile under salt
stress.
Results
Not in COGs
156
150
151
152
2483
2183
2379
2406
Analysis of homologous
coding sequences
*
*
*
*
Conclusion
These preliminary results are uncovering the mechanisms of salt stress
tolerance in Frankia
References
1. Benson DR and Silvester WB 1993 Biology of Frankia strains,actinomycete symbionts of
actinorhizal plants
2. Furnholm T et al. 2012. Development of a semi-high-throughput growth assay for filamentous
actinobacteria Frankia. Arch. Microbiol. 194 13–20
Aldehyde detoxification is important in salt stress tolerance
pH homeostasis and Phospohate
uptake are important under salt
stress
3. Hurst et al. 2014. Draft genome sequence of Frankia sp. Strain Thr. genomeA
4. Ghodhbane-Gtari F. et al. 2013 Draft Genome Sequence of Frankia sp. strain BMG5.23
genomeA
5. Mansour et al.2014. Draft genome sequence of Frankia sp. Strain CcI6. 2(1): e01205-13
Ornithine Biosynthesis is up regulated under salt stress
6. Oshone R et al. 2013. Effect of salt stress on the physiology of Frankia sp. strain
CcI6. J. Biosci. 38:699–702
7. Rengasamy P 2006 World salinization with emphasis on Australia. J. Exp. Bot. 57 1017–
1023
Acknowledgement
This research is supported in part by Hatch NH585, JGI 2012
CSP585, and by the College of Life Sciences and Agriculture at
the University of New Hampshire, Durham. Travel support was
obtained from the MCBS department, Graduate School, and the
Zsigray Fund.