To determine microbial evolutionary strategies to low-pressure (LP; 5 kPa) growth, an environmental condition not experienced on Earth until ∼20 km in altitude, a previously described evolutionary experiment was conducted. The resulting LP evolved strain WN1106, isolated from the terminus of the experiment, was shown to have several genomic mutations absent in the ancestral strain, WN624. Three of the mutations were in regulatory genes: resD, walK, and rnjB. Here we report on transcriptional microarray data from the LP-evolved WN1106 and compare those results with the previously reported ancestral WN624 transcriptional array data at either 5 or 101 kPa. At 5 kPa, WN1106 differentially expresses signals that are under the control of regulators ResD, WalK, and RnjB compared with (1) itself at ∼101 kPa and (2) WN624 at 5 kPa. These results were further confirmed by quantitative reverse transcriptase–polymerase chain reaction of a target transcript from each regulon. This work indicates that the three mutated coding regions had transcriptional control effects on each respective regulon.
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References
1.
AllenEE, FacciottiD, and BartlettDH (1999) Monounsaturated but not polyunsaturated fatty acids are required for growth of the deep-sea bacterium Photobacterium profundum SS9 at high pressure and low temperature. Appl Env Microbiol, 65:1710–1720.
2.
BisicchiaP, NooneD, LioliouE, et al. (2007) The essential YycFG two-component system controls cell wall metabolism in Bacillus subtilis. Mol Microbiol, 65:180–200.
3.
BolstadBM, IrizarryRA, ÅstrandM, et al. (2003) A comparison of normalization methods for high density oligonucleotide array data based on variance and bias. Bioinformatics, 19:185–193.
4.
BoylanR, MendelsonN, BrooksD, et al. (1972) Regulation of the bacterial cell wall: analysis of a mutant of Bacillus subtilis defective in biosynthesis of teichoic acid. J Bacteriol, 110:281–290.
5.
BoylanSA, ThomasMD, and PriceCW (1991) Genetic method to identify regulons controlled by nonessential elements: isolation of a gene dependent on alternate transcription factor sigma B of Bacillus subtilis. J Bacteriol, 173:7856–7866.
6.
BoylanSA, RedfieldAR, BrodyMS, et al. (1993) Stress-induced activation of the sigma B transcription factor of Bacillus subtilis. J Bacteriol, 175:7931–7937.
7.
BrigullaM, HoffmannT, KrispA, et al. (2003) Chill induction of the SigB-dependent general stress response in Bacillus subtilis and its contribution to low-temperature adaptation. J Bacteriol, 185:4305–4314.
8.
BrucknerJ, OsmanS, VenkateswaranK, et al. (2009) Encyclopedia of Microbiology, 3rd ed. Space microbiology: planetary protection, burden, diversity and significance of spacecraft associated. microbes.
9.
CondonC (2010) What is the role of RNase J in mRNA turnover?. RNA Biol, 7:316–321.
10.
DurandS, GiletL, BessièresP, et al. (2012) Three essential ribonucleases-RNase Y, J1, and III-control the abundance of a majority of Bacillus subtilis mRNAs. PLoS Genet, 8:e1002520.
11.
EvenS, PellegriniO, ZigL, et al. (2005) Ribonucleases J1 and J2: two novel endoribonucleases in B. subtilis with functional homology to E.coli RNase E. Nucleic Acids Res, 33:2141–2152.
12.
Fajardo-CavazosP, WatersSM, SchuergerAC, et al. (2012) Evolution of Bacillus subtilis to enhanced growth at low pressure: up-regulated transcription of des-desKR, encoding the fatty acid desaturase system. Astrobiology, 12:258–270.
13.
Fajardo-CavazosP, MorrisonMD, MillerKM, et al. (2018) Transcriptomic responses of Serratia liquefaciens cells grown under simulated Martian conditions of low temperature, low pressure, and CO2-enriched anoxic atmosphere. Sci Rep, 8:14938.
14.
HarwoodCR and CuttingSM (1990) Molecular Biological Methods for Bacillus. Hoboken, NJ: Wiley.
15.
HeckerM, Pané-FarréJ, and UweV (2007) SigB-dependent general stress response in Bacillus subtilis and related gram-positive bacteria. Annu Rev Microbiol, 61:215–236.
16.
HöperD, VölkerU, and HeckerM (2005) Comprehensive characterization of the contribution of individual SigB-dependent general stress genes to stress resistance of Bacillus subtilis. J Bacteriol, 187:2810–2826.
17.
KunstF, OgasawaraN, MoszerI, et al. (1997) The complete genome sequence of the gram-positive bacterium Bacillus subtilis. Nature, 390:249–256.
18.
Laybourn-ParryJ and PearceD (2016) Heterotrophic bacteria in Antarctic lacustrine and glacial environments. Polar Biol, 39:2207–2225.
19.
LivakKJ and SchmittgenTD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2− ΔΔCT method. Methods, 25:402–408.
20.
MäderU, ZigL, KretschmerJ, et al. (2008) mRNA processing by RNases J1 and J2 affects Bacillus subtilis gene expression on a global scale. Mol Microbiol, 70:183–196.
21.
MahaffyPR, WebsterCR, AtreyaSK, et al. (2013) Abundance and isotopic composition of gases in the Martian atmosphere from the Curiosity rover. Science, 341:263–266.
22.
MalinMC and EdgettKS (2003) Evidence for persistent flow and aqueous sedimentation on early Mars. Science, 302:1931–1934.
23.
MartínezG, NewmanC, De Vicente-RetortilloA, et al. (2017) The modern near-surface Martian climate: a review of in-situ meteorological data from Viking to Curiosity. Space Sci Rev, 212:295–338.
24.
MathyN, HébertA, MerveletP, et al. (2010) Bacillus subtilis ribonucleases J1 and J2 form a complex with altered enzyme behaviour. Mol Microbiol, 75:489–498.
25.
MattimoreV and BattistaJR (1996) Radioresistance of Deinococcus radiodurans: functions necessary to survive ionizing radiation are also necessary to survive prolonged desiccation. J Bacteriol, 178:633–637.
26.
MaughanH, CallicotteV, HancockA, et al. (2006) The population genetics of phenotypic deterioration in experimental populations of Bacillus subtilis. Evolution, 60:686–695.
27.
MeersmanF and HeremansK (2008) High hydrostatic pressure effects in the biosphere: from molecules to microbiology. In High-pressure microbiology, edited by MichielsC, BartlettDH, and AertsenA, American Society of Microbiology, Washington, DC, pp. 1–17.
28.
MillerJ (1972) Experiments in Molecular Genetics. Long Island, NY: Cold Spring Harbor.
29.
NicholsonWL (2009) Ancient micronauts: interplanetary transport of microbes by cosmic impacts. Trends Microbiol, 17:243–250.
30.
NicholsonWL (2012) Increased competitive fitness of Bacillus subtilis under nonsporulating conditions via inactivation of pleiotropic regulators AlsR, SigD, and SigW. Appl Env Microbiol, 78:3500–3503.
31.
NicholsonW and SetlowP (1990) Sporulation, germination and outgrowth. In Molecular Biological Methods for Bacillus, edited by HarwoodCR and CuttingSM, John Wiley and Sons, Hoboken, NJ.
32.
NicholsonWL, SchuergerAC, and RaceMS (2009) Migrating microbes and planetary protection. Trends Microbiol, 17:389–392.
33.
NicholsonWL, Fajardo-CavazosP, FedenkoJ, et al. (2010) Exploring the low-pressure growth limit: evolution of Bacillus subtilis in the laboratory to enhanced growth at 5 kilopascals. Appl Environ Microbiol, 76:7559–7565.
34.
NicholsonWL, KrivushinK, GilichinskyD, et al. (2013) Growth of Carnobacterium spp. from permafrost under low pressure, temperature, and anoxic atmosphere has implications for Earth microbes on Mars. Proc Natl Acad Sci, 110:666–671.
35.
OgerPM and JebbarM (2010) The many ways of coping with pressure. Res Microbiol, 161:799–809.
36.
OroseiR, LauroSE, PettinelliE, et al. (2018) Radar evidence of subglacial liquid water on Mars. Science, 361:490–493.
37.
PriceCW (2002) General stress response. In Bacillus subtilis and Its Closest Relatives, edited by SonensheinA, HochJ, and LosickR, American Society of Microbiology, Washington, DC, pp 369–384.
38.
RitchieME, PhipsonB, WuD, et al. (2015) limma powers differential expression analyses for RNA-sequencing and microarray studies. Nucleic Acids Res, 43:e47.
39.
RummelJ, StabekisP, DevincenziD, et al. (2002) COSPAR's planetary protection policy: a consolidated draft. Adv Space Res, 30:1567–1571.
40.
RutishauserA, BlankenshipDD, SharpM, et al. (2018) Discovery of a hypersaline subglacial lake complex beneath Devon Ice Cap, Canadian Arctic. Sci Adv, 4:eaar4353.
41.
SambrookJ, FritschEF, and ManiatisT (1989) Molecular Cloning: A Laboratory Manual. Long Island, NY: Cold Spring Harbor Laboratory Press.
42.
SchaefferP, MilletJ, and AubertJ-P (1965) Catabolic repression of bacterial sporulation. Proc Natl Acad Sci, 54:704–711.
43.
SchroederA, MuellerO, StockerS, et al. (2006) The RIN: an RNA integrity number for assigning integrity values to RNA measurements. BMC Mol Biol, 7:3.
44.
SchuergerAC and NicholsonWL (2006) Interactive effects of hypobaria, low temperature, and CO2 atmospheres inhibit the growth of mesophilic Bacillus spp. under simulated martian conditions. Icarus, 185:143–152.
45.
SchuergerAC and NicholsonWL (2016) Twenty species of hypobarophilic bacteria recovered from diverse soils exhibit growth under simulated martian conditions at 0.7 kPa. Astrobiology, 16:964–976.
46.
SchuergerAC, UlrichR, BerryBJ, et al. (2013) Growth of Serratia liquefaciens under 7 mbar, 0 C, and CO2-enriched anoxic atmospheres. Astrobiology, 13:115–131.
47.
SmythGK and SpeedT (2003) Normalization of cDNA microarray data. Methods, 31:265–273.
48.
SoláM, Gomis-RüthFX, SerranoL, et al. (1999) Three-dimensional crystal structure of the transcription factor PhoB receiver domain. J Mol Biol, 285:675–687.
49.
SpizizenJ (1958) Transformation of biochemically deficient strains of Bacillus subtilis by deoxyribonucleate. Proc Natl Acad Sci, 44:1072–1078.
50.
SteinmetzM and RichterR (1994) Plasmids designed to alter the antibiotic resistance expressed by insertion mutations in Bacillus subtilis, through in vivo recombination. Gene, 142:79–83.
51.
SzurmantH, BuL, BrooksCL, et al. (2008) An essential sensor histidine kinase controlled by transmembrane helix interactions with its auxiliary proteins. Proc Natl Acad Sci U S A, 105:5891–5896.
52.
VenkateswaranK, SatomiM, ChungS, et al. (2001) Molecular microbial diversity of a spacecraft assembly facility. Syst Appl Microbiol, 24:311–320.
53.
WatersSM, Robles-MartínezJA, and NicholsonWL (2014) Exposure of Bacillus subtilis to low pressure (5 kPa) induces several global regulons including the sigB-mediated general stress response. Appl Env Microbiol, 80:4788–4794
54.
WatersSM, ZeiglerDR, and NicholsonWL (2015) Experimental evolution of enhanced growth by Bacillus subtilis at low atmospheric pressure: Genomic changes revealed by whole-genome sequencing. Appl Environ Microbiol, 81:7525–7532.
55.
WinterR and JeworrekC (2009) Effect of pressure on membranes. Soft Matter, 5:3157–3173.
56.
WoodsRJ, BarrickJE, CooperTF, et al. (2011) Second-order selection for evolvability in a large Escherichia coli population. Science, 331:1433–1436.
57.
YeRW, TaoW, BedzykL, et al. (2000) Global gene expression profiles of Bacillus subtilis grown under anaerobic conditions. J Bacteriol, 182:4458–4465.
58.
ZhangX and HulettFM (2000) ResD signal transduction regulator of aerobic respiration in Bacillus subtilis: ctaA promoter regulation. Mol Microbiol, 37:1208–1219.
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