Comparative genomics of the proteostasis network in extreme acidophiles

Abstract

Extreme acidophiles thrive in harsh environments characterized by acidic pH, high concentrations of dissolved metals and high osmolarity. Most of these microorganisms are chemolithoautotrophs that obtain energy from low redox potential sources, such as the oxidation of ferrous ions. Under these conditions, the mechanisms that maintain homeostasis of proteins (proteostasis), as the main organic components of the cells, are of utmost importance. Thus, the analysis of protein chaperones is critical for understanding how these organisms deal with proteostasis under such environmental conditions. In this work, using a bioinformatics approach, we performed a comparative genomic analysis of the genes encoding classical, periplasmic and stress chaperones, and the protease systems. The analysis included 35 genomes from iron- or sulfur-oxidizing autotrophic, heterotrophic, and mixotrophic acidophilic bacteria. The results showed that classical ATP-dependent chaperones, mostly folding chaperones, are widely distributed, although they are sub-represented in some groups. Acidophilic bacteria showed redundancy of genes coding for the ATP-independent holdase chaperones RidA and Hsp20. In addition, a systematically high redundancy of genes encoding periplasmic chaperones like HtrA and YidC was also detected. In the same way, the proteolytic ATPase complexes ClpPX and Lon presented redundancy and broad distribution. The presence of genes that encoded protein variants was noticeable. In addition, genes for chaperones and protease systems were clustered within the genomes, suggesting common regulation of these activities. Finally, some genes were differentially distributed between bacteria as a function of the autotrophic or heterotrophic character of their metabolism. These results suggest that acidophiles possess an abundant and flexible proteostasis network that protects proteins in organisms living in energy-limiting and extreme environmental conditions. Therefore, our results provide a means for understanding the diversity and significance of proteostasis mechanisms in extreme acidophilic bacteria.

Fig 1. Distribution and abundance of genes that code for chaperones and proteases in the genomes of acidophilic bacterial strains.

Absence of circles indicate that encoding genes were not found within the corresponding genome.

Fig 2. Phylogenetic and gene context analysis of RidA from acidophilic bacteria.

A. Phylogenetic tree highlighting the microorganisms that share the same genetic context. B. Conserved ridA context. spoT: ppGpp synthetase II; hyp: hypothetical protein; recG: ATP-dependent DNA helicase; trp: L, D transpeptidase. Phylogenetic analysis was performed by maximum likelihood algorithm as indicated in Methods.

Fig 3. Phylogenetic and gene context analysis of SlyD from acidophilic bacteria.

A. Phylogenetic tree of SlyD B. Genetic context of slyD in genomes from Acidithiobacillia class, and Zeta-proteobacterium M. ferrooxydans O-1. kin: kinase; nuc: nuclease; fer: ferroquelatase; glgA: glycogen synthase, rok: ROK protein; gstA: glutation transferase; msrA: methionine sulfoxide reductase A; msrB: methionine sulfoxide reductase B; hyp: hypothetical protein; pse: pseudouridine synthase; pho: phosphotransferase; spoT: ppGpp synthetase II; red: reductase; dnaJ: co-chaperone protein; dnaK: chaperone protein; grpE: nucleotide exchange factor. Phylogenetic analysis was performed by maximum likelihood algorithm as indicated in Methods.

Fig 4. Phylogenetic and gene context analysis of CnoX from acidophilic bacteria.

A. Phylogenetic tree of CnoX B. Gene context of cnoX and its homologue tfp2. N-lon: N-terminal Lon protease; hyp: hypothetical protein; ycaR: protein YcaR; flhB: flagellar biosynthesis protein; fliR: flagellar biosynthesis protein; leuS: leucine tRNA ligase; ptr: phosphotransferase; folP: dihydropteroate synthase; tra: transferase; akr: aldo-keto reductase; ntrY: nitrogen regulation protein; pol: DNA polymerase; oprB: carbohydrate porin; ybbP: uncharacterized ABC transporter permease; ybbA: putative ABC transporter ATP-binding protein; ybbO: uncharacterized oxidoreductase; fetB: probable iron export permease; fetA: putative iron ABC exporter ATP-binding; htrA: serine protease; xerD: site-specific tyrosine recombinase; do: protease; crpA: protease; groES: chaperone GroES; groEL: chaperone GroEL; tfp2: thioredoxin-fold protein 2; recN: DNA repair protein. Phylogenetic analysis was performed by maximum likelihood algorithm as indicated in Methods.

Fig 5. Phylogenetic and gene context analysis of Hsp31 from acidophilic bacteria.

A. Phylogenetic tree of Hsp31 B. Gene context of hsp31 in acidophilic microorganisms. hst: homoserine transferase; tra: transferase; fmt: methionyl-tRNA formyltransferase; pdt: phosphodiesterase; envC: murein hydrolase activator; ctpA: copper exporting P-type ATPase; prcA: proteosome subunit alpha; clpS: ATP-dependent Clp protease adapter ClpS; clpA: ATP-dependent Clp protease; hst: homoserine transferase; hyp: hypothetical protein; ureG: urease accessory protein UreG; yaiL: DUF2058 domain containing protein; tra: transferase; yagQ: molybdenum cofactor insertion chaperone YagQ; panE: 2-dehydropantoate 2-reductase; thiL: thiamine-phosphate kinase; araC: L-arabinose operon transcriptional regulator; cheW: chemotaxis protein. Phylogenetic analysis was performed by maximum likelihood algorithm as indicated in Methods.

Fig 6. Phylogenetic and gene context analysis of Hsp20 from acidophilic bacteria.

A. Phylogenetic tree of Hsp20 highlighting groups I, II and III. B. Gene context of hsp20 from group I (hsp20.1), group II (hsp20.2) and group III (hsp20.3); adh: alcohol dehydrogenase; hyd: hydrolase; hyp: hypothetical protein treT: trehalose synthase; treZ: malto-oligosyltrehalose trehalohydrolase; treY: maltooligosyl trehalose synthase; syn: synthase; sppA: protease IV; luxR: transcription factor LuxR; csp: cold shock protein; kin: kinase; ABC tra: ABC transporter; rluB: ribosomal large subunit pseudouridine synthase B; orn: oligoribonuclease; yheS: putative ATP-protein YheS; yidC: membrane protein insertase YidC; mnmE: tRNA modification GtPhase MnmE; lpxL: lipid A biosynthesis lauroyl transferase; nadD: nicotinate-nucleotide adenyltransferase; rsfA: ribosomal silencing factor RsfS; ybeA: ribosomal RNA large subunit methyltransferase H; pilZ: flagellar brake protein YcgR; clsA: cardiolipin synthase A; pbp: penicillin binding protein; cbs: putative signal-transduction protein with CBSd: tnp: transposase; sel1: Sel1 repeat family protein; htrA: serine protease HtrA; dhg: dehydrogenase; cutE: apolipoprotein N-acyltransferase; prfB: peptide chain release factor 2; lon: Lon protease; end III: endonuclease III; lig: ligase; sixA: phospohistidine phosphatase SixA; yefM: antitoxin YefM; yoeB: toxin YoeB. Phylogenetic analysis was performed by maximum likelihood algorithm as indicated in Methods.

Fig 7. Schematic representation of predicted proteostasis networks in acidophiles.

The prediction of genetic products was derived from 40 genomic sequences of heterotrophic, autotrophic, and mixotrophic acidophilic microorganisms. Numbers represent the copies detected in every class. Colors represent the corresponding bacterial class.