@article{wheaton_vitko_counts_dulkis_podolsky_mukherjee_kelly_2019, title={Extremely Thermoacidophilic Metallosphaera Species Mediate Mobilization and Oxidation of Vanadium and Molybdenum Oxides}, volume={85}, ISSN={["1098-5336"]}, DOI={10.1128/AEM.02805-18}, abstractNote={In order to effectively leverage extremely thermoacidophilic archaea for the microbially based solubilization of solid-phase metal substrates (e.g., sulfides and oxides), understanding the mechanisms by which these archaea solubilize metals is important. Physiological analysis of Metallosphaera species growth in the presence of molybdenum and vanadium oxides revealed an indirect mode of metal mobilization, catalyzed by iron cycling. However, since the mobilized metals exist in more than one oxidation state, they could potentially serve directly as energetic substrates. Transcriptomic response to molybdenum and vanadium oxides provided evidence for new biomolecules participating in direct metal biooxidation. The findings expand the knowledge on the physiological versatility of these extremely thermoacidophilic archaea.}, number={5}, journal={APPLIED AND ENVIRONMENTAL MICROBIOLOGY}, author={Wheaton, Garrett H. and Vitko, Nicholas P. and Counts, James A. and Dulkis, Jessica A. and Podolsky, Igor and Mukherjee, Arpan and Kelly, Robert M.}, year={2019}, month={Mar} }
 @article{mukherjee_wheaton_counts_ijeomah_desai_kelly_2017, title={VapC toxins drive cellular dormancy under uranium stress for the extreme thermoacidophile Metallosphaera prunae}, volume={19}, ISSN={["1462-2920"]}, DOI={10.1111/1462-2920.13808}, abstractNote={When abruptly exposed to toxic levels of hexavalent uranium, the extremely thermoacidophilic archaeon Metallosphaera prunae, originally isolated from an abandoned uranium mine, ceased to grow, and concomitantly exhibited heightened levels of cytosolic ribonuclease activity that corresponded to substantial degradation of cellular RNA. The M. prunae transcriptome during ‘uranium-shock’ implicated VapC toxins as possible causative agents of the observed RNA degradation. Identifiable VapC toxins and PIN-domain proteins encoded in the M. prunae genome were produced and characterized, three of which (VapC4, VapC7, VapC8) substantially degraded M. prunae rRNA in vitro. RNA cleavage specificity for these VapCs mapped to motifs within M. prunae rRNA. Furthermore, based on frequency of cleavage sequences, putative target mRNAs for these VapCs were identified; these were closely associated with translation, transcription, and replication. It is interesting to note that Metallosphaera sedula, a member of the same genus and which has a nearly identical genome sequence but not isolated from a uranium-rich biotope, showed no evidence of dormancy when exposed to this metal. M. prunae utilizes VapC toxins for post-transcriptional regulation under uranium stress to enter a cellular dormant state, thereby providing an adaptive response to what would otherwise be a deleterious environmental perturbation.}, number={7}, journal={ENVIRONMENTAL MICROBIOLOGY}, author={Mukherjee, Arpan and Wheaton, Garrett H. and Counts, James A. and Ijeomah, Brenda and Desai, Jigar and Kelly, Robert M.}, year={2017}, month={Jul}, pages={2831–2842} }
 @article{wheaton_mukherjee_kelly_2016, title={Transcriptomes of the Extremely Thermoacidophilic Archaeon Metallosphaera sedula Exposed to Metal "Shock" Reveal Generic and Specific Metal Responses}, volume={82}, ISSN={["1098-5336"]}, DOI={10.1128/aem.01176-16}, abstractNote={ABSTRACT The extremely thermoacidophilic archaeon Metallosphaera sedula mobilizes metals by novel membrane-associated oxidase clusters and, consequently, requires metal resistance strategies. This issue was examined by “shocking” M. sedula with representative metals (Co 2+ , Cu 2+ , Ni 2+ , UO 2 2+ , Zn 2+ ) at inhibitory and subinhibitory levels. Collectively, one-quarter of the genome (554 open reading frames [ORFs]) responded to inhibitory levels, and two-thirds (354) of the ORFs were responsive to a single metal. Cu 2+ (259 ORFs, 106 Cu 2+ -specific ORFs) and Zn 2+ (262 ORFs, 131 Zn 2+ -specific ORFs) triggered the largest responses, followed by UO 2 2+ (187 ORFs, 91 UO 2 2+ -specific ORFs), Ni 2+ (93 ORFs, 25 Ni 2+ -specific ORFs), and Co 2+ (61 ORFs, 1 Co 2+ -specific ORF). While one-third of the metal-responsive ORFs are annotated as encoding hypothetical proteins, metal challenge also impacted ORFs responsible for identifiable processes related to the cell cycle, DNA repair, and oxidative stress. Surprisingly, there were only 30 ORFs that responded to at least four metals, and 10 of these responded to all five metals. This core transcriptome indicated induction of Fe-S cluster assembly (Msed_1656-Msed_1657), tungsten/molybdenum transport (Msed_1780-Msed_1781), and decreased central metabolism. Not surprisingly, a metal-translocating P-type ATPase (Msed_0490) associated with a copper resistance system (Cop) was upregulated in response to Cu 2+ (6-fold) but also in response to UO 2 2+ (4-fold) and Zn 2+ (9-fold). Cu 2+ challenge uniquely induced assimilatory sulfur metabolism for cysteine biosynthesis, suggesting a role for this amino acid in Cu 2+ resistance or issues in sulfur metabolism. The results indicate that M. sedula employs a range of physiological and biochemical responses to metal challenge, many of which are specific to a single metal and involve proteins with yet unassigned or definitive functions. IMPORTANCE The mechanisms by which extremely thermoacidophilic archaea resist and are negatively impacted by metals encountered in their natural environments are important to understand so that technologies such as bioleaching, which leverage microbially based conversion of insoluble metal sulfides to soluble species, can be improved. Transcriptomic analysis of the cellular response to metal challenge provided both global and specific insights into how these novel microorganisms negotiate metal toxicity in natural and technological settings. As genetics tools are further developed and implemented for extreme thermoacidophiles, information about metal toxicity and resistance can be leveraged to create metabolically engineered strains with improved bioleaching characteristics.}, number={15}, journal={APPLIED AND ENVIRONMENTAL MICROBIOLOGY}, author={Wheaton, Garrett H. and Mukherjee, Arpan and Kelly, Robert M.}, year={2016}, month={Aug}, pages={4613–4627} }
 @misc{wheaton_counts_mukherjee_kruh_kelly_2015, title={The Confluence of Heavy Metal Biooxidation and Heavy Metal Resistance: Implications for Bioleaching by Extreme Thermoacidophiles}, volume={5}, ISSN={["2075-163X"]}, DOI={10.3390/min5030397}, abstractNote={Extreme thermoacidophiles (Topt > 65 °C, pHopt < 3.5) inhabit unique environments fraught with challenges, including extremely high temperatures, low pH, as well as high levels of soluble metal species. In fact, certain members of this group thrive by metabolizing heavy metals, creating a dynamic equilibrium between biooxidation to meet bioenergetic needs and mechanisms for tolerating and resisting the toxic effects of solubilized metals. Extremely thermoacidophilic archaea dominate bioleaching operations at elevated temperatures and have been considered for processing certain mineral types (e.g., chalcopyrite), some of which are recalcitrant to their mesophilic counterparts. A key issue to consider, in addition to temperature and pH, is the extent to which solid phase heavy metals are solubilized and the concomitant impact of these mobilized metals on the microorganism’s growth physiology. Here, extreme thermoacidophiles are examined from the perspectives of biodiversity, heavy metal biooxidation, metal resistance mechanisms, microbe-solid interactions, and application of these archaea in biomining operations.}, number={3}, journal={MINERALS}, author={Wheaton, Garrett and Counts, James and Mukherjee, Arpan and Kruh, Jessica and Kelly, Robert}, year={2015}, month={Sep}, pages={397–451} }
 @article{mccarthy_ai_wheaton_tevatia_eckrich_kelly_blum_2014, title={Role of an Archaeal PitA Transporter in the Copper and Arsenic Resistance of Metallosphaera sedula, an Extreme Thermoacidophile}, volume={196}, ISSN={["1098-5530"]}, DOI={10.1128/jb.01707-14}, abstractNote={Thermoacidophilic archaea, such as Metallosphaera sedula, are lithoautotrophs that occupy metal-rich environments. In previous studies, an M. sedula mutant lacking the primary copper efflux transporter, CopA, became copper sensitive. In contrast, the basis for supranormal copper resistance remained unclear in the spontaneous M. sedula mutant, CuR1. Here, transcriptomic analysis of copper-shocked cultures indicated that CuR1 had a unique regulatory response to metal challenge corresponding to the upregulation of 55 genes. Genome resequencing identified 17 confirmed mutations unique to CuR1 that were likely to change gene function. Of these, 12 mapped to genes with annotated function associated with transcription, metabolism, or transport. These mutations included 7 nonsynonymous substitutions, 4 insertions, and 1 deletion. One of the insertion mutations mapped to pseudogene Msed_1517 and extended its reading frame an additional 209 amino acids. The extended mutant allele was identified as a homolog of Pho4, a family of phosphate symporters that includes the bacterial PitA proteins. Orthologs of this allele were apparent in related extremely thermoacidophilic species, suggesting M. sedula naturally lacked this gene. Phosphate transport studies combined with physiologic analysis demonstrated M. sedula PitA was a low-affinity, high-velocity secondary transporter implicated in copper resistance and arsenate sensitivity. Genetic analysis demonstrated that spontaneous arsenate-resistant mutants derived from CuR1 all underwent mutation in pitA and nonselectively became copper sensitive. Taken together, these results point to archaeal PitA as a key requirement for the increased metal resistance of strain CuR1 and its accelerated capacity for copper bioleaching.}, number={20}, journal={JOURNAL OF BACTERIOLOGY}, author={McCarthy, Samuel and Ai, Chenbing and Wheaton, Garrett and Tevatia, Rahul and Eckrich, Valerie and Kelly, Robert and Blum, Paul}, year={2014}, month={Oct}, pages={3562–3570} }
 @article{mukherjee_wheaton_blum_kelly_2012, title={Uranium extremophily is an adaptive, rather than intrinsic, feature for extremely thermoacidophilic Metallosphaera species}, volume={109}, ISSN={["0027-8424"]}, DOI={10.1073/pnas.1210904109}, abstractNote={Thermoacidophilic archaea are found in heavy metal-rich environments, and, in some cases, these microorganisms are causative agents of metal mobilization through cellular processes related to their bioenergetics. Given the nature of their habitats, these microorganisms must deal with the potentially toxic effect of heavy metals. Here, we show that two thermoacidophilic Metallosphaera species with nearly identical (99.99%) genomes differed significantly in their sensitivity and reactivity to uranium (U). Metallosphaera prunae, isolated from a smoldering heap on a uranium mine in Thüringen, Germany, could be viewed as a "spontaneous mutant" of Metallosphaera sedula, an isolate from Pisciarelli Solfatara near Naples. Metallosphaera prunae tolerated triuranium octaoxide (U(3)O(8)) and soluble uranium [U(VI)] to a much greater extent than M. sedula. Within 15 min following exposure to "U(VI) shock," M. sedula, and not M. prunae, exhibited transcriptomic features associated with severe stress response. Furthermore, within 15 min post-U(VI) shock, M. prunae, and not M. sedula, showed evidence of substantial degradation of cellular RNA, suggesting that transcriptional and translational processes were aborted as a dynamic mechanism for resisting U toxicity; by 60 min post-U(VI) shock, RNA integrity in M. prunae recovered, and known modes for heavy metal resistance were activated. In addition, M. sedula rapidly oxidized solid U(3)O(8) to soluble U(VI) for bioenergetic purposes, a chemolithoautotrophic feature not previously reported. M. prunae, however, did not solubilize solid U(3)O(8) to any significant extent, thereby not exacerbating U(VI) toxicity. These results point to uranium extremophily as an adaptive, rather than intrinsic, feature for Metallosphaera species, driven by environmental factors.}, number={41}, journal={PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA}, author={Mukherjee, Arpan and Wheaton, Garrett H. and Blum, Paul H. and Kelly, Robert M.}, year={2012}, month={Oct}, pages={16702–16707} }