Genomic Features for Desiccation Tolerance and Sugar Biosynthesis in the Extremophile Gloeocapsopsis sp. UTEX B3054 The MIT Faculty has made this article openly available. Please share how this access benefits you. Your story matters. Citation Urrejola, Catalina et al. "Genomic Features for Desiccation Tolerance and Sugar Biosynthesis in the Extremophile Gloeocapsopsis sp. UTEX B3054." Frontiers in Microbiology 10 (May 2019): 950 © 2019 Frontiers Media As Published http://dx.doi.org/10.3389/fmicb.2019.00950 Publisher Frontiers Media Version Final published version Citable link https://hdl.handle.net/1721.1/123543 Terms of Use Creative Commons Attribution 4.0 International license Detailed Terms https://creativecommons.org/licenses/by/4.0/ fmicb-10-00950 May 3, 2019 Time: 16:51 # 1 ORIGINAL RESEARCH published: 07 May 2019 doi: 10.3389/fmicb.2019.00950 Genomic Features for Desiccation Tolerance and Sugar Biosynthesis in the Extremophile Gloeocapsopsis sp. UTEX B3054 Catalina Urrejola1, Jaime Alcorta2, Loreto Salas1, Mónica Vásquez3, Martin F. Polz4, Rafael Vicuña1 and Beatriz Díez2* Edited by: 1 Department of Molecular Genetics and Microbiology, Pontificia Universidad Católica de Chile, Santiago, Chile, Dale Anthony Casamatta, 2 Laboratorio de Ecología Microbiana de Sistemas Extremos, Department of Molecular Genetics and Microbiology, University of North Florida, Pontificia Universidad Católica de Chile, Santiago, Chile, 3 Laboratorio de Ecología Microbiana y Toxicología Ambiental, United States Department of Molecular Genetics and Microbiology, Pontificia Universidad Católica de Chile, Santiago, Chile, Reviewed by: 4 Department of Civil and Environmental Engineering, Massachusetts Institute of Technology, Cambridge, Virginie Chapon, MA, United States Commissariat à l’Énergie Atomique et aux Énergies Alternatives (CEA), France For tolerating extreme desiccation, cyanobacteria are known to produce both Chelsea Denise Villanueva, compatible solutes at intracellular level and a copious amount of exopolysaccharides Cleveland State University, as a protective coat. However, these molecules make cyanobacterial cells refractory United States to a broad spectrum of cell disruption methods, hindering genome sequencing, and *Correspondence: Beatriz Díez molecular studies. In fact, few genomes are already available from cyanobacteria [email protected] from extremely desiccated environments such as deserts. In this work, we report the Specialty section: 5.4 Mbp draft genome (with 100% of completeness in 105 contigs) of Gloeocapsopsis This article was submitted to sp. UTEX B3054 (subsection I; Order Chroococcales), a cultivable sugar-rich and Evolutionary and Genomic hardly breakable hypolithic cyanobacterium from the Atacama Desert. Our in silico Microbiology, a section of the journal analyses focused on genomic features related to sugar-biosynthesis and adaptation Frontiers in Microbiology to dryness. Among other findings, screening of Gloeocapsopsis genome revealed a Received: 04 May 2018 unique genetic potential related to the biosynthesis and regulation of compatible solutes Accepted: 15 April 2019 Published: 07 May 2019 and polysaccharides. For instance, our findings showed for the first time a novel Citation: genomic arrangement exclusive of Chroococcaceae cyanobacteria associated with the Urrejola C, Alcorta J, Salas L, recycling of trehalose, a compatible solute involved in desiccation tolerance. Additionally, Vásquez M, Polz MF, Vicuña R and we performed a comparative genome survey and analyses to entirely predict the Díez B (2019) Genomic Features for Desiccation Tolerance and Sugar highly diverse pool of glycosyltransferases enzymes, key players in polysaccharide Biosynthesis in the Extremophile biosynthesis and the formation of a protective coat to dryness. We expect that this Gloeocapsopsis sp. UTEX B3054. Front. Microbiol. 10:950. work will set the fundamental genomic framework for further research on microbial doi: 10.3389/fmicb.2019.00950 tolerance to desiccation and to a wide range of other extreme environmental conditions. Frontiers in Microbiology| www.frontiersin.org 1 May 2019| Volume 10| Article 950 fmicb-10-00950 May 3, 2019 Time: 16:51 # 2 Urrejola et al. Genomics of Desiccation-Tolerant Cyanobacterium The study of microorganisms like Gloeocapsopsis sp. UTEX B3054 will contribute to expand our limited understanding regarding water optimization and molecular mechanisms allowing extremophiles to thrive in xeric environments such as the Atacama Desert. Keywords: Atacama Desert, compatible solutes, cyanobacteria, desiccation tolerance, DNA extraction, exopolysaccharide, glycosyltransferase, trehalose INTRODUCTION Klähn and Hagemann, 2011) and EPS (Grilli Caiola et al., 1993, 1996; Hill et al., 1997; Tamaru et al., 2005; Knowles and The Atacama Desert is the driest warm desert on Earth (Houston Castenholz, 2008; Mager and Thomas, 2011) might play in and Hartley, 2003; Hartley et al., 2005). Located in Northern desiccation tolerance in cyanobacteria. However, comprehensive Chile, for many years it was thought to be a sterile territory, genomic analyses of the mechanisms for tolerating extreme unable to give shelter to any kind of living organism (McKay desiccation in unicellular cyanobacteria are still missing. In that et al., 2003; Navarro-González et al., 2003). We have recently sense, we decided to sequence and to study the genome of learned that in the Atacama Desert the occasional water inputs Gloeocapsopsis sp. UTEX B3054, a unicellular cyanobacterium coming from the coastal fog and dew sustain the scarce microbial that we demonstrate belongs to the Chroococcaceae family, life thriving under its characteristic extreme environmental and which possesses few known cultivable and sequenced conditions (Houston and Hartley, 2003; McKay et al., 2003). representatives. This strain was obtained by cell-sorting from Microbial life has developed a variety of physical and Gloeocapsopsis sp. AAB1 culture, an enrichment initially collected molecular strategies to overcome the high solar radiation and from a quartz rock in the Atacama Desert and described temperatures, as well as to maximize the efficiency in the use to be extremely tolerant to desiccation (Azúa-Bustos et al., of the low amount of water available. Indeed, most microbial 2014). Besides improving the genome coverage of this family life in deserts is somehow associated to rocks, developing either of cyanobacteria, our study aimed to predict and analyze within or underneath them (Chan et al., 2012; Pointing and the genomic mechanisms likely associated to the desiccation Belnap, 2012; Cowan et al., 2014; Davila et al., 2015; Wierzchos tolerance of Gloeocapsopsis sp. UTEX B3054. In particular, et al., 2015). These microbial communities are dominated by we focused on identifying the genetic potential and genomic primary producers, represented mainly by morphological and mechanisms likely involved in the biosynthesis of compatible metabolically diverse cyanobacteria (Pointing et al., 2009; Wong solutes and EPS, molecules that play a key role in microbial et al., 2010; Wierzchos et al., 2015; Wei et al., 2016). Most of these tolerance to dryness. desert cyanobacteria produce copious extracellular structures, a feature that is thought to constitute both the architectural and metabolic basis for the microbial community and its tolerance MATERIALS AND METHODS to extreme environmental conditions (Knowles and Castenholz, 2008; Colica et al., 2014; Rossi and De Philippis, 2015). Strain Isolation and DNA Extraction Specifically, cyanobacteria organized in packet-like structures The strain used in this study, Gloeocapsopsis sp. UTEX B3054, such as Chroococcidiopsis and Gloeocapsa, dominate lithic was obtained from the non-axenic Gloeocapsopsis sp. AAB1 communities found in warm deserts (Warren-Rhodes et al., 2006; enrichment culture initially isolated from the Atacama Desert Bahl et al., 2011; Chan et al., 2012; Wierzchos et al., 2015; (Azúa-Bustos et al., 2014). To massively eliminate contaminant Crits-Christoph et al., 2016). Although these microorganisms heterotrophic bacteria, a single cyanobacterial cell was sorted have been identified by microscopy and by 16S-rDNA surveys, into BG11 media using an Influx Mariner Cell Sorter (Cytopeia, a significant barrier to progress in the study of cyanobacteria in Seattle, WA, United States). Chlorophyll-containing cells were general has been the difficulties encountered in obtaining axenic detected based on red fluorescence (692–40 nm; fluorescence cultures, as well as the presence of copious exopolysaccharide filters are specified here by the wavelength of maximum (EPS) that hinders sequencing of their genomes (Tillett and transmission and spectral width of bandpass) excited with a Neilan, 2000; Chrismas et al., 2016). To illustrate the latter, 488 nm laser, while triggering was based on side light scatter only 1,110 from a total of 76,299 genomes available in the (SSC) to allow the exclusion of non-fluorescent cells. The clone Integrated Microbial Genomes and Microbiomes (IMG/JGI) of Gloeocapsopsis sp. UTEX B3054 culture was deposited in the database correspond to cyanobacteria. Solely the marine genus UTEX Culture Collection of Algae under the accession code Prochlorococcus concentrates a 55.85% of the cyanobacterial UTEX B3054, and it is publicly available. genomes already available. Moreover, merely three genomes Several treatments were further implemented in order to correspond to cyanobacteria isolated from desert environments, mechanically,