Publication:
Physiological and transcriptome-based profile of glycosyl hydrolases from the Halophilic archaeon Halogeometricum borinquense
Physiological and transcriptome-based profile of glycosyl hydrolases from the Halophilic archaeon Halogeometricum borinquense
dc.contributor.advisor | Montalvo-Rodríguez, Rafael R. | |
dc.contributor.author | Cuebas-Irizarry, Mara F. | |
dc.contributor.college | College of Arts and Sciences - Sciences | en_US |
dc.contributor.committee | Siritunga, Dimuth | |
dc.contributor.committee | Rodríguez-Minguela, Carlos | |
dc.contributor.department | Department of Biology | en_US |
dc.contributor.representative | Collins, Dana | |
dc.date.accessioned | 2019-03-01T12:10:03Z | |
dc.date.available | 2019-03-01T12:10:03Z | |
dc.date.issued | 2017 | |
dc.description.abstract | Since the discovery of the Archaea domain, a lot of interest has been given to study regulation of gene expression in these microorganisms. However, few reports are available that provides knowledge about how essential pathways can occur in extreme conditions. For halophilic archaea, little is known about genes involved in carbohydrate metabolism and how they are regulated. Therefore, the interest in this research project is to understand the function of glycosyl hydrolases involved in carbohydrate utilization at high salinity. To achieve this goal, the haloarchaeon Halogeometricum borinquense was selected as model since it has shown shorter generation times than other haloarchaea in different media. Also, H. borinquense is able to use glucose, mannose, fructose, xylose, maltose, trehalose, cellobiose, raffinose and glycerol as carbon sources. Cells of H. borinquense were grown in rich media, and minimal media containing three different sugars as its only carbon source: 0.4% (w/v) glucose, 0.4% (w/v) sucrose and 0.6% (w/v) maltose. Enzymatic assays for three different glycosyl hydrolases (α-glucosidase, β-glycosidase and β-galactosidase) were performed under each condition. The results were expressed in fold-difference or how much more or less was the activity in one condition versus another. A decrease in fold-difference in rich media was observed in response to glucose showing that glycosyl hydrolases are less expressed in rich media than glucose. Regarding to maltose and sucrose fold difference for each enzyme respectively was: α-glucosidase’s, 2-, 3- fold; β-glycosidase, 2-, 4- fold; and for β-galactosidase, 6-, 7- fold higher when compared to glucose enzyme levels. Molecular analyses through RNA-seq were performed identifying 39 putative glycosyl hydrolases, and revealed evidence that these enzymes were transcribed differentially as judged by differences across the transcriptome under distinct growth conditions. This investigation remarks the first attempt to address the topic of carbon catabolite repression in hypersaline environments. Specifically, demonstrating that this phenomenon is performed differently than bacterial and eukaryotic organisms, a comparison based on the sugar preference acting as a repressor. Is suggested the yeast extract as a potential repressor in halophilic archaea, such as in thermophilic archaea. In addition to. here is the first transcriptome analysis in H. borinquense which could lead future studies with this organism as model for gene expression and regulation in haloarchaea. | en_US |
dc.description.abstract | Desde el descubrimiento del dominio Archaea, se le hadado mucho interés a estudiar la regulación de expresión de genes en estos microorganismos. Sin embargo, pocos reportes están disponibles donde se provea conocimiento acerca de procesos esenciales que se lleven a cabo en condiciones extremas. En cuanto a arqueas halofílicas, poco se sabe acerca de los genes involucrados en el metabolismo de carbohidratos y cómo se regulan. Por esto, nuestro interés es entender la función de las glicolasas hidrolíticas involucradas en la utilización de carbohidratos a alta salinidad. Para llevar a cabo este objetivo, la arquea halofílica Halogeometricum borinquense fue seleccionada como modelo, ya que tiene un tiempo de generación más corto que otras arqueas halofílicas in diferentes medios. También H. borinquense tiene la habilidad de usar glucosa, manosa, fructosa, xilosa, y glicerol como fuentes de carbono. Las células de H. borinquense fueron crecidas en medio rico, y medio mínimo con tres azúcares distintas como su única fuente de carbono: 0.4% (v/w) glucosa, 0.4% (v/w) sacarosa y 0.6% (v/w) maltosa. Ensayos enzimáticos para tres glicolasas hidrolíticas fueron realizados en cada condición. Una disminución en la diferencia en el doblez de actividad enzimática en medio rico comparado con glucosa. En maltosa y sacarosa, la diferencia en doblez para cada enzima respectivamente fue: α-glucosidasa 2-, 3- dobleces; β-glycosidasa, 2-, 4- dobleces; and for β-galactosidasa, 6-, 7- dobleces más que cuando se compara con los niveles enzimáticos de glucosa. Se realizó un análisis molecular a través de “RNA-seq” donde se identificaron 39 posibles glicolasas hidrolíticas y se mostró evidencia de que esas enzimas eran diferentes a nivel de transcripcional, dado sus diferencias en el transcriptoma en las distintas condiciones de crecimiento. Esta investigación resalta el primer intento de estudiar represión catabólica de carbono en ambientes hipersalinos. Específicamente, demostrando que este fenómeno se lleva a cabo diferente a organismos bacterianos y eucariotas. Esta comparación se basa en la preferencia de azúcar que a su vez actúa como represor. Se sugiere que el extracto de levadura actúa como un represor potencial para arqueas halofílicas, así como también sucede en arqueas termofílicas. De manera que se recomienda seguir investigando estas enzimas para entender su function en distintas fuentes de carbono. Finalmente, dado que este es el primer análisis de transcriptoma en H. borinquense, se podría utilice para realizar estudios futuros como organism modelo de expresión y regulación de genes en arqueas halofílicas. | en_US |
dc.description.graduationSemester | Spring | en_US |
dc.description.graduationYear | 2017 | en_US |
dc.description.sponsorship | Howard Hughes Medical Institute (HHMI) “Enhancing Advanced Educational Opportunities in STEM Fields for Minority Students at UPRM” Grant Number 52007566; National Science Foundation Award number: 1517797 “Collaborative Research: Chromatin Modification in Archaea and its Role in Gene Expression” | en_US |
dc.identifier.uri | https://hdl.handle.net/20.500.11801/1816 | |
dc.language.iso | en | en_US |
dc.rights.holder | (c) 2017 Mara F. Cuebas-Irizarry | en_US |
dc.rights.license | All rights reserved | en_US |
dc.subject | Glycosyl hydrolases | en_US |
dc.subject | Halogeometricum borinquense | en_US |
dc.subject.lcsh | Archaobacteria | en_US |
dc.subject.lcsh | Halophilic microorganisms | en_US |
dc.subject.lcsh | Microbial genetics | en_US |
dc.subject.lcsh | Glycosidares | en_US |
dc.subject.lcsh | Archaebacteria -- Metabolism | en_US |
dc.subject.lcsh | Halobacterium | en_US |
dc.title | Physiological and transcriptome-based profile of glycosyl hydrolases from the Halophilic archaeon Halogeometricum borinquense | en_US |
dc.type | Thesis | en_US |
dspace.entity.type | Publication | |
thesis.degree.discipline | Biology | en_US |
thesis.degree.level | M.S. | en_US |
Files
Original bundle
1 - 1 of 1
No Thumbnail Available
- Name:
- BIOL_CuebasIrizarryM_2017.pdf
- Size:
- 2.58 MB
- Format:
- Adobe Portable Document Format
- Description:
License bundle
1 - 1 of 1
No Thumbnail Available
- Name:
- license.txt
- Size:
- 1.64 KB
- Format:
- Item-specific license agreed upon to submission
- Description: