Browsing by Author "Vargas-Chacoff, Luis"

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    Discovery of BbX transcription factor in the patagonian blennie: Exploring expression changes following combined bacterial and thermal stress exposure
    (2023) Martinez, Danixa; Nualart, Daniela; Loncoman, Carlos; Opazo, Juan C.; Zabala, Kattina; Morera, Francisco J.; Mardones, Gonzalo A.; Vargas-Chacoff, Luis
    High-Mobility Group (HMG) proteins are involved in different processes such as transcription, replication, DNA repair, and immune response. The role of HMG proteins in the immune response of fish has been studied mainly for HMGB1, where its expression can be induced by the stimulation of viral/bacterial PAMPs and can act as a proinflammatory mediator and as a global regulator of transcription in response to temperature. However, for BbX this role remains to be discovered. In this work, we identified the BbX of E. maclovinus and evaluated the temporal expression levels after simultaneous challenge with P. salmonis and thermal stress. Phylogenetic analysis does not significantly deviate from the expected organismal relationships suggesting orthologous relationships and that BbX was present in the common ancestor of the group. BbX mRNA expression levels were very high in the intestinal tissue of E. maclovinus (foregut, midgut, and hindgut). Nevertheless, the protein levels analyzed by WB showed the highest levels of BbX protein in the liver (constitutive expression). On the other hand, the mRNA expression levels of BbX in the liver of E. maclovinus injected with P. salmonis and subjected to thermal stress showed an increase at days 16 and 20 in all treatments applied at 12 degrees C and 18 degrees C. Meanwhile, the protein levels quantified by WB showed a statistically significant increase in the HMG-Bbx at all experimental times (4, 8, 12, 16, and 20 dpi). However, at 4 dpi the HMG-Bbx protein levels were much higher than the other days evaluated. The results suggest that BbX protein may be implicated in the response mechanism to temper-ature and bacterial stimulation in the foregut, midgut, hindgut, and liver, according to our findings at the level of mRNA and protein. Furthermore, our WB analysis suggests an effect of P. salmonis on the expression of this protein that can be observed in condition C+ 12 degrees C compared to C- 12 degrees C. Then, there is an effect of temperature that can be evidenced in the condition AM 18 degrees C and SM 18 degrees C, compared to AB 18 degrees C and SB 18 degrees C at 4, 8, and 12 dpi. We found not differences in the levels of this protein if the thermal stress is achieved through acclimatization or shock. More research is necessary to clarify the importance of this type of HMG in the immune response and thermal tolerance in fish.
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    How Many Sirtuin Genes Are Out There? Evolution of Sirtuin Genes in Vertebrates With a Description of a New Family Member
    (2023) Opazo, Juan Carlos; Vandewege, Michael W.; Hoffmann, Federico G.; Zavala, Kattina; Melendez, Catalina; Luchsinger, Catalina; Cavieres, Viviana A.; Vargas-Chacoff, Luis; Morera, Francisco J.; Burgos Hitschfeld, Patricia Verónica; Tapia-Rojas, Cheril; Mardones, Gonzalo A.
    Studying the evolutionary history of gene families is a challenging and exciting task with a wide range of implications. In addition to exploring fundamental questions about the origin and evolution of genes, disentangling their evolution is also critical to those who do functional/structural studies to allow a deeper and more precise interpretation of their results in an evolutionary context. The sirtuin gene family is a group of genes that are involved in a variety of biological functions mostly related to aging. Their duplicative history is an open question, as well as the definition of the repertoire of sirtuin genes among vertebrates. Our results show a well-resolved phylogeny that represents an improvement in our understanding of the duplicative history of the sirtuin gene family. We identified a new sirtuin gene family member (SIRT3.2) that was apparently lost in the last common ancestor of amniotes but retained in all other groups of jawed vertebrates. According to our experimental analyses, elephant shark SIRT3.2 protein is located in mitochondria, the overexpression of which leads to an increase in cellular levels of ATP. Moreover, in vitro analysis demonstrated that it has deacetylase activity being modulated in a similar way to mammalian SIRT3. Our results indicate that there are at least eight sirtuin paralogs among vertebrates and that all of them can be traced back to the last common ancestor of the group that existed between 676 and 615 millions of years ago.
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    Revealing the Salmo salar NLRP3 Inflammasome: Insights from Structural Modeling and Transcriptome Analysis
    (MDPI, 2023) Acevedo, Waldo; Moran-Figueroa, Rodrigo; Vargas-Chacoff, Luis; Morera Galleguillos, Francisco Javier; Pontigo, Juan Pablo
    The NLRP3, one of the most heavily studied inflammasome-related proteins in mammals, remains inadequately characterized in Atlantic salmon (Salmo salar), despite the significant commercial importance of this salmonid. The NLRP3 inflammasome is composed of the NLRP3 protein, which is associated with procaspase-1 via an adapter molecule known as ASC. This work aims to characterize the Salmo salar NLRP3 inflammasome through in silico structural modeling, functional transcript expression determination in the SHK-1 cell line in vitro, and a transcriptome analysis on Atlantic salmon. The molecular docking results suggested a similar arrangement of the ternary complex between NLRP3, ASC, and caspase-1 in both the Atlantic salmon and the mammalian NLRP3 inflammasomes. Moreover, the expression results confirmed the functionality of the SsNLRP3 inflammasome in the SHK-1 cells, as evidenced by the lipopolysaccharide-induced increase in the transcription of genes involved in inflammasome activation, including ASC and NLRP3. Additionally, the transcriptome results revealed that most of the inflammasome-related genes, including ASC, NLRP3, and caspase-1, were down-regulated in the Atlantic salmon following its adaptation to seawater (also known as parr-smolt transformation). This is correlated with a temporary detrimental effected on the immune system. Collectively, these findings offer novel insights into the evolutionarily conserved role of NLRP3.