Browsing by Author "Salinas, Francisco"
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- ItemClonal Heterogeneity Influences the Fate of New Adaptive Mutations(2017) Vazquez-Garcia, Ignacio; Salinas, Francisco; Li, Jing; Fischer, Andrej; Barre, Benjamin; Hallin, Johan; Bergstrom, Anders; Alonso-Perez, Elisa; Warringer, Jonas; Mustonen, Ville; Liti, GianniThe joint contribution of pre-existing and de novo genetic variation to clonal adaptation is poorly understood but essential to designing successful antimicrobial or cancer therapies. To address this, we evolve genetically diverse populations of budding yeast, S. cerevisiae, consisting of diploid cells with unique haplotype combinations. We study the asexual evolution of these populations under selective inhibition with chemotherapeutic drugs by time-resolved whole-genome sequencing and phenotyping. All populations undergo clonal expansions driven by de novo mutations but remain genetically and phenotypically diverse. The clones exhibit widespread genomic instability, rendering recessive de novo mutations homozygous and refining pre-existing variation. Finally, we decompose the fitness contributions of pre-existing and de novo mutations by creating a large recombinant library of adaptive mutations in an ensemble of genetic backgrounds. Both pre-existing and de novo mutations substantially contribute to fitness, and the relative fitness of preexisting variants sets a selective threshold for new adaptive mutations.
- ItemFungal light-oxygen-voltage domains for optogenetic control of gene expression and flocculation in yeast(2018) Salinas, Francisco; Rojas Jorquera, Vicente Alberto; Delgado Hernández, Verónica Melissa; López Salinas, Javiera C.; Agosin T., Eduardo; Larrondo Castro, Luis Fernando
- ItemIndirect monitoring of TORC1 signalling pathway reveals molecular diversity among different yeast strains(2019) Kessi‐Pérez, Eduardo I.; Salinas, Francisco; Molinet, Jennifer; González, Asier; Muñiz, Sara; Guillamón, José M.; Hall, Michael N.; Larrondo Castro, Luis Fernando; Martínez, Claudio
- ItemInteractions between Core Elements of the Botrytis cinerea Circadian Clock Are Modulated by Light and Different Protein Domains(2022) Rojas, Vicente; Salinas, Francisco; Romero, Andres; Larrondo, Luis F.; Canessa, PauloBotrytis cinerea possesses a complex light-sensing system composed of eleven photoreceptors. In B. cinerea, bcwcl1 encodes for the BcWCL1 protein, the orthologue of the blue-light photoreceptor WC-1 from Neurospora crassa. The functional partner of BcWCL1 is the BcWCL2 protein, both interacting in the nucleus and forming the B. cinerea white collar complex (BcWCC). This complex is required for photomorphogenesis and circadian regulation. However, no molecular evidence shows a light-dependent interaction between the BcWCC components or light-sensing capabilities in BcWCL1. In this work, by employing a yeast two-hybrid system that allows for the in vivo analysis of protein-protein interactions, we confirm that BcWCL1 and BcWCL2 interact in the absence of light as well as upon blue-light stimulation, primarily through their PAS (Per-Arnt-Sim) domains. Deletion of the PAS domains present in BcWCL1 (BcWCL1(PAS increment )) or BcWCL2 (BcWCL2(PAS increment )) severely impairs the interaction between these proteins. Interestingly, the BcWCL1(PAS increment ) protein shows a blue-light response and interacts with BcWCL2 or BcWCL2(PAS increment ) upon light stimulation. Finally, we demonstrate that BcWCL1 and BcWCL1(PAS increment ) respond to blue light by introducing a point mutation in the photoactive cysteine, confirming that both proteins are capable of light sensing. Altogether, the results revealed the complexity of protein-protein interactions occurring between the core elements of the B. cinerea circadian clock.
- ItemModular and Molecular Optimization of a LOV (Light-Oxygen-Voltage)-Based Optogenetic Switch in Yeast(2021) Romero, Andres; Rojas, Vicente; Delgado, Veronica; Salinas, Francisco; Larrondo, Luis F.Optogenetic switches allow light-controlled gene expression with reversible and spatiotemporal resolution. In Saccharomyces cerevisiae, optogenetic tools hold great potential for a variety of metabolic engineering and biotechnology applications. In this work, we report on the modular optimization of the fungal light-oxygen-voltage (FUN-LOV) system, an optogenetic switch based on photoreceptors from the fungus Neurospora crassa. We also describe new switch variants obtained by replacing the Gal4 DNA-binding domain (DBD) of FUN-LOV with nine different DBDs from yeast transcription factors of the zinc cluster family. Among the tested modules, the variant carrying the Hap1p DBD, which we call "HAP-LOV", displayed higher levels of luciferase expression upon induction compared to FUN-LOV. Further, the combination of the Hap1p DBD with either p65 or VP16 activation domains also resulted in higher levels of reporter expression compared to the original switch. Finally, we assessed the effects of the plasmid copy number and promoter strength controlling the expression of the FUN-LOV and HAP-LOV components, and observed that when low-copy plasmids and strong promoters were used, a stronger response was achieved in both systems. Altogether, we describe a new set of blue-light optogenetic switches carrying different protein modules, which expands the available suite of optogenetic tools in yeast and can additionally be applied to other systems.
- ItemNatural Variation in Non-coding Regions Underlying Phenotypic Diversity in Budding Yeast(2016) Salinas, Francisco; De Boer, Carl G.; Abarca, Valentina; García, Verónica; Cuevas, Mara; Araos, Sebastian; Larrondo Castro, Luis Fernando; Martínez, Claudio; Cubillos, Francisco A.
- ItemPowerful decomposition of complex traits in a diploid model(NATURE PUBLISHING GROUP, 2016) Hallin, Johan; Maertens, Kaspar; Young, Alexander I.; Zackrisson, Martin; Salinas, Francisco; Parts, Leopold; Warringer, Jonas; Liti, GianniExplaining trait differences between individuals is a core and challenging aim of life sciences. Here, we introduce a powerful framework for complete decomposition of trait variation into its underlying genetic causes in diploid model organisms. We sequence and systematically pair the recombinant gametes of two intercrossed natural genomes into an array of diploid hybrids with fully assembled and phased genomes, termed Phased Outbred Lines (POLs). We demonstrate the capacity of this approach by partitioning fitness traits of 6,642 Saccharomyces cerevisiae POLs across many environments, achieving near complete trait heritability and precisely estimating additive (73%), dominance (10%), second (7%) and third (1.7%) order epistasis components. We map quantitative trait loci (QTLs) and find nonadditive QTLs to outnumber (3:1) additive loci, dominant contributions to heterosis to outnumber overdominant, and extensive pleiotropy. The POL framework offers the most complete decomposition of diploid traits to date and can be adapted to most model organisms.
- ItemRIM15 antagonistic pleiotropy is responsible for differences in fermentation and stress response kinetics in budding yeast(2016) Kessi Pérez, Eduardo I.; Araos, Sebastián; García, Verónica; Salinas, Francisco; Abarca, Valentina; Larrondo Castro, Luis Fernando; Martínez, Claudio; Cubillos, Francisco A.