Comparative sequence analyses were performed with members of the Solanaceae and the Brassicaceae. These studies investigated genomic organization, determined levels of microcolinearity, identified orthologous genes and investigated the molecular basis of trait differences. The first analysis was performed by comparison of tomato (Solanum lycopersicum) genomic sequence (119 kb) containing the JOINTLESS1 (J1) locus with orthologous sequences from two potato species, a diploid, Solanum bulbocastanum (800-900 Mb, 2N=2X=24), and a hexaploid, Solanum demissum (2,700 Mb, 2N=6X=72). Gene colinearity was well maintained across all three regions. Twelve orthologous open reading frames were identified in identical order and orientation and included three putative J1 orthologs with 93-96% amino acid sequence identity in both potato species. Although these regions were highly conserved, several local disruptions were detected and included small-scale expansion/contraction regions with intergenic sequences, non-colinear genes and transposable elements. Three putative Solanaceous-specific genes were also identified in this analysis. The second analysis was performed by comparison of a Thellungiella halophila (T. halophila) genomic sequence (193 kb) containing the SALT OVERLY SENSITIVE1 (SOS1) locus with the orthologous sequence (146 kb) in Arabidopsis thaliana (Arabidopsis). T. halophila is a halophytic relative of Arabidopsis thaliana that exhibits extreme salt tolerance. Twenty-five genes, including the putative T. halophila SOS1 (ThSOS1), showed a high degree of colinearity with Arabidopsis genes in the corresponding region. Although the two sequences were significantly colinear, several local rearrangements were detected which were caused by tandem duplications and inversions. Three major expansion/contraction regions in T. halophila contained five LTR retrotransposons which contributed to genomic size variation in this region. ThSOS1 shares similar gene structure and sequence with Arabidopsis SOS1 (AtSOS1), including 11 transmembrane domains and a cyclic nucleotide-binding domain. Three Simple Sequence Repeats (SSRs) were detected within a 540 bp region upstream of the putative translational start site in ThSOS1. The (CTT)n repeat is present in different copy numbers in ThSOS1 (18 repeats) and AtSOS1 (3 repeats). When present in the 5' UTRs of some Arabidopsis genes, (CTT)n serves as a putative salicylic acid responsive element. These SSRs may serve as cis-acting elements affecting differential mRNA accumulation of SOS1 in the two species.

El trabajo realizado en la Tesis Doctoral llega a las siguientes conclusiones:1.- T. halophila muestra una relación en peso raíz/parte aérea mayor que A. thaliana, y esta diferencia se ve incrementada en condiciones de estrés salino, lo que podría suponer una ventaja para afrontarlo.2.- El estrés salino produce en A. thaliana mayores reducciones en la absorción y en las concentraciones internas de K+ que en T. halophila, a la vez que T. halophila presenta menor absorción de Na+ y transporte a la parte aérea que A. thaliana. Ambas circunstancias resultan en una mayor relación K+/Na+ en T. halophila, lo que puede suponer una mayor tolerancia a la salinidad.3.- El gen ThHAK5 codifica para un transportador que media un transporte de K+ de alta afinidad en levaduras similar al observado en las plantas de T. halophila lo que sugiere que este transportador juega un papel fundamental en la absorción de K+ en el rango de la alta afinidad en esta especie vegetal.4.- Aunque AtHAK5 y ThHAK5 presentan una gran homología de secuencia y unas características funcionales similares, la regulación de los genes que los codifican difieren en condiciones salinas. Así, la salinidad reduce en menor medida la inducción de ThHAK5 por ayuno de K+. En consecuencia, la absorción de K+ de alta afinidad está menos afectada por la presencia de NaCl en el medio externo en T. halophila.5.- La mutagénesis al azar permite encontrar aminoácidos importantes para la función de las proteínas y ésta ha permitido identificar dos versiones mutantes del transportador de K+ de alta afinidad AtHAK5 más eficientes, capaces de transportar K+ a concentraciones externas de Na+ muy elevadas (0.1 mM K+ y 800 mM Na+). / The work done in this Thesis provides some interesting conclusions:1.- Thellungiella halophila show a weight ratio root/shoot bigger than Arabidopsis thaliana, and this difference arise under salt stress, what might provide an effective mechanism of salt tolerance to T. halophila.2.- In A. thaliana, salt stress induces a bigger reduction of K+ uptake and tissue concentrations than in T. halophila, and at the same time T. halophila shows a reduced Na+ uptake and Na+ transport to the shoot. Both properties enable a higher ratio K+/Na+ in T. halophila which might be another mechanism of salt tolerance. 3.- The ThHAK5 gene isolated in this Thesis, encode a K+ transporter that mediates high affinity K+ transport in Saccharomyces cerevisiae similar to the observed in intact plants of T. halophila, which suggest a key role of this transporter in the high affinity range of concentrations.4.- Although AtHAK5 and ThHA5 shows high sequence homology and similar functional properties, gene regulation is different under salt stress. Thus, salinity reduces to a lesser extent the K+-starvation ThHAK5 induction. As a consequence, high affinity K+ uptake is less affected by NaCl in T. halophila. 5.- Random mutagenesis allows the identification of important aminoacids for protein function, and with this technique two more efficient mutant versions of AtHAK5 have been isolated. The evolved AtHAK5 mutant versions are able to transport K+ at high Na+ external concentrations (0.1 mM K+ and 800 mM Na+) in yeast.