RESUMEN
Different superstructural features of the upstream regulatory regions of the two pea genes rbcS-3A and rbcS-E9 have been derived using a theoretical method, developed in our laboratory a few years ago, which evaluates the DNA distortion energy from a matrix of the deviations of the 16 possible dinucleotides from the standard B conformation. The theoretical analysis, which predicts different flexibilities of the regulatory regions of the two genes, is in satisfactorily good agreement with experimental evaluations from gel electrophoretic mobility and cyclization kinetics, suggesting a possible model to explain the largely different transcription efficiencies of the two genes.
Asunto(s)
ADN/química , Fabaceae/genética , Plantas Medicinales , Secuencias Reguladoras de Ácidos Nucleicos , Ribulosa-Bifosfato Carboxilasa/genética , Secuencia de Bases , Cinética , Datos de Secuencia Molecular , Conformación de Ácido NucleicoRESUMEN
Two light responsive elements (LREs), DNA sequences, 62 base pairs long, relevant to the light control during transcription of the pea genes rbcS-3A and rbcS-3.6, were synthesized and ligated to obtain multimers with defined superstructural features. Their gel electrophoretic mobilities were studied in the presence of the tetracation, spermine, since it was previously suggested, on the basis of theoretical analysis, that spermine can increase DNA bending and thus could be useful in revealing DNA superstructural features. In fact, the difference between the curvatures of the two LREs, derived from gel electrophoresis retardation ratios, increases in the presence of spermine. Circular dichroism spectra of the complexes between spermine and the two LREs, at different neutralization ratios, show that the polyamine is able to induce the formation of asymmetric arrangements of complexes of molecules. The chirality of these complexes appears dramatically different for the two LREs, suggesting that their different superstructural features give rise to different interactions with the polyamine.
Asunto(s)
ADN/química , Fabaceae/genética , Genes de Plantas/efectos de los fármacos , Plantas Medicinales , Ribulosa-Bifosfato Carboxilasa/genética , Espermina/farmacología , Animales , Secuencia de Bases , Dicroismo Circular , ADN/genética , ADN/aislamiento & purificación , Electroforesis en Gel de Poliacrilamida/métodos , Genes de Plantas/efectos de la radiación , Luz , Datos de Secuencia Molecular , Conformación de Ácido Nucleico , Oligodesoxirribonucleótidos , Homología de Secuencia de Ácido NucleicoRESUMEN
Analyses of the DNA sequences, named Light Responsive Elements (LREs), relevant in the photoactivation of the two pea genes rbcS-3A and rbcS-3.6, encoding the same protein but differently expressed, have been carried out, taking advantage of two complementary methods; a theoretical analysis, based on conformational energy calculations, and an experimental evaluation of LREs curvature, derived from gel electrophoretic mobilities of multimers of the LREs and of oligonucleotides corresponding to the three boxes in which they can be dissected. Theoretical and experimental analyses show that the curvature of the rbcS-3A LRE is larger than that of rbcS-3.6 LRE, and seems to be correlated with rbcS-3A higher transcription efficiency.
Asunto(s)
Fabaceae/genética , Genes de Plantas , Plantas Medicinales , Ribulosa-Bifosfato Carboxilasa/genética , Secuencia de Bases , Fabaceae/enzimología , Genes de Plantas/efectos de la radiación , Luz , Datos de Secuencia Molecular , Conformación de Ácido Nucleico , Oligodesoxirribonucleótidos/síntesis químicaRESUMEN
Superstructural features of the upstream regulative regions of the five pea rbcS genes have been studied, using theoretical curvature profiles and electrophoretic properties of synthetic oligonucleotides as models of the DNA curved regions. The curvatures derived from both methods are in good agreement and seem correlated with the different transcription efficiency of the five genes.