Functional Dissection of 5′ and 3′ Extragenic Control Regions of Human tRNAval Genes Reveals Two Different Regulatory Effects

ABSTRACT Two natural human tRNAVal genes, pHtV1 and pHtV3, differ in their transcription efficiency by an order of magnitude. The extragenic control regions (ECRs) responsible for this effect were compared with respect to the kinetics and thermodynamics of transcription complex formation. The 5′ ECR of pHtV1 acts by increasing both the rate of stable complex formation and the equilibrium constant of association between tDNA and at least one transcription factor present in the stable complex. The stability of the preinitiation complexes is not affected by ECRs. For the formation of a stable preinitiation complex, we suggest a two-step mechanism, comprising (i) the ECR-controlled association of at least one transcription factor (TFIIIC) with the tDNA, and (ii) an ECR-independent conformational change of this tDNA–protein complex. The function of 3′ ECRs could be discriminated from the 5′ ECR-mediated effects by transcriptional analysis of two chimeric constructs derived from pHtV1 and pHtV3. Surprisingly, t...