Characterization of genomic poly(dT-dG).poly(dC-dA) sequences: structure, organization, and conformation.

Abstract

genomes. We have isolated and characterized TG-elements from different locations in the human genome: from randomly isolated clones, associated with the actin gene family, and linked to another repeated element. The results indicate that the following features are typical of these TGelements: (i) the elements consist of 20 to 60 base pairs of (dT-dG)" * (dC-dA)., (ii) the sequences characterized in our study were not flanked by direct or inverted repeats, (iii) the sequences are interspersed rather than in satellite blocks, (iv) the elements are not usually associated with other repeated elements, and (v) some of the elements are found near coding sequences or in introns. Studies on the conformation of a genomic TG-element in a supercoiled plasmid indicate several distinct properties of the TG-element: (i) it is in the Z-form only at low ionic strength, (ii) S1 nuclease recognizes its Z-form with a marked preference for one of the B-Z junctions, and (iii) the sensitive region extends for 20 base pairs near the B-Z junction. In contrast to the result with the supercoiled plasmid, S1 nuclease failed to recognize the TG-element in minichromosomes.Sequences of alternating purines and pyrimidines such as poly(dG-dC) * poly(dG-dC) and poly(dT-dG) * poly(dC-dA) may assume a left-handed conformation (Z-DNA) under certain conditions (12,22,23,41). Studies with specific antibodies indicate that Z-DNA can be found in acid-fixed chromosomes (2, 20) and supercoiled plasmid and viral DNAs (21, 23), and several interesting biological roles for Z-DNA have been proposed (20,23). Hybridization studies with synthetic poly(dT-dG) * poly(dC-dA) suggest that homologous sequences are abundant and interspersed in the genomes of many, if not all, eucaryotes (8,9). In addition, sequence analysis of various cloned genes occasionally reveals the presence of poly(dT-dG) * poly(dC-dA) in the vicinity of coding regions (e.g., see references 1, 10, 18, 19, 33, 34, and 40). From these data a partial characterization of the structure and organization of this class of poly(dTdG) * poly(dC-dA) sequences has emerged, and on the basis of some of these data it has been proposed that the poly(dTdG) * poly(dC-dA) sequences spread throughout the genome by insertion (26).If the genomic poly(dT-dG) -poly(dC-dA) sequences (TGelements) have a biological function, it is probably because of their conformational peculiarity. The Z-form of poly(dGdC) * poly(dG-dC), another potential Z-DNA sequence, has been extensively studied. The Z-form of poly(dG-dC)-poly(dG-dC) is recognized by specific antibodies (14) and is sensitive to S1 nuclease (31) when in a supercoiled plasmid. The poly(dT-dG) -poly(dC-dA) sequence, when in supercoiled plasmids, also has been shown to be in Z-form (12,22), although its conformational properties are not known in detail. Supercoiled plasmids with genomic TG-elements thus are useful substrates for characterizing the conformational properties of these elements.In this report we present the results of a series of experi-* Corresponding ...

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