Antibodies which are specific to the Z-DNA conformation have been purified and characterized on the basis of their binding to three different DNA polymers which can form this left-handed helix. These antibodies bind specifically to polytene chromosomes of Drosophila melanogaster as visualized by fluorescent staining. The staining is found in the interband regions and its intensity varies among different interbands in a reproducible manner. This is the first identification of the Z-DNA conformation in material of biological origin.

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The construction of knotted topologies is a key goal of stereochemistry. In order to measure the chiral properties of knotted molecules, it is necessary to produce both enantiomers of a knot from the same molecule. A molecule containing the same backbone structure that is an amphichiral knot can provide a useful control molecule for such measurements. In the case Of molecules with chiral backbones, configurational chirality, exclusive of the chirality due to knotting, must be measured from the circle of the same sequence. Trefoil knots of both chiralities, an amphichiral knot, and an unknotted circular molecule have all been constructed by enzymatic closute of the same linear DNA molecule. The molecule contains two double helical domains that can be induced to assume the righthanded B conformation or the left-handed Z conformation under selected solution conditions. The molecules expected to contain left-handed DNA have been shown to bind an anti-Z-DNA antibody in gel-retention assays.Knotted topologies have a long history in both biology1-5 and chemistry,6-9 but it is only recently that it has been possible to construct particular knotted molecules. Sauvage and his colleagues have reported the synthesis of a mixture of both chiralities of trefoil knots10'11 from small molecules, and we have reported the synthesis of trefoil12-14 and figure-815 knots from single-stranded DNA. The physical properties of knots are of great interest.16 However, they are not readily available from chemical species, because of the difficulties associated with their syntheses. In order to use physical means to probe chirality resulting from knots of opposite handedness, it is necessary to produce and isolate both enantiomers of the same molecule. Liang and Mislow17 have used the term amphicheiral, to describe knots that are topologically achiral; in discussing knots constructed from chiral components (D-nucleotides), we use the term "cheirality" here to describe the chirality that refers

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Idiotypic cross-reactions were evaluated in 60 polynucleotide-binding monoclonal lupus autoantibodies produced by human-human hybridomas that were derived from seven unrelated patients with SLE. Three antiidiotype reagents were prepared by immunization of rabbits or a mouse with monoclonal autoantibodies from two patients. Binding of the three reagents to their corresponding idiotypes was inhibited by one or more polynucleotides, an indication that the antiidiotypes reacted with the variable regions of the autoantibodies. Each antiidiotype appeared to detect a different idiotypic determinant. Of the 60 monoclonal autoantibodies tested, 40 reacted in one or more competitive immunoassays; 15 reacted with one antiidiotype, 10 reacted with two antiidiotypes and 15 reacted with three antiidiotypes. A monoclonal antiidiotype reagent cross-reacted with autoantibodies from six of the seven patients. The idiotypic cross-reactions of immunoglobulins from unrelated patients suggest that the autoantibodies are derived from related families of germ line genes that are expressed by patients with SLE.

Idiotype (Id) 16/6 marks a variable (V) region structure that occurs frequently in the human immunoglobulin repertoire. The basis of the Id has been traced to a germline heavy chain gene segment, V.18/2 (V.26). To pursue the molecular basis for the frequency of Id 16/6, we have analyzed polymerase chain reaction-generated C/z, C% and V.3 family V gene libraries derived from the circulating and tonsillar B cells of four normal individuals and from the B cells of two patients with active systemic lupus erythematosus (SLE). The frequency of V.18/2 in these libraries was compared with three control V. genes, V.56P1, V.21/28, and V.A57. Plaque lifts from C/z and C3' V. cDNA libraries were screened with gene-specific oligonucleotide probes. The frequency of V.18/2 ranged from 4 to 10% ofJ. + plaques (two to five times that of control V. genes). In four V.3 family-specific libraries derived from rearranged DNA, V.18/2 represented 19-33% of V.3 + plaques. Hybridizing V.18/2 plaques were 98-100% homologous to the germline V. gene; mutations when present were often in framework 3. Extensive variation was seen in the complementarity determining region 3 sequences of these rearranged V genes. The high frequency of V.18/2 expression in the B cell repertoire was confirmed by sequencing randomly picked J.+ plaques. In two patients with active SLE the frequency of use of V.18/2 was not greater than that observed in normal subjects. These results show that V.18/2 is overrepresented in the B cell repertoire of normal subjects and suggest that the immune repertoire may be dominated by relatively few V genes.

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 ...

Abstract. Sera from 55 patients with systemic lupus erythematosus were studied to clarify the significance of the patterns of nuclear fluorescence observed. The sera in which the IgG fraction produced a peripheral pattern of nuclear fluorescence were found to contain complement-fixing antibodies to native DNA and to DNA-histone complexes. This correlation did not exist when complement-fixing activity was compared to the IgM nuclear patterns. Sera which contained only complement-fixing antibodies to denatured DNA and which did not react with native DNA or nucleoprotein did not produce the peripheral pattern of nuclear fluorescence. The data suggest that single strands of DNA were not the reactive groups in the nucleus responsible for the peripheral pattern. The results support the conclusion that DNA within a DNA-protein complex may be the nuclear antigen responsible for the peripheral pattern of nuclear fluorescence.Analysis of the clinical data revealed that a close correlation existed between the presence of IgG peripheral pattern, complement-fixing antibodies to DNA and histone-DNA complexes, and clinical manifestation of active disease. IntroductionThe presence of antinuclear antibodies in sera from patients with systemic lupus erythematosus (SLE) has been described by many investigators (1-4). These antibodies have been shown to be-