Previously we have demonstrated that the U2 snRNA genes from the higher plant Arabidopsis thaliana contain two upstream elements, the USE with sequence RTCCCACATCG and a -30 'TATA' box, which are essential for transcription by RNA polymerase II, and that the conserved spacing of about four helical DNA turns between these elements is important for optimal promoter function. We have now isolated three genes encoding U6 RNA in Arabidopsis. Transcription of these genes in transfected protoplasts of Nicotiana plumbaginifolia is resistant to alpha-amanitin indicating that they are transcribed by RNA polymerase III. The upstream regions of three Arabidopsis U6 genes contain USE and -30 TATA-like elements similar to those found to be important for transcription of U2 RNA genes but the spacing between the two elements is about 10 bp closer than in the U2 genes. Using synthetic U6 genes we demonstrate that the USE and TATA elements are indispensable for their transcription, the TATA boxes of U2 and U6 genes are interchangeable, and that the intragenic A box-like sequence of U6 gene is not essential. Increasing the distance between the USE and TATA by 10 bp inactivates U6 gene transcription, demonstrating that proper positioning of the elements is also important for transcription by RNA polymerase III. The data indicate that the structure of U-snRNA gene promoters and the determinants of polymerase specificity are completely different between vertebrates and plants.
Although most eukaryotic genes known to be transcribed by RNA polymerase III have intragenic promoter elements, some are similar to genes transcribed by RNA polymerase II in that they have upstream promoters (reviewed in refs 1-4). Transcription of the vertebrate U6 and 7SK RNA genes by RNA polymerase III depends exclusively upon upstream signals, some of which are indistinguishable from the elements essential for polymerase II-specific genes. In the plant Arabidopsis thaliana the promoter elements for the U6 and U2 small nuclear RNA genes, transcribed by RNA polymerases III and II respectively, are identical, comprising a -30 TATA box and an upstream element specific for small nuclear RNA genes. The distance between these elements differs, however. Here we report evidence that this separation is crucial in determining whether the genes are transcribed by polymerase II or III.
A search for Y-specific DNA sequences has been performed in a sample of seven 46,XX true hermaphrodites and one 45,X mixed gonadal dysgenesis case and compared with a sample of 11 XX males. Using six Y-specific DNA probes no hybridization signal was obtained in the hermaphrodite group; in contrast, all XX males gave a positive signal with at least one probe. This difference is statistically highly significant. We conclude that the aetiology of true hermaphroditism is different from that of the XX male syndrome. As all cases of the hermaphrodite group are positive for the serological sex-specific antigen (Sxs) it is concluded that this antigen can be present even in the absence of Y-specific DNA.
A patient described as a 45,X male (Forabosco et al. 1977) was examined for the presence of Y-specific DNA by using various probes detecting restriction fragments from different regions of the Y chromosome. Positive hybridization signals were obtained for Yp fragments only. In situ hybridization with two different probes, pDP31 and the pseudoautosomal probe 113F, led to a clear assignment of the Yp sequences to the short arm of one chromosome 18. Cytogenetically, the presence of all of Yp including the Y centromere on 18p could be demonstrated replacing a segment of similar size of 18p. Thus, the Y/18 translocation chromosome is dicentric structurally, but it was shown to be monocentric functionally with the no. 18 centromere active. Gene dosage studies with the probe B74 defining a sequence at 18p11.3 demonstrated a single dose of this sequence in the patient. In agreement with these observations, the patient shows clinical signs of the 18p-syndrome. It is concluded that in XO males in general, the X is of maternal origin while the maleness is due to a de novo Y/autosome translocation derived from the father. Depending on the nature of the autosomal deficiency caused by the Y/autosome translocation, the patient may have congenital malformations.
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