In vivo footprinting studies suggest a role for chromatin in transcription of the human 7SK gene.

Mutation and deletion analyses of mammalian class III small nuclear RNA genes transcribed by RNA polymerase (pol) III have defined three functional promoter elements: a distal sequence element (DSE) at around -220, a proximal sequence element (PSE) at around -60 and a TATA box at around -30. Although binding studies have identified factors that bind to these sites in vitro, it is not known exactly how proteins interact with the promoters of these genes in vivo. In this study, we have used dimethyl sulphate and DNase I treatment of HeLa cells and nuclei, respectively, followed by linker-mediated polymerase chain reaction, to obtain in vivo footprints of proteins binding to the promoter of the Pol III-transcribed 7SK gene. Our results show that most of the characterised promoter elements of this gene are protected in vivo in these cells, and the pattern of DNase I protection suggests that a nucleosome lies between the DSE and the PSE. Methylation protection was also seen upstream of the DSE over a sequence corresponding to the binding site of a POZ domain-containing protein, ZID, which interacts with components of histone deacetylase complexes. These findings suggest that chromatin structure plays a role in the cascade of protein-DNA interactions that regulate expression of this pol III-transcribed gene.

Transcription of the human U2 snRNA genes continues beyond the 3' box in vivo.

The 3' box of the human class II snRNA genes is required for proper 3' processing of transcripts, but how it functions is unclear. Several lines of evidence suggest that termination of transcription occurs at the 3' box and the terminated transcript is then a substrate for processing. However, using nuclear run-on analysis of endogenous genes, we demonstrate that transcription continues for at least 250 nucleotides beyond the 3' box of the U2 genes. Although in vivo footprinting analysis of both the U1 and U2 genes detects no protein-DNA contacts directly over the 3' box, a series of G residues immediately downstream from the 3' box of the U1 gene are clearly protected from methylation by dimethylsulfate. In conjunction with the 3' box of the U1 gene, this in vivo footprinted region causes termination of transcription of transiently transfected U2 constructs, whereas a 3' box alone does not. Taken together, these results indicate that the 3' box is not an efficient transcriptional terminator but may act as a processing element that is functional in the nascent RNA.

[Genetic diagnosis of Williams syndrome]. / A Williams-szindróma genetikai diagnózisa.

Williams syndrome is a complex developmental disorder. The major cardiovascular component of Williams syndrome is supravalvular aortic stenosis, a progressive disease that may need surgical repair. Williams syndrome is associated with heterozygous microdeletion in the chromosomal region 7q11.23 encompassing the elastin gene. We have identified a new, highly informative tetranucleotide repeat polymorphism within the human elastin gene. This marker together with other, previously described elastin gene markers was used to show deletion of the elastin gene in nine sporadic Williams syndrome patients from Hungary. Application of polymorphisms within and flanking the elastin gene on chromosome 7 provides a fast, polymerase chain reaction based method for mutational analysis of Williams syndrome patients.

Functional redundancy of promoter elements ensures efficient transcription of the human 7SK gene in vivo.

Deletion and mutation studies of the human 7SK gene transfected into HeLa cells have identified three functional regions of the promoter corresponding to the TATA box at -25, the proximal sequence element (PSE) between -49 and -65 and the distal sequence element (DSE) between -243 and -210. These elements show sequence homology to equivalent regions in other snRNA genes and are functionally analogous. Unlike the DSEs of many snRNA genes however, the 7SK DSE does not contain a consensus binding site for the transcription factor Oct-1 but rather, contains two non-consensus Oct-1 binding sites that can function independently of one another to enhance transcription. Unusually, the 7SK PSE can retain function even after extensive mutation and removal of the conserved TGACC of the PSE has little effect in the context of the whole promoter. However, the same mutation abolishes transcription in the absence of the DSE suggesting that protein/protein interactions between DSE and PSE binding factors can compensate for a mutant PSE. Mutation of the 7SK TATA box allows snRNA type transcription by RNA polymerase II to occur and this is enhanced by the DSE, indicating that both the DSE and the PSE can also function with pol II. In addition, mutation of the TATA box does not abolish pol III dependent transcription, suggesting that other sequence elements may also play a role in the determination of polymerase specificity. Although the human 7SK gene is transcribed efficiently in Xenopus oocytes, analysis of the 7SK wild-type gene and mutants in Xenopus oocytes gives significantly different results from the analysis in HeLa cells indicating that the recognition of functional elements is not the same in the two systems.

Additional upper Pleistocene human remains from Vindija cave, Croatia, Yugoslavia.

This report presents a morphological description of ten hominid skeletal fragments found at Vindija cave, northwestern Yugoslavia, in 1980-1981. Eight of the specimens (seven cranial fragments and one complete hand proximal phalanx) were excavated from level G3, a stratum correlated to the Lower Würm stadial and containing Mousterian lithic elements. The salient morphological features of these new specimens are similar to those of previously described hominids from this same stratum, indicating that the new specimens are also remains of archaic H. sapiens (Neandertals). One parietal fragment was excavated from the stratigraphically more recent F complex. The F complex contains Upper Paleolithic lithic elements, and the new hominid specimen from this complex appears to exhibit features similar to other early modern H. sapiens in southcentral Europe.