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R Tjian

Publications and source records attributed to R Tjian.

212 records · Page 12Linked to original sources
PubMed1975

Antibody directed against Bacillus subtilis rho factor purified by sodium dodecyl sulfate slab gel electrophoresis. Effect on transcription by RNA polymerase in crude extracts of vegetative and sporulating cells.

Antibody directed against rho factor from vegetative Bacillus subtilis was prepared by immunizing a rabbit with denaturated rho polypeptide isolated by electrophoresis of partially purified DNA-dependent RNA polymerase on a sodium dodecyl sulfate-polyacrylamide slab gel. Antiserum to rho reacted specifically with native rho polypeptide but not with core RNA polymerase as judged by complement fixation and by an immunodiffusion assay. Anti-rho antibody also inhibited the ability of rho to stimulate transcription of phage phie DNA but failed to inhibit transcription of poly(dA-dT) by core enzyme. Specific antibody was also raised against a mixture of the beta and beta' subunits of RNA polymerase purified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The effect of the anti-rho gamma-globulin on the transcription of phage phie DNA by RNA polymerase in crude extracts of vegetative and sporulating cells was examined. Anti-rho antibody markedly inhibited the transcription of phage DNA by RNA polymerase partially purified from vegetative bacteria by ammonium sulfate fractionation but had little effect on transcription of the phage DNA template by enzyme from sporulating cells. Addition of purified rho to a vegetative extract that had been depleted of rho by treatment with the anti-rho antibody restored active transcription of phage DNA. However, addition of purified rho to an antibody-treated extract of sporulating cells had little effect on phie RNA synthesis. These findings suggest that sporulating cells contain a component that interferes with the activity of the rho subunit of RNA polymerase.

Antibodies, Bacterial↗
PubMed1974

Chloramphenicol restores sigma factor activity to sporulating Bacillus subtilis.

The sigma subunit of RNA polymerase from sporulating Bacillus subtilis is markedly inhibited in its ability to direct active transcription of phage varphie DNA in vitro. Treatment of sporulating bacteria with chloramphenicol rapidly restores sigma activity, suggesting that sporulating cells contain an inhibitor of sigma that is physiologically unstable or that becomes unstable after drug treatment. The hypothetical inhibitor is depleted exponentially with an apparent half-life of 11 min at 37 degrees .

Bacillus subtilis↗
PubMed1974

An immunological assay for the sigma subunit of RNA polymerase in extracts of vegetative and sporulating Bacillus subtilis.

The activity of the sigma subunit of Bacillus subtilis RNA polymerase decreases markedly during the first hours of sporulation [T.G. Linn et al. (1973) Proc. Nat. Acad. Sci. USA 70, 1865-1869]. We have prepared antibody against RNA polymerase holoenzyme to determine the fate of sigma polypeptide during spore formation. This antiserum specifically and independently precipitates sigma and core polymerase from crude extracts of B. subtilis as judged by both sodium dodecyl sulfate and urea gel electrophoresis of the precipitates. We report that crude extracts of sporulating cells lacking sigma activity contain as much sigma polypeptide as extracts of vegetative cells. However, sigma polypeptide in extracts from sporulating cells is apparently only weakly associated with RNA polymerase, as indicated by the failure of sigma to co-purify efficiently with core enzyme during phase partitioning. The loss of sigma activity and the weak binding of sigma to core enzyme occurs normally in a mutant blocked at an intermediate stage of sporulation (SpoII-4Z) and in wild-type bacteria sporulating in 121B medium, Difco sporulation medium, or Sterlini-Mandelstam resuspension medium. In contrast, sigma in two mutants (SpoOa-5NA and SpoOb-6Z) blocked at an early stage of spore formation remains active and tightly associated with RNA polymerase during stationary phase.

Animals↗
PubMedDate not supplied

Multiple specific contacts between a mammalian transcription factor and its cognate promoters.

The human transcription factor Sp1 binds upstream of certain viral and cellular promoters and activates initiation of RNA synthesis from these promoters by RNA polymerase II. The Sp1 binding sites of both simian virus 40 and a related monkey promoter contain multiple copies of the sequence GGGCGG, three or four of these sequences forming close contacts with Spl. The clustered contacts fall on one strand of the DNA and are arranged similarly in the major groove of the DNA helix in both promoters.

Animals↗
PubMedDate not supplied

Comparative chromosome mapping of a conserved homoeo box region in mouse and human.

Specific genes are assumed to regulate pattern formation in the mammalian embryo, but as yet none has been identified unequivocally. It is possible that such genes in mammals may be identified by virtue of a conserved coding sequence, because many of the Drosophila melanogaster homoeotic and segmentation genes, which have crucial roles in the regulation of segmental pattern formation during embryonic development, contain a 180-base pair (bp) DNA sequence, the homoeo box, and that sequences homologous to the Drosophila homoeo box are also present in 6-10 copies in higher animals, including mammals. Although the assumption that the homoeo box identifies genes responsible for pattern formation in mammals remains to be validated, it is a particularly attractive hypothesis given the strong conservation of homoeo boxes over vast evolutionary distances. Here we report the localization of a human homoeo box region, previously cloned and shown to contain two homoeo boxes within a sequence of 5-kilobases (kb), to the long arm of chromosome 17. We show that two single-copy homoeo box-flanking probes derived from this region strongly hybridize to single-copy restriction fragments in mouse genomic DNA and that these conserved homoeo box-flanking sequences map to mouse chromosome 11. This may be significant as several genes that map to chromosome 17 in human also map to chromosome 11 in the mouse, implying that a segment of mouse chromosome 11 is homologous to a region of human chromosome 17. Taken together, these data suggest that the homoeo box region detected with our probes is highly conserved in human and mouse.

Animals↗
PubMedDate not supplied

Control of eukaryotic messenger RNA synthesis by sequence-specific DNA-binding proteins.

The enzymatic machinery that carries out RNA synthesis provides the cell with the means to adjust the patterns of transcription in response to environmental and developmental signals. In eukaryotes, this regulation is mediated in part by promoter-specific transcription factors, which are DNA-binding proteins with the ability to discriminate between distinctive DNA sequence elements found in the promoter regions of different genes. The presence of these factors bound to DNA enables other components of the transcriptional machinery, including the RNA polymerase, to initiate transcription with selectivity and accuracy.

Base Sequence↗
PubMedDate not supplied

Sp1 binds to promoter sequences and activates herpes simplex virus 'immediate-early' gene transcription in vitro.

During a herpes simplex virus (HSV-1) lytic infection, three classes of viral genes are transcribed in a temporally regulated manner. The HSV-1 'immediate-early' (IE) promoter sequences contain multiple copies of a hexanucleotide sequence, GGGCGG, known as a GC box, and one or more copies of an 11-base pair (bp) conserved A + T-rich element, designated TAATGARAT. The TAATGARAT elements are thought to mediate the trans-activation of IE RNA synthesis by a virion-associated protein(s), and the flanking G + C-rich sequences appear both to potentiate this induction and to direct IE promoter activity in vivo. The similarity of the herpesvirus GC box repeats to those of the simian virus 40 (SV40) early promoter prompted the in vitro analysis of HSV IE transcription reported here. We show that the mammalian gene-specific transcription factor Sp1 binds to eight distinct regions of the HSV short terminal repeat and stimulates transcription 25-fold from the divergent IE-3 (ICP-4) and IE-4/5 promoters.

Binding Sites↗
PubMedDate not supplied

Transcription factor Sp1 recognizes a DNA sequence in the mouse dihydrofolate reductase promoter.

Human (HeLa) cells contain a host-cell-encoded transcription factor, Sp1, which is required for transcription of simian virus 40 (SV40) promoters. Since the discovery of Sp1 we have been interested in learning what role this factor plays in uninfected cells. A monkey cellular gene promoter interacts with Sp1, but no gene products linked to this promoter have yet been identified. The finding that the sequence of the 5'-flanking DNA of the mouse dihydrofolate reductase (DHFR) gene contains several regions showing strong homology to the Sp1 binding region of simian virus 40 (SV40) prompted us to undertake experiments with dhfr. We report here that Sp1 binds to these regions in the dhfr promoter, and that Sp1-containing preparations stimulate transcription from the dhfr promoter in an in vitro reaction. Our results suggest that in addition to its interactions with the SV40 viral promoter, one function of Sp1 is to direct the expression of the cellular DHFR gene.

Animals↗
PubMedDate not supplied

Activation of transcription by two factors that bind promoter and enhancer sequences of the human metallothionein gene and SV40.

Genetic analysis of eukaryotic transcriptional promoters has revealed that protein-coding genes often contain a complex array of cis-control elements consisting of upstream activator sequences and enhancer elements. The metallothionein genes provide a useful example for dissecting the action of multiple interspersed control elements that govern both basal level and regulated expression in animal cells. The human metallothionein (hMTIIA) promoter has been analysed in detail and found to contain no less than five distinct control elements in the 5' flanking regions of the gene that mediate specificity and regulation of transcription. These different control elements can be functionally subdivided into two categories: basal and induced elements. There are several distinct basal recognition sequences, which include a TATA-box, a GC-box, and at least two basal level enhancer (BLE) sequences, that function like classical enhancer elements. The hMTIIA gene also responds to induction by heavy metals and by steroid hormones through the action of metal regulatory elements (MRE) and glucocorticoid responsive elements (GRE). Here we report the identification of two cellular DNA-binding proteins that interact selectively with sequences governing the basal level expression of hMTIIA. One of these factors is a novel activator protein (AP1) that interacts with sequences in the BLE of hMTIIA and also binds to a site within the 72-base pair (bp) repeats of the simian virus 40 (SV40) enhancer region. The second protein has been purified to homogeneity and shown to be transcription factor Sp1 which recognizes and binds to a single GC-box element within the hMTIIA promoter.

Base Sequence↗
PubMedDate not supplied

Selectivity of chromatin-remodelling cofactors for ligand-activated transcription.

An array of regulatory protein and multi-subunit cofactors has been identified that directs eukaryotic gene transcription. However, establishing the specific functions of various related cofactors has been difficult owing to the limitations inherent in assaying transcription in animals and cells indirectly. Here we describe, using an integrated chromatin-dependent reconstituted transcription reaction, the purification and identification of a multi-subunit cofactor (PBAF) that is necessary for ligand-dependent transactivation by nuclear hormone receptors. A highly related cofactor, human SWI/SNF, and the ISWI-containing chromatin-remodelling complex ACF both fail to potentiate transcription. We also show that transcriptional activation mediated by nuclear hormone receptors requires TATA-binding protein (TBP)-associated factors (TAFs) as well as the multi-subunit cofactors ARC/CRSP. These studies demonstrate functional selectivity amongst highly related complexes involved in gene regulation and help define a more complete set of factors and cofactors required to activate transcription.

Adenosine Triphosphate↗