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Biomedical subjects

J W Roberts

Publications and source records attributed to J W Roberts.

At least 91 records · Page 5Linked to original sources
PubMed1985

Rho-dependent transcription termination at lambda R1 requires upstream sequences.

In order to define the DNA sequence requirements for rho-dependent transcription termination, we have constructed and analyzed two classes of mutants affecting the cro-tR1 region of phage lambda: one class contains progressively larger deletions in the region upstream of the rho-dependent terminator tR1, and the other consists of small insertions or deletions generated by the linker scanning procedure. We show that rho-dependent termination at lambda tR1 in vitro requires contribution from sequences far upstream of the sites of termination. No single linker scanning mutant completely abolishes transcription termination, indicating that there exists no unique sequence that serves as an obligatory rho utilization site. Our results suggest that rho can interact with a large portion of the RNA transcript, including sequences that would be translated in vivo. Moreover, rho action appears to require a minimal transcript length, about 150-190 nucleotides for the tR1 termination sites. We also examine the effect of the auxiliary transcription factor NusA on transcription of these mutants, and we provide support for the proposal that specific effects of NusA on transcription termination require a sequence known as "boxA" (Olson, E.R., Flamm, E.L., and Friedman, D.I. (1982) Cell 31, 61-70).

Bacterial Proteins↗
PubMed1985

Spontaneous rupture of hepatocellular carcinoma.

Spontaneous rupture of a primary hepatocellular carcinoma is an extremely rare event in the western hemisphere. Only a handful of single-case reports have been authored in the continental United States. A ruptured hepatoma carries a dismal prognosis and is usually beyond a "resection for cure" stage. In this report, two cases of spontaneously ruptured primary hepatocellular carcinoma are described. Both cases involved cirrhotic livers, and the tumor in each case was resected to attempt cure. One patient survived nearly 2 years; the other is alive and well at this time.

Aged↗
PubMed1985

Evolutionary conservation of DNA sequences expressed in sea urchin eggs and early embryos.

DNA sequence divergence measurements indicate that Strongylocentrotus franciscanus is more distinct from S. purpuratus and S. drobachiensis than these two species are from each other, in agreement with paleontological and morphological evidence. The evolutionary divergence of several classes of expressed DNA sequences was compared with that of total single-copy DNA. Between S. franciscanus and S. purpuratus the divergence of cDNA made from gastrula cytoplasmic poly(A)+ RNA is about half that of total single-copy DNA. Similar results were obtained for cDNA made from unfertilized egg poly(A)+ RNA. In contrast, sequences expressed in gastrula nuclear RNA have diverged almost as much as total single-copy DNA.

Animals↗
PubMed1985

Phage lambda gene Q antiterminator recognizes RNA polymerase near the promoter and accelerates it through a pause site.

The positive regulator encoded by phage lambda gene Q is a transcription antiterminator that affects RNA polymerase initiating at the phage late gene promoter, but not at other promoters. We show that this nucleotide-sequence-specific interaction of Q protein and RNA polymerase can occur while the enzyme is pausing after 16 nucleotides of the late gene transcript have been made. Furthermore, Q protein chases RNA polymerase from this early pause site, so that it both recognizes the enzyme and changes its transcription properties at this site. We suggest that the ability of Q-modified RNA polymerase to escape this pause reflects the change that allows it to go through terminators. We also show that NusA protein is required for efficient Q protein activity in vitro.

Bacterial Proteins↗
PubMed1985

Rectal prolapse.

Rectal prolapse occurs mostly in the geriatric female patient and can be a very disabling condition. The etiology is intussusception of the rectosigmoid secondary to excessive and prolonged straining. Medical therapy for this disease process is not helpful and patients will require a surgical procedure. The two best surgical procedures for the correction of rectal prolapse are low anterior resection of the rectosigmoid and proctopexy. A few patients who are unfit for laparotomy may require the Thiersch Wire procedure. Two unresolved problems after surgical therapy are continuing constipation and incontinence. Constipation is treated by dietary measures, stool softeners, and periodic enemas. Laxatives are to be discouraged. Incontinence in patients with rectal prolapse improves in most patients after a procedure to correct the prolapse. In those patients in which incontinence persists, no form of therapy has been found to be uniformly successful.

Adult↗
PubMed1984

A potential stem-loop structure and the sequence CAAUCAA in the transcript are insufficient to signal rho-dependent transcription termination at lambda tR1.

It has been suggested that a sequence in the RNA transcript that can form a stem and loop structure, followed by the sequence CAAUCAA, is the signal for rho-dependent transcription termination. We tested this hypothesis by synthesizing a DNA duplex whose sequence corresponds to a region of the lambda tR1 terminator that contains these structural features. We cloned this synthetic DNA fragment under the control of the lacUV5 promoter, and showed that it does not cause rho-dependent termination in vitro. RNA polymerase pauses during in vitro transcription across the synthetic sequence, although less efficiently than at the corresponding sequence on the lambda template. No rho-mediated termination was detected even under conditions that prolonged transcriptional pausing at the synthetic site, indicating that the synthetic sequence is defective as a transcript release site. We suggest that unlike rho-independent terminators, rho-dependent terminators require sequences in addition to those immediately before the sites of termination.

Base Sequence↗
PubMed1984

Pathological rupture of the spleen associated with hepatitis A.

Spontaneous rupture of the spleen refers to rupture of a normal spleen in the absence of trauma. Atraumatic rupture of a diseased spleen is termed pathological rupture and has been described in a variety of pathological conditions. This report describes a case of pathological splenic rupture associated with hepatitis A confirmed by positive IgM antibodies. To the best of our knowledge, this case represents the sole verified example of this previously unrecognized life-threatening complication of hepatitis.

Adult↗
PubMed1984

RecA protein--promoted lambda repressor cleavage: complementation between RecA441 and RecA430 proteins in vitro.

Induction of prophage lambda occurs in recA441 mutant lysogens after a shift to 42 degrees C in the presence of adenine. If the synthesis of RecA441 protein is maintained at a low basal level by the presence of a second mutation in the recA441 gene, recA453, induction of prophage lambda is prevented. The ability to induce prophage lambda is restored by the introduction, on a transducing phage, of a second recA gene carrying the recA430 mutation; by itself, the RecA430 protein is devoid of activity against the lambda repressor (Rebollo et al. 1984). In order to explain how the RecA430 protein might complement the RecA441 protein to provide lambda repressor cleavage in a recA453-441 (recA430) diploid lysogen, we characterized the cleavage reaction catalysed by a mixture of these proteins in vitro. Our results suggest that, in the presence of dATP, the RecA441 and RecA430 proteins form mixed multimers on single-stranded DNA, in which the RecA441 protein molecules enhance the DNA binding affinity of RecA430 protein molecules, but RecA430 protein molecules support no cleavage of the lambda repressor. Although the effects of the RecA430 and single-strand binding (SSB) proteins are similar in vitro, we show that the SSB protein cannot substitute for the RecA430 protein in restoring lambda repressor cleavage in a recA453-441 lysogen. Comparison of the stimulatory effect of long single-stranded DNA with that of (dA)14 oligonucleotides on the RecA441 protein-directed cleavage of the lambda repressor in the presence of various nucleoside triphosphates (NTPs) indicates that the cooperative binding of the RecA441 protein to single-stranded DNA stabilizes the RecA protein-DNA complexes so that they remain intact long enough to support cleavage of the lambda repressor. We conclude that the low basal level of the RecA441 protein in a recA453-441 cell is sufficient to cleave the lambda repressor, under conditions where a normal basal level of RecA430 protein is also present allowing the formation of mixed multimers on single-stranded DNA regions normally present in the cell.

Bacteriophage lambda↗
PubMed1983

Purification of the bacteriophage lambda late gene regulator encoded by gene Q.

The product of bacteriophage lambda gene Q is a transcription antiterminator that activates phage late gene expression. We report a method to purify the lambda Q protein to near homogeneity. We have followed during purification both activity of Q protein to provoke in vitro synthesis of the lambda late protein endolysin in the DNA-dependent protein-synthesizing system, and radioactivity in a polypeptide that we show by genetic criteria to be lambda Q. These co-purify throughout the procedure.

Bacteriophage lambda↗
PubMed1983

RNA polymerase pausing and transcript release at the lambda tR1 terminator in vitro.

Transcription of the lambda rightward operon encounters multiple tandem p-dependent termination sites at a region known as tR1, yielding the mRNA for the lytic repressor cro. We show that in the absence of p factor, RNA polymerase pauses at each of three p-dependent termination sites (I, II, and III) at tR1 during in vitro transcription. We demonstrate that p factor itself does not affect transcriptional pausing at these sites. Relative occupancies of the pause sites during initial transcription have been measured. The lambda cin-1cnc-1 mutations, which abolish or vastly reduce termination at site II of tR1, nevertheless elicit transcriptional pausing at that site. The NusA protein (L factor) prolongs these transcriptional pauses to varying degrees. Transcriptional pausing at termination site II is dramatically enhanced by NusA in lambda wild type, but not in lambda cin-1cnc-1. NusA enhances pausing at site I but inhibits termination at that site, suggesting that it may also affect transcript release. We show that NusA does alter the kinetics of transcript release from isolated ternary complexes. We suggest that the ability of the NusA protein to increase or decrease termination efficiencies at various sites is attributed to its ability to modulate transcriptional pausing as well as transcript release.

Bacterial Proteins↗
PubMed1983

Mitochondrial DNA sequences in the nuclear genome of Strongylocentrotus purpuratus.

Two sea urchin embryo complementary DNA clones representing mitochondrial 16 S ribosomal RNA and cytochrome oxidase subunit I messenger RNA have been characterized. The cloned cDNAs are colinear with sea urchin mitochondrial DNA, and their identification is based on cross-hybridization with known restriction fragments of human mitochondrial DNA, and on nucleotide sequence determinations. The mitochondrial cDNA clones also displayed an unexpected reaction with specific genomic DNA sequences in gel blot hybridizations. Genomic phage lambda recombinants containing sequences hybridizing with the mitochondrial clones were isolated and the arrangement of these sequences was determined. The genomic region studied contains a sequence homologous with the 3' end of the mitochondrial 16 S rRNA gene, flanked on one side by what is possibly a complete copy of the cytochrome oxidase subunit I gene, and on the other by a duplication of a fragment of this gene. The nucleotide sequence divergence between the mitochondrial and nuclear homologues of the cytochrome oxidase subunit I gene varies for different regions of the gene, from about 13% to 25%, while there is about 8% sequence divergence between nuclear and mitochondrial versions of the 3' 16S rRNA sequence. The structure of the genomic mitochondrial sequence homologues indicates that during sea urchin evolution there occurred a germ-line transposition of a fragment of the mitochondrial genome into the nuclear DNA, followed by rearrangements and single nucleotide substitutions.

Animals↗
PubMed1983

Transcripts of three mitochondrial genes in the RNA of sea urchin eggs and embryos.

cDNA clones representing mitochondrial 16 S rRNA, and mRNAs for cytochrome oxidase I and an unidentified reading frame were used to measure the prevalence and stability of these transcripts in gastrula stage embryos. The 16 S rRNA is the most prevalent embryo poly(A) RNA, and is synthesized about four times more rapidly than is the mRNA for cytochrome oxidase. The relative prevalence of the two mRNAs is largely determined by their turnover rates.

Animals↗
PubMed1983

Urethral hemangioma: treatment by total excision and grafting.

Capillary hemangiomas of the urethra are rare benign tumors that may behave in a persistently recurrent manner. Radical extirpative surgery may be necessary to cure the patient. We describe a new 2-stage technique for replacing long segments of urethra.

Adolescent↗
PubMed1983

Comparison of sea urchin and human mtDNA: evolutionary rearrangement.

Clones of full-length mtDNA have been isolated from a Strongylocentrotus franciscanus recombinant DNA library by screening a cDNA clone of cytochrome oxidase subunit 1 mRNA. Restriction fragment cross-hybridization analysis shows the following difference in gene arrangement between sea urchin and human mtDNA. The 16S rRNA and cytochrome oxidase subunit 1 genes are directly adjacent in sea urchin mtDNA. These two genes are separated in human and other mammalian mtDNAs by the region containing unidentified reading frames 1 and 2. In spite of the difference in gene order, gene polarity appears to have been conserved. We conclude that the difference in gene order reflects a rearrangement that took place in the sea urchin lineage since sea urchins and mammals last shared a common ancestor.

Animals↗
PubMed1982

Comparison of the polyoma virus early and late promoters by transcription in vitro.

Polyoma virus DNA was transcribed in the HeLa whole cell extract in vitro system (1). Early region transcripts with the same 5'-ends as in vivo mRNAs, located 31 +/- 2bp from 'TATA'-boxes, were synthesized by RNA polymerase II. Sequences sufficient for efficient expression of the early promoter were present in a substitution mutant lacking viral DNA from a position 55bp before the principal cap sites. Late region transcripts were synthesised inefficiently. Only one (at nt5129 +/- 2) of the many late mRNA cap sites functioned as an in vitro initiation point. This was the one 5'-end located 31 +/- 2bp from a sequence resembling the 'TATA' consensus. The proportion of late to early region RNA polymerase II transcripts decreased dramatically at suboptimal template concentrations. An hypothesis to explain the regulation of late gene expression in vivo based on these results is proposed. A linear templates were transcribed only by RNA polymerase II, transcripts with the same sense as late mRNAs and 5'-ends at nt5076 +/- 2 were produced from superhelical template by an alpha amanitin resistant enzyme.

Base Sequence↗
PubMed1982

The phage lambda Q gene product: activity of a transcription antiterminator in vitro.

Phage lambda gene Q encodes a positive regulator of phage late gene expression. We show that purified Q gene protein functions as a transcription antiterminator in vitro. It acts at the terminator for lambda 6S RNA, a small constitutive transcript that precedes the late genes, thus opening transcription into the entire late gene region. It is active in a purified transcription system consisting otherwise of only RNA polymerase, L factor and DNA containing the region encoding the 6S RNA. Q protein thus acts directly on the transcription complex to alter its response to transcription terminators.

Bacteriophage lambda↗
PubMed1982

Symptomatic hyperparathyroidism.

In view of our last 100 patients operated on for hyperparathyroidism we found the following: 1. The operation is safe, with no mortality and a 2 per cent morbidity from transient hypoparathyroidism. 2. The operation is effective (99 per cent in our series) with removal of only clinically enlarged glands. 3. Most patients have symptomatic hypercalcemia (73 per cent in our series) and are improved with parathyroidectomy.

Age Factors↗