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Biomedical subjects

K Bloom

Publications and source records attributed to K Bloom.

At least 163 records · Page 9Linked to original sources
PubMed1988

Chromatin structure of altered yeast centromeres.

We have investigated the chromatin structure of wild-type and mutationally altered centromere sequences in the yeast Saccharomyces cerevisiae by using an indirect end-labeling mapping strategy. Wild-type centromere DNA from chromosome III (CEN3) exhibits a nuclease-resistant chromatin structure 220-250 base pairs long, centered around the conserved centromere DNA element (CDE) III. A point mutation in CDE III that changes a central cytidine to a thymidine and completely disrupts centromere function has lost the chromatin conformation typically associated with the wild-type centromere. A second conserved DNA element, CDE I, is spatially separated from CDE III by 78-86 A + T-rich base pairs, which is termed CDE II. The sequence and spatial requirements for CDE II are less stringent; alterations in CDE II length and sequence can be tolerated to a limited extent. Nuclease-resistant cores are altered in dimension in two CDE II CEN3 mutations. Two CDE I deletion mutations that retain partial centromere function also show nuclease-resistant regions of reduced size and intensity. The results from a number of such altered centromeres indicate a correlation between the presence of a protected core and centromere function.

Base Sequence↗
PubMed1988

Selective excision of the centromere chromatin complex from Saccharomyces cerevisiae.

We have taken advantage of the known structural parameters associated with centromere DNA in vivo to construct a CEN fragment that can be selectively excised from the chromatin DNA with restriction endonucleases. CEN3 DNA is organized in chromatin such that a 220-250-bp region encompassing the elements of centromere homology is resistant to nuclease digestion. Restriction enzyme linkers encoding the Bam HI-recognition site were ligated to a 289 base pair DNA segment that spans the 220-250-bp protected core (Bloom et al., 1984). Replacement of this CEN3-Bam HI linker cassette into a chromosome or plasmid results in formation of a complete structural and functional centromeric unit. A centromere core complex that retains its protected chromatin conformation can be selectively excised from intact nuclei by restriction with the enzyme Bam HI. The centromeric protein-DNA complex is therefore not dependent upon the intact torsional constrains on linear chromosomes for its structural integrity. Isolation of this complex provides a novel approach to characterizing authentic centromeric proteins bound to DNA in their native state.

Cell Nucleus↗
PubMed1987

UV-induced damage and repair in centromere DNA of yeast.

The centromere is the region within a chromosome that is required for proper segregation during mitosis and meiosis. Lesions in this sequence represent a unique type of damage, as loss of function could result in catastrophic loss of the genetic material of an entire chromosome. We have measured the induction by ultraviolet (UV) light of pyrimidine dimers in a 2550-bp restriction fragment that includes the centromere region of chromosome III in Saccharomyces cerevisiae. Yeast cells were exposed to ultraviolet light, cellular DNA was gently extracted, and subsequently treated with a UV-specific endonuclease to cleave all pyrimidine dimers. The sites of UV-specific nuclease scission within the centromere were determined by separating the DNA according to molecular weight, transferring the fragments to nitrocellulose, and hybridizing to a radiolabeled 624-bp fragment homologous to the centromere DNA from chromosome III. Several hotspots were identified in chromatin DNA from cells, as well as in irradiated deproteinized DNA. Double strand damage due to closely opposed pyrimidine dimers was also observed. At biological doses (35% survival) there are approximately 0.1 to 0.2 pyrimidine dimers per centromere. These dimers are efficiently repaired in the centromere and surrounding region.

Base Sequence↗
PubMed1987

Genetic manipulation of centromere function.

A conditional centromere was constructed in Saccharomyces cerevisiae by placing the centromere of chromosome III immediately downstream from the inducible GAL1 promoter from S. cerevisiae. By utilizing growth conditions that favor either transcriptional induction (galactose-carbon source) or repression (glucose-carbon source) from the GAL1 promoter, centromere function can be switched off or on, respectively. With the conditional centromere we were able to radically alter the mitotic transmission pattern of both monocentric and dicentric plasmids. Moreover, it was possible to selectively induce the loss of a single chromosome from a mitotically dividing population of cells. We observed that the induction of chromosome III aneuploidy resulted in a dramatic change in cell morphology. The construction of a conditional centromere represents a novel way to create conditional mutations of cis-acting DNA elements and will be useful for further analysis of this important stabilizing element.

Centromere↗
PubMed1987

Manifestations of sepsis.

The clinical manifestations of sepsis may be flagrant or subtle. Awareness of the signs and symptoms of sepsis allows early recognition and prompt, appropriate management. The clinical presentation, relative frequency, and current pathophysiologic understanding of the manifestations of sepsis are reviewed. Special emphasis is placed on the cardiopulmonary manifestations, which are examined in a temporal sequence of preshock, early shock, and late shock states. While therapy for the underlying infection (such as antibiotics and drainage of abscesses) is often sufficient, therapy for the specific manifestations of sepsis may also be necessary. Guidelines for therapy for these manifestations of sepsis are given.

Bacterial Infections↗
PubMed1986

Work of breathing through different sized endotracheal tubes.

The ability to breathe spontaneously through an endotracheal tube is a usual prerequisite before an intubated patient can have it removed. Other researchers have measured air flow resistance through endotracheal tubes. In this study, we evaluated work of breathing in joules per min and tension-time index while three normal volunteers breathed through different sized endotracheal tubes. Four 27.5-cm endotracheal tubes were used. Subjects breathed with a constant tidal volume of 500 ml. By increasing respiratory frequency, minute ventilation was increased from 5 to 30 L/min. As tube diameter decreased, work and the tension-time index increased. Changes were magnified at higher minute ventilations through the 6- and 7-mm endotracheal tubes, and the tension-time index critical fatigue level of 0.15 was approached or exceeded.

Equipment Design↗
PubMed1986

Tightly centromere-linked gene (SPO15) essential for meiosis in the yeast Saccharomyces cerevisiae.

We used DNA fragments from the centromere regions of yeast (Saccharomyces cerevisiae) chromosomes III and XI to examine the transcriptional activity within this chromosomal domain. DNA transcripts were found 200 to 300 base pairs from the 250-base-pair centromere core and lie within an ordered chromatin array. No transcripts were detected from the functional centromere region. We examined the cellular function of one of these tightly centromere-linked transcripts. (CEN11)L, by disrupting the coding sequences in vivo and analyzing the phenotype of the mutant yeast cell. Diploids heterozygous for the (CEN11)L disruption sporulated at wild-type levels, and the absence of the (CEN11)L gene product had no effect on the viability or mitotic growth of haploid cells. Diploids homozygous for the (CEN11)L disruption were unable to sporulate when induced by the appropriate nutritional cues. The mutant cells were competent for intragenic recombination and appeared to be blocked at the mononucleate stage. The temporal ordering of (CEN11)L function with respect to the sporulation mutant spo13 suggests that the (CEN11)L gene product may be required at both the first and second meiotic cell divisions. This new sporulation gene has been termed SPO15.

Centromere↗
PubMed1986

Palliation of bronchogenic carcinoma with 198Au implantation using the fiberoptic bronchoscope.

The majority of cases of bronchogenic carcinoma remain incurable, and many of these patients require palliation of the effects of the tumor on the airway. We have developed a technique for implanting radioactive (198Au) seeds via the fiberoptic bronchoscope. We now retrospectively review the results obtained in 111 procedures in 54 patients. Response was assessed by improvement in symptoms, chest roentgenogram, or bronchoscopic appearance. Nineteen of 29 (66 percent) patients with occluding endobronchial lesions benefitted. Twenty of 22 (91 percent) with hemoptysis improved. All six patients with tracheal lesions benefitted. Two of six (33 percent) patients with nonoccluding endobronchial lesions responded. Complications directly related to the procedure were rarely of major consequence, although a single patient had an exsanguinating hemoptysis four days following the last of multiple implantations. The simplicity, relative safety, and potential wide availability coupled with low equipment costs would suggest an increasing role for this technique in the palliation of endobronchial neoplasms.

Adult↗
PubMed1985

Characterization of a tightly centromere-linked gene essential for meiosis in the yeast Saccharomyces cerevisiae.

The centromere region in the yeast Saccharomyces cerevisiae is characterized by short DNA fragments, less than 1,000 bp in length, that are capable of stabilizing entire chromosomes throughout mitotic and meiotic cell divisions. The CEN fragments are organized in a unique chromatin structure and are surrounded by ordered arrays of nucleosomal subunits. RNA transcripts are found 200-300 bp from the centromere, and lie within this ordered chromatin array. No transcripts have been detected through the centromere itself. We have examined the expression and cellular function of a tightly centromere-linked transcript on chromosome 11, (CEN11)L. The (CEN11)L transcript is present at constitutive levels throughout the mitotic and meiotic cell cycles. Disruption of the coding sequences in vivo has no effect on cell viability or mitotic growth, but the cells are unable to sporulate. Genetic complementation with known mutants in sporulation (spo10, spo13) has defined (CEN11)L as a new locus that appears to be required during both meiotic segregation divisions.

Base Sequence↗
PubMed1984

Staphylococcal abscess complicating endocardial aneurysmectomy.

Subendocardial left ventricular aneurysmectomy relieved previously intractable ventricular tachycardia in a 68-year-old man with severe coronary artery disease. Staphylococcal septicemia developed postoperatively; an infected venoclysis site may have provided the portal of entry. Autopsy confirmed staphylococcal abscess at the epicardial aspect of the ventriculotomy, constituting the first reported case, to our knowledge, of this complication following endocardial surgery.

Abscess↗
PubMed1984

Stable transfer and expression of exogenous human globin genes in human erythroleukemia (K562) cells.

To study the expression of globin genes in human cells, human epsilon-globin genes were transferred into a K562 cell line, Bos, which synthesizes very low amounts of epsilon-globin mRNA. A plasmid (pSV2neo-epsilon) containing a complete epsilon-globin gene and 2 kilobases (kb) of 5' flanking DNA as well as a neomycin-resistance gene and a simian virus 40 origin of replication was transfected into Bos cells; the compound G418, a neomycin analogue, was used to select transformed cells. The presence of unique bands by DNA restriction analysis shows that 11 of 14 of the G418-resistant clones have at least one copy of an integrated epsilon-globin gene. RNA expression measured by RNA blotting shows significantly more epsilon-globin mRNA sequences than in untransfected Bos cells in 10 of 11 lines; in most lines, epsilon-globin mRNA was additionally increased in the presence of hemin. In two lines, epsilon-globin mRNA expression with hemin was comparable to that of a high epsilon-globin producing cell line, K562 clone 2. The one G418-resistant line without epsilon-globin genes had no epsilon-mRNA expression. The high epsilon-mRNA expression in several of the lines suggests that exogenous epsilon-globin genes with only 2-kb 5' flanking DNA may be sufficient to be appropriately expressed in these homologous erythroid cells. These results have implications for the potential success of transfer of normal human genes to human bone marrow cells as an approach to the treatment of inherited anemias.

Base Sequence↗
PubMed1983

Inhibition of airway reactivity by nifedipine in patients with coronary artery disease.

Nine patients with coronary artery disease were challenged with aerosolized methacholine before and 3 days after the daily oral administration of 30 mg of nifedipine. Resting airway tone appeared to be generally unaffected by nifedipine, as there were no significant changes in the baseline ventilatory functions. Serial measurements of spirometry and airway resistance were made after cumulative methacholine doses of 1.25, 3.75, and 8.75 mg. Identical doses of methacholine induced smaller changes in specific airway conductance (p less than 0.02), airway resistance (p less than 0.001), FEV1 (p less than 0.02), and FEF25-75% (p less than 0.01) after nifedipine therapy than before. These differences were significant at all methacholine dosage levels by paired t test except for FEV1 at the 3.75 mg dose. This study demonstrated that the bronchospastic response to methacholine in patients with coronary artery disease is reduced after the administration of nifedipine. This salutary effect on ventilatory function may be of importance to cardiac patients with concomitant chronic airway obstruction who are intolerant of beta-adrenergic blocking agents.

Airway Resistance↗