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Biomedical subjects

D A Wolf

Publications and source records attributed to D A Wolf.

At least 19 recordsLinked to original sources
PubMed2001

The fission yeast COP9/signalosome is involved in cullin modification by ubiquitin-related Ned8p.

BACKGROUND: The function of the fission yeast cullins Pcu1p and Pcu4p requires modification by the ubiquitin-related peptide Ned8p. A recent report by Lyapina et al. shows that the COP9/signalosome (CSN), a multifunctional eight subunit complex, regulates Ned8p modification of Pcu1p. Disruption of caa1/csn1, which encodes subunit 1 of the putative S. pombe CSN, results in accumulation of Pcu1p exclusively in the modified form. However, it remained unclear whether this reflects global control of all cullins by the entire CSN complex. RESULTS: We demonstrate that multiple CSN subunits control Ned8p modification of Pcu3p, another fission yeast cullin, which, in complex with the RING domain protein Pip1p, forms a ubiquitin ligase that functions in cellular stress response. Pcu3p is modified by Ned8p on Lys 729 and accumulates exclusively in the neddylated form in cells lacking the CSN subunits 1, 3, 4, and 5. These CSN subunits co-elute with Pcu3p in gel filtration fractions corresponding to approximately 550 kDa and specifically bind both native and Ned8p-modified Pcu3p in vivo. While CSN does not influence the subcellular localization of Pcu3p, Pcu3p-associated in vitro ubiquitin ligase activity is stimulated in the absence of CSN. CONCLUSIONS: Taken together, our data suggest that CSN is a global regulator of Ned8p modification of multiple cullins and potentially other proteins involved in cellular regulation.

COP9 Signalosome Complex↗
PubMed2001

Promotion of NEDD-CUL1 conjugate cleavage by COP9 signalosome.

SCF ubiquitin ligases control various processes by marking regulatory proteins for ubiquitin-dependent proteolysis. To illuminate how SCF complexes are regulated, we sought proteins that interact with the human SCF component CUL1. The COP9 signalosome (CSN), a suppressor of plant photomorphogenesis, associated with multiple cullins and promoted cleavage of the ubiquitin-like protein NEDD8 from Schizosaccharomyces pombe CUL1 in vivo and in vitro. Multiple NEDD8-modified proteins uniquely accumulated in CSN-deficient S. pombe cells. We propose that the broad spectrum of activities previously attributed to CSN subunits--including repression of photomorphogenesis, activation of JUN, and activation of p27 nuclear export--underscores the importance of dynamic cycles of NEDD8 attachment and removal in biological regulation.

3T3 Cells↗
PubMed2001

Heimlich trauma: a violent maneuver.

The Heimlich maneuver is a life-saving technique for dislodging foreign material from the respiratory tract. This report illustrates intraabdominal injuries, including a large mesenteric laceration, mesenteric contusions, and intraperitoneal hemorrhage, that occurred in a recipient of a vigorously applied Heimlich maneuver. The potential for confusing such injuries with homicidally inflicted injuries is emphasized.

Airway Obstruction↗
PubMed2001

Hyperostosis cranii ex vacuo in adults: a consequence of brain atrophy from diverse causes.

Hyperostosis cranii ex vacuo is diffuse thickening of the bones of the cranium occurring after successful ventricular shunting in hydrocephalic children, presumably as a compensatory phenomenon. We present three adults with severe brain atrophy and correspondingly severe skull thickening. In each, the cause of cerebral atrophy was well defined, and none had undergone ventricular shunting. In two, brain atrophy resulted from different temporally discrete insults sustained in adult life, ischemic in one and traumatic in the other. In the third case. progressive brain atrophy resulted from a primary neurodegenerative disorder, Hallervordan Spatz disease. Our observations suggest that hyperostosis cranii ex vacuo is a more general phenomenon than has been previously recognized, and point to a relationship between dynamic changes in brain size and skull thickness. We suggest that such relationships should be taken into account in anthropometric evaluation of the skull.

Adult↗
PubMed2000

Mechanical culture conditions effect gene expression: gravity-induced changes on the space shuttle.

Three-dimensional suspension culture is a gravity-limited phenomenon. The balancing forces necessary to keep the aggregates in suspension increase directly with aggregate size. This leads to a self-propagating cycle of cell damage by balancing forces. Cell culture in microgravity avoids this trade-off. We determined which genes mediate three-dimensional culture of cell and tissue aggregates in the low-shear stress, low-turbulent environment of actual microgravity. Primary cultures of human renal cortical cells were flown on the space shuttle. Cells grown in microgravity and ground-based controls were grown for 6 days and fixed. RNA was extracted, and automated gene array analysis of the expression of 10, 000 genes was performed. A select group of genes were regulated in microgravity. These 1,632 genes were independent of known shear stress response element-dependent genes and heat shock proteins. Specific transcription factors underwent large changes in microgravity including the Wilms' tumor zinc finger protein, and the vitamin D receptor. A specific group of genes, under the control of defined transcription factors, mediate three-dimensional suspension culture under microgravity conditions.

Animals↗
PubMed1999

F-box/WD-repeat proteins pop1p and Sud1p/Pop2p form complexes that bind and direct the proteolysis of cdc18p.

Ubiquitin-dependent proteolysis plays an important role in cell-cycle control [1] [2]. In budding yeast, the protein Skp1p, the cullin-family member Cdc53p, and the F-box/WD-repeat protein Cdc4p form the SCFCdc4p ubiquitin ligase complex, which targets the cyclin-dependent kinase (Cdk) inhibitor Sic1p for proteolysis [3] [4] [5] [6] [7] [8]. Sic1p is recruited to the SCFCdc4p complex by binding to the WD-repeat region of Cdc4p [5] [6], while Skp1p binds to the F-box of Cdc4p [9]. In fission yeast, two distinct Cdc4p-related proteins, Pop1p/Ste16p [10] [11] and the recently identified Sud1p/Pop2p [12], regulate the stability of the replication initiator Cdc18p and the Cdk inhibitor Rum1p. We show here that, despite their structural and functional similarities, the pop1 and pop2 genes fail to complement each other's deletion phenotypes, indicating that they perform non-redundant, but potentially interdependent, functions in proteolysis. Consistent with this hypothesis, Pop1p and Pop2p formed heterooligomeric complexes when overexpressed, and binding of Cdc18p to Pop2p was dependent on Pop1p. The Pop1p-Pop2p interaction was mediated by the amino-terminal domain of Pop2p which, when fused to full-length Pop1p, rescued the phenotype of a Deltapop1Deltapop2 double mutant. Thus, close physical proximity of two distinct F-box/WD-repeat proteins directs proteolysis mediated by the SCFPop ubiquitin ligase complex.

Binding Sites↗
PubMed1999

Tumor necrosis factor-alpha contributes to ischemia- and reperfusion-induced endothelial activation in isolated hearts.

-During myocardial reperfusion, polymorphonuclear neutrophil (PMN) adhesion involving the intercellular adhesion molecule-1 (ICAM-1) may lead to aggravation and prolongation of reperfusion injury. We studied the role of early tumor necrosis factor-alpha (TNF-alpha) cleavage and nuclear factor-kappaB (NF-kappaB) activation on ICAM-1 expression and venular adhesion of PMN in isolated hearts after ischemia (15 minutes) and reperfusion (30 to 480 minutes). NF-kappaB activation (electromobility shift assay) was found after 30 minutes of reperfusion and up to 240 minutes. ICAM-1 mRNA, assessed by Northern blot, increased during the same interval. Functional effect of newly synthesized adhesion molecules was found by quantification (in situ fluorescence microscopy) of PMN, given as bolus after ischemia, which became adherent to small coronary venules (10 to 50 microm in diameter). After 480 minutes of reperfusion, ICAM-1-dependent PMN adhesion increased 2.5-fold compared with PMN adhesion obtained during acute reperfusion. To study the influence of NF-kappaB on PMN adhesion, we inhibited NF-kappaB activation by transfection of NF-kappaB decoy oligonucleotides into isolated hearts using HJV-liposomes. Decoy NF-kappaB but not control oligonucleotides blocked ICAM-1 upregulation and inhibited the subacute increase in PMN adhesion. Similar effects were obtained using BB 1101 (10 microg), an inhibitor of TNF-alpha cleavage enzyme. These data suggest that ischemia and reperfusion in isolated hearts cause liberation of TNF-alpha, activation of NF-kappaB, and upregulation of ICAM-1, an adhesion molecule involved in inflammatory response after ischemia and reperfusion.

Animals↗
PubMed1999

Differentiation of microorganisms based on pyrolysis-ion trap mass spectrometry using chemical ionization.

The ability to differentiate microorganisms using pyrolysision trap mass spectrometry was demonstrated for five Gram-negative disease-causing organisms: Brucella melitensis, Brucella suis, Vibrio cholera, Yersinia pestis, and Francisella tularensis. Bacterial profiles were generated for gamma-irradiated bacterial samples using pyrolytic methylation and compared for electron ionization and chemical ionization using several liquid reagents with increasing proton affinities. Electron ionization combined with pyrolysis caused extensive fragmentation, resulting in a high abundance of lower mass ions and diminishing the diagnostic value of the technique for compound identification and bacterial profiling. Chemical ionization reduced the amount of fragmentation due to ionization while enhancing the molecular ion region of the fatty acids. As the proton affinity of the reagent increased, the protonated molecular ions of the fatty acids became the predominant ions observed in the mass spectrum. As a result, chemical ionization was shown to be more effective than electron ionization in bacterial profiling. Whereas the bacteria could be distinguished at the Genera level using electron ionization, further differentiation to the subspecies level was possible using chemical ionization. The greatest separation among the five test organisms, in terms of Euclidean distances, was obtained using ethanol as the chemical ionization reagent and using pooled masses representing specific fatty acid biomarkers rather than total ion profiles.

Fatty Acids↗
PubMed1999

Regulation of c-myc and immunoglobulin kappa gene transcription by promoter-proximal pausing of RNA polymerase II.

In normal cells, the proto-oncogene c-myc is regulated by promoter-proximal pausing of RNA polymerase II (pol II). In Burkitt lymphoma cells, c-myc is chromosomally translocated to one of the three immunoglobulin (Ig) gene loci and its transcription is driven constitutively by Ig enhancers. Promoter-proximal pausing of pol II is abolished on the translocated c-myc allele. This raised the question whether induction of Ig gene transcription also involves activation of promoter-proximal paused pol II. Here we have studied the transcriptional activation of a functionally rearranged Ig kappa gene in the mouse pre B cell line 70Z/3. We show that pol II pauses approximately 50 bp downstream of the transcriptional start site of the uninduced Ig kappa gene.

Animals↗
PubMed1999

Budding yeast Cdc6p induces re-replication in fission yeast by inhibition of SCF(Pop)-mediated proteolysis.

In fission yeast, overexpression of the replication initiator protein Cdc18p induces re-replication, a phenotype characterized by continuous DNA synthesis in the absence of cell division. In contrast, overexpression of Cdc6p, the budding yeast homolog of Cdc18p, does not cause re-replication in S. cerevisiae. However, we have found that Cdc6p has the ability to induce rereplication in fission yeast. Cdc6p cannot functionally replace Cdc18p, but instead interferes with the proteolysis of both Cdc18p and Rum1p, the inhibitor of the protein kinase Cdc2p. This activity of Cdc6p is entirely contained within a short N-terminal peptide, which forms a tight complex with Cdc2p and the F-box/WD-repeat protein Sud1p/Pop2p, a component of the SCF(Pop) ubiquitin ligase in fission yeast. These interactions are mediated by two distinct regions within the N-terminal region of Cdc6p and depend on the integrity of its Cdc2p phosphorylation sites. The data suggest that disruption of re-replication control by overexpression of Cdc6p in fission yeast is a consequence of sequestration of Cdc2p and Pop2p, two factors involved in the negative regulation of Rum1p, Cdc18p and potentially other replication proteins.

CDC2 Protein Kinase↗
PubMed1999

The family as provider of long-term care: efficiency, equity, and externalities.

The passage of the baby-boom generation into old age raises the prospect of intense pressures on public programs benefiting the elderly, limiting any contemplated expansion of programs serving those needing sustained personal care. This necessitates consideration of comparative efficiency of alternative resources for elder care. I focus on two distinct aspects of such efficiency: productive--the relationship between inputs and outputs--and target--the coincidence of served and those viewed as needing services. I argue that for theoretical reasons family members, specifically children, may be more productive and efficient carers than paid helpers. Furthermore, even if no more efficient than formal providers, care provided by children reduces public expenditures on long-term care. In view of the value to society of children's caregiving activities, if a collective program of long-term care insurance were to be adopted, it should be configured to target its financing and benefits according to family composition.

Caregivers↗
PubMed1999

Time? Money? Both? The allocation of resources to older parents.

We provide estimates of a reduced-form model of the allocation of household time and money resources. We consider four demands for these resources: time spent working, time spent providing care for noncoresident elderly parents, time spent performing housework, and monetary transfers to noncoresident elderly parents. We focus on the effects of wage rates and parental characteristics on the allocation decisions of adult children and their households concerning these four demands. We find that households with individuals earning high wages rely relatively more on cash transfers and relatively less on time transfers than do lower-wage households. We also find evidence consistent with an unmeasured tendency of some families to provide multiple sources of support.

Adult↗
PubMed1999

Wischnewski ulcers and acute pancreatitis in two hospitalized patients with cirrhosis, portal vein thrombosis, and hypothermia.

Accidental hypothermia has been described in the forensic literature but reports of occurrence in hospitalized patients are rare. Associated anatomic lesions include acute hemorrhagic pancreatitis and characteristic acute gastric ulcers termed Wischnewski ulcers. We report here two patients with cirrhosis and ascites; one also had hepatocellular carcinoma. Portal vein thrombosis, acute hemorrhagic pancreatitis and Wischnewski ulcers were present in both. The clinical records documented hypothermia that progressed over several days. Temperature nadirs of 31.0 degrees C (87.8 degrees F) and 32.2 degrees C (90.0 degrees F) were recorded in each patient, respectively, one day before death, although each transiently reached temperatures that did not register on standard monitoring devices. This is the first report that chronicles antemortem body temperatures in hypothermic patients with Wischnewski ulcers and pancreatitis at autopsy. Also, the association of these findings with portal vein thrombosis and cirrhosis has not been previously described. We discuss this constellation of findings with regard to possible mechanistic interrelations.

Acute Disease↗
PubMed1998

Cell cycle: oiling the gears of anaphase.

The anaphase-promoting complex (APC) or cyclosome directs the ubiquitination and destruction of proteins that control specific steps in mitosis. Recent studies show that APC activity requires WD40 domain proteins, and that one of these proteins is part of the checkpoint control that ensures accurate chromosome segregation.

Anaphase↗
PubMed1998

Death following minor head trauma in two adult individuals with the Chiari I deformity.

Reports of sudden death associated with minor head trauma in adults with the Chiari I malformation are rare. We describe two such cases, occurring in a 71-year-old individual and in a 22-year-old individual. In both cases, postmortem examination revealed prominent cerebellar tonsillar herniation and, in one, associated tonsillar sclerosis and hydrocephalus. Evidence of trauma was minimal in both, and was not sufficient to explain these fatalities. We describe the neuropathologic findings and discuss the differential diagnostic considerations in these cases of sudden death. The implications regarding manner of death are also emphasized.

Adult↗
PubMed1997

Nitric oxide attenuates reoxygenation-induced ICAM-1 expression in coronary microvascular endothelium: role of NFkappaB.

Enhanced leukocyte adhesion has been shown to occur in post-ischemic reperfused hearts due to the upregulation of specific cell-surface adhesion molecules. Therefore, we investigated the influence of 4 h of reoxygenation after 20 h of hypoxia on ICAM-1 induction in primary cultures of rat coronary microvascular endothelial cells (CMEC). ICAM-1 surface expression as well as oxygen free radical formation were measured by flow cytometry. Changes in ICAM-1 mRNA levels were assessed by Northern blot and activation of NFkappaB and AP-1 signalling were analysed by electrophoretic mobility shift assays (EMSA) in CMEC lysates. Although hypoxia alone did not affect cell-surface ICAM-1 expression, 4 h of reoxygenation induced a significant upregulation of ICAM-1. ICAM-1 mRNA could not be found after hypoxia alone, but could be detected as early as 1 h following reoxygenation. Unlike AP-1, the activation of which could be detected in CMEC lysates following hypoxia alone, NFkappaB binding activity was induced only following reoxygenation, concurrent with an increase in the formation of reactive oxygen species (ROS). A proteasome inhibitor, nor-Leu (25 microM) inhibited NFkappaB activation by reoxygenation and ICAM-1 expression. Blockade of endogenous nitric oxide (NO) synthesis in CMEC with L-nitroarginine (10 microM) accentuated post-reoxygenation ICAM-1 expression. Finally, an exogenous NO donor, S-nitrosoacetyl-penicillamine (SNAP, 100 microM), suppressed the generation of ROS upon reoxygenation, and blocked the activation of NFkappaB and the upregulation of ICAM-1. Thus, ICAM-1 upregulation in CMEC primary cultures is not induced by hypoxia alone, but appears shortly after reoxygenation in the absence of exogenous cytokines or inflammatory cells. Because upregulation of AP-1 through hypoxia alone did not affect ICAM-1 expression, we conclude that redox-sensitive NFkappaB activation triggers ICAM-1 upregulation. NO inhibits reoxygenation-specific ICAM-1 upregulation, most likely by diminishing oxidative stress that leads to NFkappaB activation.

Animals↗
PubMed1997

The division of family labor: care for elderly parents.

We consider the division of caregiving efforts among the children of older, functionally limited parents. Our model of parental care assumes that care decisions are made in the context of an extended family, with each child taking into account not only the parent's needs and the child's own circumstances, but also the characteristics and actual care behavior of siblings. We propose a simultaneous-Tobit statistical framework that embodies these assumptions. The model is estimated using data from the 1993 Asset and Health Dynamics Among the Oldest Old (AHEAD) study. The findings indicate that a child's hours of parent care are reduced, but on much less than a one-for-one basis, as the parent-care hours of siblings increase. We also find that a child's supply of parent-care hours is reduced by having sisters, holding constant the care efforts of siblings.

Aged↗