Telomerase in kinetoplastid parasitic protozoa.

We have identified telomerase activity in extracts of three evolutionarily diverse kinetoplastid species: Trypanosoma brucei, Leishmania major, and Leishmania tarentolae. Telomerase activity was initially detected in extracts from insect form cells of all three kinetoplastid species by using a modification of the one-tube telomere repeat amplification protocol [Kim, N., et al. (1994) Science 266, 2011-2015], although better results were subsequently achieved with the two-tube telomere repeat amplification protocol [Autexier, C., Pruzan, R., Funk, W. & Greider, C. (1996) EMBO J. 15, 5928-5935]. The activity in T. brucei extracts was sufficiently robust to enable its detection in a direct assay of telomerase; enzyme processivity was found to be relatively low. The in vitro properties of telomerase suggest a possible templating domain sequence for the telomerase RNA of T. brucei. Telomerase activity is likely to contribute to telomere maintenance in these parasitic organisms and provides a new target for chemotherapeutic intervention.

Dungan model of dynamic integration.

TOPIC: The Dungan Model of Dynamic Integration (DMDI), which views humans as having three synergistic dimensions of body, mind, and spirit. PURPOSE: To present a new nursing model for practice and education. SOURCE: This is the author's model and it is composed of concepts and constructs that are well-documented in nursing literature. CONCLUSIONS: Dynamic Integration is a broad conceptual model that allows for testing a number of mid-range theories to add to the science and discipline of nursing.

The SLA RNA gene of Trypanosoma brucei is organized in a tandem array which encodes several small RNAs.

We have recently identified the Spliced Leader Associated RNA (SLA RNA) which is implicated in pre-messenger RNA splicing in Trypanosoma brucei by virtue of its interaction with the 5' splice site of the trans spliced spliced leader RNA (SL RNA) in vivo. Southern analyses reveal that the SLA RNA gene is found in a tandem array of 10-11 copies per haploid genome in T. brucei. Each repeat unit in the array encodes three additional small RNAs of unknown function. RNA polymerase inhibition studies are consistent with transcription of all four genes by the same polymerase, but do not clearly differentiate between RNA polymerases II and III. The SLA RNA has homologs in other kinetoplastid protozoa and we have determined the sequence from two additional species. Trypanosoma cruzi and Crithidia fasciculata. Features of both secondary structure and sequence are conserved in these organisms. One conserved element, 5'-UGUAGUG-3', has the potential to base-pair to the SL RNA upstream of the 5' splice site. This potential interaction is consistent with the sites of SL RNA to SLA RNA psoralen cross-linking in vivo [1].

Managing spiritual distress in patients with advanced cancer pain.

Cancer threatens our very existence; when pain is also present, suffering is compounded and leads to spiritual distress. This article presents a nursing strategy to deal with the responses of clients and families in such complex situations. Four case studies were analyzed to describe the methodology and applicability of an approach based upon the Dungan model of dynamic integration. The goal of the first appointment was to teach clients how to effectively deal with the advanced cancer pain that brought them to the center. Presence, caring, genuine concern, and active listening facilitated the therapeutic relationship and were the hallmarks of nursing modalities. Daily appointments with the nurse continued for 6-10 days while clients were also being treated by medical staff. Trust was established by reaching the mutual goal of giving the client control over pain. Spiritual assessment confirmed the diagnosis. Grief was a characteristic of spiritual distress experienced by all clients. Therefore, grief counseling was instituted early and reinforced daily. Spiritual counseling, cognitive reframing, crisis intervention, and reminiscence were used selectively. Because of the complexity of problems faced by families, multi-disciplinary referrals were required by all clients. Outcomes were evaluated positively by participants. All clients increased hope, participated in the grief process, and integrated the possibility of death in such a way as to promote personally defined "quality of life" through their waning days.

Evidence for the presence of a small U5-like RNA in active trans-spliceosomes of Trypanosoma brucei.

The existence of the Trypanosoma brucei 5' splice site on a small RNA of uniform sequence (the spliced leader or SL RNA) has allowed us to characterize the RNAs with which it interacts in vivo by psoralen crosslinking treatment. Analysis of the most abundant crosslinks formed by the SL RNA allowed us previously to identify the spliced leader-associated (SLA) RNA. The role of this RNA in trans-splicing, as well as the possible existence of an analogous RNA interaction in cis-splicing, is unknown. We show here that the 5' splice site region of the SL RNA is also crosslinked in vivo to a second small RNA. Although it is very small and lacks a 5' trimethylguanosine (TMG) cap, the SLA2RNA possesses counterparts of the conserved U5 snRNA stem-loop 1 and internal loop 1 sequence elements, as well as a potential trypanosome snRNA core protein binding site; these combined features meet the phylogenetic definition of U5 snRNA. Like U5, the SLA2 RNA forms an RNP complex with the U4 and U6 RNAs, and interacts with the 5' splice site region via its putative loop 1 sequence. In a final analogy with U5, the SLA2 RNA is found crosslinked to a molecule identical to the free 5' exon splicing intermediate. These data present a compelling case for the SLA2 RNA not only as an active trans-spliceosomal component, but also for its identification as the trypanosome U5 structural homolog. The presence of a U5-like RNA in this ancient eukaryote establishes the universality of the spliceosomal RNA core components.

Health maintenance for the independent frail older adult: can it improve physical and mental well-being?

This descriptive study measured outcomes of a health maintenance programme (HMP) of regular exercise, health teaching and group participation on physical and mental well-being in a convenience sample of frail older adults living independently. Hand strength, range of motion (flexibility) and blood pressure were used as indicators of physical well-being. Self-esteem and life satisfaction were used as indicators of mental health. A pre-test/post-test design was used taking all measurements before starting the programme and after a 6-month interval of participation. Paired t-tests of mean change scores, with each participant acting as his or her own control, showed statistical improvement in systolic blood pressure and range of motion in right ankle and in self-esteem and life satisfaction using visual analogue scales. Clinical improvement was demonstrated in all measurements and by participant evaluation of the programme.

A holistic protocol for management of fluid volume excess in hemodialysis patients.

Patients on hemodialysis as treatment for end stage renal disease (ESRD), are at high risk for ineffective management of their therapeutic regimen because it entails pervasive changes in lifestyle. This article describes the experiences of five patients who participated in a nursing protocol of 8-10 therapeutic interviews associated with dialysis treatment over a 5-week period. The Dungan Model of Dynamic Integration provided the conceptual framework for holistic management strategies of teaching, counseling, and social support to assist patients to integrate required changes into their lifestyles. Fluid volume excess (FVE) was used as an indicator of ineffective management of therapeutic regimens. Presence of FVE, indicated by weight gain, presence of peripheral edema, and abnormal lung sounds were assessed before each dialysis treatment. Findings confirmed that these patients were at high risk for ineffective self-management and that the approach of augmented professional support used in the protocol was valuable to patients and families during this difficult transition.

A relaxation protocol to reduce patient anxiety.

Relaxation and music therapy have shown promise as anxiety-reducing interventions for patients in critical care settings. The challenge for nurses is to incorporate these modalities of care into effective clinical strategies. This article describes one method of introducing relaxation therapy to the anxious patient within the context of the Dungan Model of Dynamic Integration.

Identification of a small RNA that interacts with the 5' splice site of the Trypanosoma brucei spliced leader RNA in vivo.

In vivo psoralen cross-linking of the trypanosome spliced leader (SL) RNA has led to the discovery of a small RNA that we provisionally call the spliced leader-associated (SLA) RNA. The 72 nt SLA RNA is unlike any known small RNA except for a small region that resembles U5 snRNA. The SL/SLA RNA cross-links map to two regions, the predominant interactions occurring between the 5' splice site region of the SL RNA and a CUUUUA sequence in the SLA RNA. The resemblance between these cross-links and interactions of U5 snRNA with cis-spliced pre-mRNAs suggests that the SLA RNA may be the trans-splicing analog of U5 snRNA in trypanosomes.

Nursing strategy for initial emotional response to cancer diagnosis.

Nursing strategy for patients' initial emotional response to cancer was conducted with five clients and evaluated for effectiveness using a case study approach. This clinical evaluation of a management strategy was conducted in an acute-care setting during the phase of initiating a therapeutic relationship. The nurse identified stressors, support systems, usual coping strategies, and level of knowledge about the disease. Two meetings a week were conducted with each client during hospitalization. Evaluation was completed on four of the five clients. The management strategy facilitated beginning reintegration of body, mind, and spirit resulting in a smoother transition phase. The aim of this protocol was to initiate effective coping responses to a potentially devastating diagnosis.

Thermally perturbed rhodanese can be protected from inactivation by self-association.

A fluorescence-detected structural transition occurs in the enzyme rhodanese between 30-40 degrees C that leads to inactivation and aggregation, which anomalously decrease with increasing protein concentration. Rhodanese at 8 micrograms/ml is inactivated at 40 degrees C after 50 min of incubation, but it is protected as its concentration is raised, such that above 200 micrograms/ml, there is only slight inactivation for at least 70 min. Inactivation is increased by lauryl maltoside, or by low concentrations of 2-mercaptoethanol. The enzyme is protected by high concentrations of 2-mercaptoethanol or by the substrate, thiosulfate. The fluorescence of 1,8-anilinonaphthalene sulfonate reports the appearance of hydrophobic sites between 30-40 degrees C. Light scattering kinetics at 40 degrees C shows three phases: an initial lag, a relatively rapid increase, and then a more gradual increase. The light scattering decreases under several conditions; at increased protein concentration; at high concentrations of 2-mercaptoethanol; with lauryl maltoside; or with thiosulfate. Aggregated enzyme is inactive, although enzyme can inactivate without significant aggregation. Glutaraldehyde cross-linking shows that rhodanese can form dimers, and that higher molecular weight species are formed at 40 degrees C but not at 23 degrees C. Precipitates formed at 40 degrees C contain monomers with disulfide bonds, dimers, and multimers. We propose that thermally perturbed rhodanese has increased hydrophobic exposure, and it can either: (a) aggregate after a rate-limiting inactivation; or (b) reversibly dimerize and protect itself from inactivation and the formation of large aggregates.

A chaperone-mimetic effect of serum albumin on rhodanese.

Reactivation of denatured rhodanese (thiosulfate:cyanide sulfurtransferase, EC 2.8.1.1) was found to be aided by the presence of serum albumin. Both the rate and the extent of reactivation of the urea-denatured enzyme were optimal at low rhodanese and moderate serum albumin concentrations. Similarly, stabilization of the sulfurtransferase activity of rhodanese that had been partially unfolded at 40 degrees C was aided by the presence of serum albumin. All the observations are in accord with a model in which enzyme that has been partially refolded from the urea-denatured state or partially unfolded thermally interacts directly with serum albumin in a way that prevents rhodanese self-association. Serum albumin thus acts as a molecular chaperone in these systems.

Chylothorax: a complication of thoracic trauma or surgery.

Chylothorax is a troublesome complication of thoracic trauma and surgery. The critical care nurse's assessment of respiratory changes is frequently the first sign that this serious complication is occurring. Chylothorax may cause a previously stable thoracic patient to develop sudden, potentially serious respiratory problems.

Long-term lifestyle management.

The Clinical Nurse Specialist impacts the client with chronicity by strategic management of cost and quality outcomes. Three enabling strategies: counseling, social support, and teaching-learning are the tools the nurse and client use to implement long-term lifestyle adjustment to chronicity. Reduced health care system utilization and increased self-esteem and self-care abilities are the result of the strategies. Lifestyle management warrants further nursing research.

Processive recombination by the phage Mu Gin system: implications for the mechanisms of DNA strand exchange, DNA site alignment, and enhancer action.

The Gin DNA invertase of bacteriophage Mu carries out processive recombination in which multiple rounds of exchange follow synaptic complex formation. The stereostructure of the knotted products determined by electron microscopy establishes critical features of site synapsis and DNA exchange. Surprisingly, the invertase knots substrates with directly repeated sites as well as those with inverted sites. The results suggest that the Gin synaptic complex contains three mutually perpendicular dyads; one is the axis of site rotation during exchange, and they cause inverted and direct site substrates to form a similar synaptic complex. The extensive knotting by Gin has implications for the energetics of recombination and shows that the enhancer for recombination is required only at an early stage, and thus may normally operate in a hit-and-run fashion.

Discovery of a predicted DNA knot substantiates a model for site-specific recombination.

The mechanism of site-specific genetic recombination mediated by Tn3 resolvase has been investigated by a topological approach. Extrapolation of a detailed model of synapsis and strand exchange predicts the formation of an additional DNA product with a specific knotted structure. Two-dimensional gel electrophoresis of DNA reacted in vitro revealed a product, about 0.1 percent of the total, with the appropriate mobility. A technique for determining DNA topology by electron microscopy was improved such that less than a nanogram of DNA was required. The structure of the knot was as predicted, providing strong evidence for the model and showing the power of the topological method.