The TP1 isolate of feline sarcoma virus encodes a fgr-related oncogene lacking gamma actin sequences.

We have isolated a new feline sarcoma virus, TP1-FeSV. The virus encodes a myristilated 83 kD gag-onc fusion protein displaying tyrosine kinase activity. We have established nonproducer cell lines lacking the TP1-FeSV associated helper virus (FeLV) and TP1-FeSV transfected NIH cell lines. Southern Blot analysis of genomic DNA and Northern Blot analysis of RNA isolated from these cell lines revealed that the oncogene of the TP1-FeSV isolate is related to the fgr oncogene of the GR-FeSV, but shows no hybridization to the gamma actin homologous sequences of the GR-FeSV. We have isolated TP1-FeSV specific clones from a genomic library. Restriction enzyme and sequence analysis showed that the TP1-FeSV genome consists of the first 1651 nucleotides of the gag gene, followed directly by fgr sequences. The TP1-FeSV fgr sequence starts 43 nucleotides after the beginning of the GR-FeSV fgr sequence. In contrast to the GR-FeSV fgr which has lost 13 amino acids of the c-fgr carboxy terminus, the TP1-FeSV fgr contains the complete carboxy terminus of the cellular fgr gene. The TP1-FeSV fgr sequence is followed by a unique 328 nucleotide long sequence of unknown origin. The 3' recombination site occurs within the pol gene, 460 nucleotides from the start of the env leader sequence. Comparison of the subcellular localization of the transforming proteins of TP1-FeSV and GR-FeSV show no striking difference; both molecules are in part associated with subcellular membrane/cytoskeletal fractions and form complexes with the cellular pp90 and pp50.

Plasmodium falciparum calcium-dependent protein kinase phosphorylates proteins of the host erythrocytic membrane.

The unusual Ca(2+)-dependent protein kinase from Plasmodium falciparum (PfCPK) [1], whose gene structure and expression in bacteria have been reported [1], was purified to homogeneity. The purified recombinant kinase has a native molecular mass of 62,000, is activated by Ca2+ (K0.5 = 15 microM) in the presence of Mg2+ or Mn2+, and can associate with 45Ca2+. The activation by Ca2+ could be partially replaced by Mn2+, but not by Zn2+ or Mg2+. PfCPK preferentially phosphorylated casein and histone H1. The Km and Vmax for Mg2+ ATP were 26 microM and 70 nmol min-1 mg-1, respectively, with casein as substrate; and 34 microM and 143 nmol min-1 mg-1, respectively, with histone H1 as substrate. The kinase undergoes autophosphorylation on both serine and threonine residues. Calmodulin antagonists (calmidazolium, trifluoperazine, N-[6-aminohexyl]-5-chloro-1-napthalene-sulfonamide, and ophiobolin A) could inhibit the kinase activation, but much higher concentrations of the antagonists are needed than was required to inhibit calmodulin-mediated effects. PfCPK preferentially phosphorylates proteins of the host erythrocytic membrane in vitro but phosphorylates parasitic proteins only to a minor extent. The selectivity of the phosphorylation may be partially controlled by phosphatidylserine which is bound to some of the erythrocytic membrane proteins. Using a rabbit polyclonal antiserum against the recombinant protein, the kinase was found to be mainly expressed in the ring and schizont stages, and mainly localized in the parasitic membrane-organelle fraction and partially localized on the erythrocytic membrane.

Localization and stage specific phosphorylation of Plasmodium falciparum phosphoproteins during the intraerythrocytic cycle.

Fifty-nine Plasmodium falciparum specific phosphoproteins with molecular weights between 15,000 and 192,000 were analyzed by SDS-PAGE and two-dimensional gel electrophoresis. 40 phosphoproteins were identified by [gamma-32P]ATP labeling of cell lysates, 19 by [32P]orthophosphate labeling of parasitic cultures in vivo. Changes in the phosphorylation pattern during the infectious erythrocytic cycle were determined for all proteins. In parallel, cell fractionation studies were done to follow up possible changes in the cellular distribution of these proteins. Several phosphoproteins are associated with the membrane fraction of infected erythrocytes. One pair of proteins of approx. 88 kDa and a pI of about 5.0 was further characterized. Both proteins are located in the parasitic fractions as well as in the membrane of infected erythrocytes during the entire cycle. Phosphorylation of these proteins, however, is restricted to the trophozoite and schizont stages. Peptide mapping studies demonstrated that both proteins are identical with the exception of minor modifications which are probably not the result of differences in phosphorylation.

Plasmodium falciparum protein kinase 5 and the malarial nuclear division cycles.

In the course of our studies on cell cycle regulation mechanisms of Plasmodium falciparum, we investigated expression pattern, kinase activity, and localization of PfPK5, a putative malarial member of the family of cyclin-dependent protein kinase (cdks). The kinase was immunoprecipitated from parasites of selected stages and from parasites blocked with the cell-cycle inhibitor aphidicolin. An elevated kinase activity of PfPK5 from aphidicolin-blocked cells suggested that the enzyme might be implicated in the regulation of the parasite's S-phase. To further investigate this hypothetical function, parasite cultures were treated with the specific cdk inhibitors flavopiridol and olomoucine, which act on PfPK5 in vitro at similar concentrations as on other cdks. When applied during the nuclear division cycles of the parasite, both drugs markedly inhibited the DNA synthesis, as predicted from our proposition that PfPK5 is necessary to activate or maintain the parasite S-phase. Immunolocalization studies provide further evidence for this potential role of PfPK5.

A Plasmodium falciparum protein kinase with two unusually large kinase inserts.

A protein kinase gene (PfPK1) has been isolated from the human parasite Plasmodium falciparum by using a mixed oligonucleotide pool which corresponds to a highly conserved region of serine/threonine protein kinases. The gene, which contains one intron, encodes a protein with a predicted length of 909 amino acids. The predicted protein contains all the conserved sequences characteristic of a protein kinase catalytic domain. These sequences are discontinuous, however, since they are separated by two large kinase inserts with 178 and 330 amino acids in size. Specific antisera were raised against recombinant fragments of the protein and a PfPK1-specific peptide. Using one of these antibodies, a functional protein kinase was precipitated from malarial lysates and this kinase recognized casein as an exogenous substrate. PfPK1 was expressed in a stage-specific fashion and also had a stage-specific cellular localization. During the intraerythrocytic life cycle, PfPK1 shifts from the parasite cytosol to the parasite membrane fraction. An unusual feature of PfPK1 is its electrophoretic mobility on SDS-PAGE. Whereas the predicted protein size is about 100 kDa, the apparent size is about 70 kDa. There are no indications for RNA processing and we could exclude proteolytic processing as an explanation.

Two major phosphoproteins of Plasmodium falciparum are heat shock proteins.

Two major phosphoproteins of Plasmodium falciparum could be identified by partial amino acid sequencing as the plasmodial members of the hsp 70 heat shock protein family, Pfhsp and Pfgrp. According to phosphoamino acid analyses of Pfhsp and Pfgrp isolated from [32P]orthophosphate-labeled malarial cultures, both proteins were phosphorylated in Ser and Thr. While Pfhsp contains higher amounts of labeled phosphoserine, Pfgrp contains higher amounts of phosphothreonine. Phosphorylation of both proteins increased throughout the entire erythrocytic growth cycle. At the trophozoite and schizont stages Pfhsp and Pfgrp are the most prominent phosphoproteins of Plasmodium falciparum. Using multiply redundant oligonucleotides directed against the N-terminus of Pfgrp we cloned and sequenced the entire Pfgrp gene. The gene encodes a product with a predicted length of 652 amino acids. The deduced amino acid sequence has identities of 65.5% and 65.0% to the human and rat grp78 proteins, respectively. Pfgrp possesses a classical N-terminal leader sequence. The published grp78 related gene sequences of Plasmodium falciparum are all fragments of the same plasmodial gene.

Stereotypes concerning normal and handicapped children.

Individuals' attitudes were assessed toward various groups of children. In study 1 the respondents were 65 male and female teachers from across the state of Kansas. In study 2 the respondents were 89 men and women in attendance at the 1978 International Conference of the Association for Children with Learning Disabilities. In both studies the evaluations of the labels "gifted children," "normal children," and "physically handicapped children" were found to be significantly more positive than the evaluations of the labels "mentally retarded children," "learning disabled children," and "emotionally disturbed children." These results seem to indicate that definite negative stereotypes are held toward the latter three groups of children. In study 1 these findings were found to occur generally regardless of the respondents' sex, age, educational level attained, and amount of previous mainstreaming experience.

Quantitative observations of hymens in prepubescent females selected for non-abuse.

The maximum hymenal opening was evaluated quantitatively in 111 prepubescent females during routine physical examinations in a pediatric subspecialty office. For comparison, an additional 53 females referred by child protective agencies were also examined. Analysis of data show "non-abused" groups may be separated statistically from "abused" groups on the basis of area of hymenal opening. The mean area for the "non-abused, non-masturbate" group was 6.4 mm2. The upper limit of area (mean + 3 S.D.) of hymenal opening in this group was 24.1 mm2. A child having a hymenal opening diameter of 6.94 mm or less has a 99% chance of being in the "non-abused" group. The area of hymenal opening for "non-abused" groups did not change with increasing age, height or weight. A skilled pediatrician knowledgeable in the area of sexual abuse may obtain clinically relevant information with ordinary office equipment and trained personnel. Regular and repeated observations of genitalia during routine health maintenance examinations are vital baseline measurements for the physical and mental health of young female patients.

Psychological and psychophysiological factors in prevention and treatment of cold injuries.

Cold injured patients in Alaska come from many sources. Although sport and work continues to provide large numbers of cold injured, most severe repeat injuries tend to reflect other biopsychosocial consequences. Certain behaviors can increase the probability of injury, however all persons living in cold climates are potential candidates. One can decrease risk by education, knowledge and intelligent behavior. Proper respect for adequate protection and hydration seem to be critical factors. Understanding the psychological, physiological and psychophysiological aspects of the cold environment performer helps refine the prevention and treatment strategies for cold injury. Skill training with bio-behavioral methods, such as thermal biofeedback, and the value of medical psychotherapy appear to offer continued promise by facilitating physiologic recovery from injury, as well as assisting in long term rehabilitation. Both approaches increase the likelihood of a favorable healing response by soliciting active patient participation. Medical Psychotherapy for traumatic injuries can also help identify and manage cognitive emotional issues for families and patients faced with the permanent consequences of severe thermal injuries. Thermal biofeedback therapy has the potential benefit of encouraging greater self-reliance and responsibility for self-regulating overall health by integrating self-management skills regarding physiology, diet and lifestyle. Inpatient and outpatient biofeedback training offers specific influence over vascular responses for healing, as well as providing an effective tool for pain management. Interest in cold region habitation has continued to expand our study of human tolerance to harsh, extreme environments. Biological, psychological, sociological, and anthropological views on adaptation, habituation, acclimatization, and injury in cold environments acknowledges the role of development, learning and educated responses to cold environments. The study of health, performance, and injury prevention in extreme isolated cold environments has important strategic and scientific implications. What is learned from behavioral studies of cold survival provides an opportunity to increase our scientific knowledge and understanding. These cold research findings can assist in our future exploration of cold, underwater farming at great depths, and to far distance space travel to cold planets. The relatively new research frontier "Polar Psychology" has evolved to study how interactions with cold environments can have both positive and/or negative consequences. This research simulates the psychological factors likely to be encountered while exploring isolated cold regions of distant galaxies. The psychological and psychophysiological correlates of cold experience appear to be a function of four interactive issues: the environment, genetic predisposition, learning or experience, and finally perception or cognition. Individual cold tolerance seems to relate heavily on sensation, perception and behavior.(ABSTRACT TRUNCATED AT 400 WORDS)

Effects of neutral posture on muscle tension during computer use.

This study focused on developing a new approach to seated work positions was conducted on 67 office workers who use a Visual Display Terminal (VDT) as a major function of their working day. Muscle tension was measured by surface electromyography (sEMG) while participants were asked to adopt 4 selected working postures. Pain was measured before and after ergonomic intervention on the Nordic scale, which was modified for this study. Adjustable workstations were used to place participants in desired positions during the clinical testing sessions and the extended intervention period. Results indicate the effects of this ergonomic intervention may have positive effects on muscle tension and pain, significant enough to encourage employers to implement training and workstation modifications following these guidelines.

Sequence effects of relaxation training, EMG, and temperature biofeedback on anxiety, symptom report, and self-concept.

Assessed the effects of particular treatment combinations of relaxation training, temperature, and EMG biofeedback on state-trait anxiety, symptom report, and self-concept. The four groups received one of the following sequences: (a) relaxation training, temperature, and EMG biofeedback; (b) temperature, EMG biofeedback, and relaxation; (c) temperature followed by EMG biofeedback; (d) EMG biofeedback followed by temperature. A sample of 37 volunteers participated in 16 20-minute training sessions over an 11-week period, which totaled 800 appointments. Training was found equally effective for decreasing frontalis EMG and increasing finger skin temperature, regardless of sequence. Most substantial improvement occurred after 8 sessions, whereas little improvement was found after 16 sessions. Each group became increasingly homogeneous over time on all measures.

Contingent vs. noncontingent EMG feedback and hand temperature in relation to anxiety and locus of control.

This study was designed to measure the effects of contingent and noncontingent EMG feedback on hand temperature, anxiety, and locus of control. Two groups of six subjects each were selected on the basis of high test-anxiety scores. The groups participated in a reverse design study in which Group 1 received five sessions of contingent EMG ffedback followed by five sessions of noncontingent feedback. Group 2 received noncontingent feedback followed by contingent feedback. Results indicate a significant order of treatment effect. Subjects who received contingent feedback first produced lower EMG readings, lower test-anxiety scores, and higher hand temperatures during noncontingent feedback sessions. Receiving noncontingent feedback first may actually have interfered with utilizing contingent feedback.

Cognitive and physiological feedback on cold pain tolerance.

Results supported the relevancy of cognitive information effects on pain tolerance, in that subjects who were given a rational and accurate explanation of what to expect showed greater tolerance than those who received irrelevant information. Accurate monitoring of hand temperature did not seem necessarily advantagous as an influence on pain tolerance. It appears merely watching a monitor, regardless of the specific contents of the screen, resulted in longer hand immersion times when compared to no monitor. The monitors seem to serve as distractors and specificity of physiological information was not particularly useful. However, neither information nor physiological monitoring emerged as the primary influence on pain tolerance in this study. Instead, the strongest predictors found were motivation and self-efficacy. The subject's own self prediction of anticipated performance with cold induced pain was closely consistent with actual performance. Although these results alone may not generalize to extended field situations, this study does reinforce the general findings of previous research: namely Bandura's (10) evidence on self-efficacy. While it is obvious cold temperatures have measurable physiological consequences, the experience of pain is also psychologically mediated. Pain associated with cold injury and frostbite in hospital studies show personality correlates are significantly related to the frequency, severity and tragedy of subsequent results (15). A replication of this study will include male subjects even though it is anticipated that findings will be consistent, with perhaps longer immersion times. Future research may want to develop training strategies aimed at teaching self-efficacy and realistic expectations of potential consequences in cold environments rather than scare tactics regarding physiological and psychological cold pain tolerance.

Biofeedback vs. video games: effects on impulsivity, locus of control and self-concept with incarcerated juveniles.

Research has found hyperactivity, poor impulse control, impaired sustained attention and low self-concept to be behavioral deficits common to juvenile delinquents. Limited opportunities for exercising self-control while incarcerated may encourage helplessness. If biofeedback training enhances self-regulation skills, then perhaps these behaviors can be taught in confinement. A sample of 12 felonious juvenile residents (aged 15-18) from a highly restricted environment were assigned randomly to a biofeedback or video game group and trained for 10 half-hour sessions. Results indicated virtually no significant differences between biofeedback and video game training. However, pre and post differences for both groups combined demonstrated significant gains in impulsivity, EMG, and self-concept. Both groups rated themselves equally on self-control ability, regardless of training. Further comparisons between other institutionalized residents (N = 14) and staff counselors (N = 10) as non-treatment controls were made. On each measure, both training groups improved consistently and became more like their less restricted counterparts.

Gene structure and expression of an unusual protein kinase from Plasmodium falciparum homologous at its carboxyl terminus with the EF hand calcium-binding proteins.

An unusual protein kinase gene, termed PfCPK, was isolated from Plasmodium falciparum. The gene, which contains five exons and four introns, encodes a product with a predicted length of 524 amino acids. The amino-terminal segment of the predicted protein contains all of the conserved sequences characteristic of a protein kinase catalytic domain and has a high homology to several protein serine-threonine kinase subfamilies (30-41% amino acid identities). These subfamilies include calcium/calmodulin-dependent protein kinases, calcium-dependent protein kinase, ribosomal S6 protein kinase, cyclic nucleotide-dependent protein kinases, protein kinase C, and the yeast SNF1 subfamily. All of these protein kinases are relatively close in the phylogeny tree and within the kinase catalytic domains have about 35% amino acid identities to each other, suggesting that PfCPK is also in this region of the phylogeny tree. An unusual feature of PfCPK is that its carboxyl-terminal segment displays homology to the EF hand calcium-binding proteins, for example 34% amino acid identity to chicken fast skeletal muscle troponin C and 35% amino acid identity to human calmodulin. Like troponin Cs and calmodulins, PfCPK also contains four EF hand calcium-binding motifs. Furthermore, the four introns in the PfCPK gene are all located in the carboxyl-terminal putative EF hand calcium-binding region (EF hand calcium-binding proteins from higher eukaryotes generally contain multiple introns). This combination of a protein kinase and an EF hand calcium-binding protein in a single polypeptide implies that PfCPK may be directly activated by calcium. Constructs containing the full-length PfCPK cDNA have been expressed in Escherichia coli at a high level to generate a 60-kDa recombinant protein. Compared with similar fractions from control cells, the fraction containing PfCPK recombinant protein exhibited an elevated protein kinase activity which was Ca(2+)-dependent.

The cell cycle in protozoan parasites.

Research into cell cycle control in protozoan parasites, which are responsible for major public health problems in the developing world, has been hampered by the difficulties in performing classical genetic analysis with these organisms. Nevertheless, in a large part thanks to the data gathered in other eukaryotic systems and to the acquisition of the sequences of parasite genes homologous to cell cycle regulators, many molecular tools required for an in-depth study of the cell cycle in protozoan parasites have been collected over the past few years. Despite the considerable phylogenetic divergence between these organisms and other eukaryotes, and notwithstanding important specificities such as the apparent lack of checkpoints during cell cycle progression, available data indicate that the major families of cell cycle regulators appear to operate in protozoan parasites. Functional studies are now needed to define the precise role of these regulators in the life cycle of the parasites, and to possibly validate cell cycle control elements as potential targets for chemotherapy.