Introns are essential for growth-regulated expression of the mouse thymidylate synthase gene.

The thymidylate synthase (TS) gene is expressed at much higher levels in proliferating cells than in quiescent cells. We have been studying the sequences that are important for regulating the mouse TS gene. We previously showed that DNA sequences upstream of the essential promoter elements as well as downstream of the ATG codon are both necessary (but neither is sufficient) for normal regulation in growth-stimulated cells. In the present study, we examined the possible roles of the coding region, polyadenylation signal, and introns as downstream regulatory elements. Minigenes consisting of 1 kb of the TS 5'-flanking region, the coding region (with or without various introns at their normal locations), and polyadenylation signals from the TS gene, the human beta-globin gene, and the bovine growth hormone gene were stably transfected into wild-type mouse 3T6 cells. Minigenes that contained introns 5 and 6, 1 and 2, or 1 alone were regulated regardless of which polyadenylation signal was included. A minigene that contained an internally deleted version of intron 1 was also regulated in response to growth stimulation. However, when all introns were omitted, there was little if any change in the level of minigene expression as cells progressed from G1 through S phase. These observations indicate that TS introns contain sequences that are necessary for normal growth-regulated expression of the mouse TS gene. These sequences appear to be associated with sequences that are important for splicing and to function in cooperation with upstream regulatory elements to bring about normal S-phase-specific expression.     

Combining specialist and primary health care teams for HIV positive patients: retrospective and prospective studies.

OBJECTIVE: To develop and evaluate a model of health care for HIV positive patients involving specialist, hospital based teams and primary health care teams. DESIGN: One year retrospective and a 2 1/2 year prospective study. SETTING: Two hospitals in West London and 88 general practitioners in 72 general hospitals. SUBJECTS: 209 adults with HIV infection. INTERVENTION: General practitioners enrolled in the project were faxed structured outpatient clinic summaries. When hospital inpatients were discharged, a brief discharge summary was faxed. General practitioners had access to consultant physicians skilled in HIV medicine through a 24 hour mobile telephone service. An HIV/AIDS management and treatment guide containing relevant local information was produced. Quarterly discussion forums for general practitioners were held, and a regular newsletter was produced. MAIN OUTCOME MEASURES: Hospital attendance and general practitioner consultations; perceived benefits and problems of patients and general practitioners. RESULTS: The average length of a hospital inpatient stay was halved for those patients who had participated in the project for two years, and the average number of visits to the outpatient clinic per month fell for patients with AIDS. There was a substantial increase in the number of visits to general practitioners by patients with AIDS and symptomatic HIV infection. Patients and general practitioners both felt that the standard of health care provided had improved. CONCLUSIONS: This model of health care efficiently and effectively utilised existing teams of hospital and primary health care professionals to provide care for HIV positive patients. Simple, prompt, and regular communication systems which provided information relevant to the needs of general practitioners were central to its success.     

Sulfoxide configuration in sparsomycin determines time-dependent and competitive inhibition of peptidyl transferase

Sparsomycin, S_cR_s configuration, was the most potent of the four possible stereoisomers as a competitive inhibitor of peptide bond formation. In addition, the configuration of the two chiral centers dictated whether the compound exhibited time- and temperature-dependent inhibition of peptidyl transferase when incubated with polysomes prior to enzyme assay. The data corroborate the thesis that a peptidyl transferase-mediated acylation of the pivotal sulfoxide moiety and subsequent Pummerer rearrangement play a significant role in the inhibitory properties of sparsomycin.     

Use of the avidin-biotin complex for the localization of actin and myosin with fluorescence microscopy

A new indirect method for fluorescence localization of proteins making use of the avidin-biotin complex is described. We have prepared both a biotin-modified rabbit heavy meromyosin (BHMM) and a biotin-modified antibody to a smooth muscle myosin. After fixation, cells can be treated with either BHMM, which binds to actin, or the biotinyl antibody, which binds to myosin. In a second step the cell are treated with a fluorescent derivative of avidin (Fl-avidin) which binds to the biotinyl proteins and thus indirectly reveals the location of the cellular action or myosin.     

Increased susceptibility to infection with Brugia pahangi in aged female jirds (Meriones unguiculatus)

Female Mongolian jirds, Meriones unguiculatus, from 5 age groups of 2, 12, 16, 21, and 28 months, were infected with Brugia pahangi. Infections were followed for 125 days by weekly bleedings beginning 55 days postinoculation. Jirds were then killed and adult parasites recovered. Results showed a significant shortening of the prepatent period in the 12-, 21-, and 28-month-old groups. The proportion of gravid female worms did not vary significantly among the 5 groups. Similarly, the ratio of females to male worms showed little variation from group to group. Microfilaremia data for the 5 infection groups show an age-associated increase in numbers of circulating microfilariae. Some individuals in the 28-month group demonstrated 1,500 to 3,000 microfilariae per 0.25 ml peripheral blood, a level that was not approached by young females.     

Novel shuttle plasmid vehicles for Escherichia-Streptococcus transgeneric cloning

A novel plasmid vector that is able to replicate both in Escherichia coli and in Streptococcus sanguis is described. This 9.2-kb plasmid, designated pVA856, carries Cm^r, Tc^r and Em^r determinants that are expressed in E. coli. Only the Em^r determinant is expressed in S. sanguis. Both the Cm^r and the Tc^r of pVA856 may be insertionally inactivated. This plasmid affords several different cleavage-ligation strategies for cloning in E. coli followed by subsequent introduction of chimeras in to S. sanguis. In addition, we have modified a previously described E. coli-S. sanguis shuttle plasmid [pVA838; Macrina et al., Gene 19 (1982) 345–353], so that it is unable to replicate in S. sanguis. The utility of such a plasmid for cloning and selecting sequences enabling autonomous replication in S. sanguis is demonstrated.     

THE SYSTEMATIC POSITION OF EOGINKGOITES

Eoginkgoites is a fan-shaped, imparipinnate leaf with a short rachis and long petiole. It was first described from the Upper Triassic Newark Group of Pennsylvania by Bock who assigned the fossil to the Ginkgoales. The fossil has also been found in the Upper Triassic Chinle Formation in Utah and Arizona and in the Newark Group in North Carolina. Investigation of the well-preserved specimens found in the Chinle Formation shows that the leaf has anastomosing venation, a marginal vein and paracytic (syndetocheilic) stomata. These characters indicate that the leaf is bennettitalean and Eoginkgoites is reassigned to the Bennettitales although its shape is perplexing. Eoginkgoites may be an important index fossil to the lower Upper Triassic (middle Carnian) rocks of North America.     

Optimum concentration and pH of phosphate buffer for assay of cytochrome c oxidase in intact plant mitochondria

For measurement of cytochrome c oxidase activity in intact plant mitochondria the optimum concentration of K-Pi buffer and pH in the reaction was found to be 75 mM and 7.4 respectively. The suitable concentration of K-Pi buffer for suspending and storing mitochondria, however, was found to be 20 mM or lower. These requirements applied equally well for mitochondria from wheat (Triticum aestivum L.), barley (Hordeum vulgare L.), maize (Zea mays L.), and snap bean (Phaseolus vulgaris L.).     

Intact protein (antigen) absorption in fishes: mechanism and physiological significance

Although the literature indicates that there is no longer any reason to doubt that the fish gut possesses the capacity to absorb intact protein macromolecules, neither the exact mode of protein absorption nor the physiological significance of this phenomenon has been established. A review of the literature relating to evidence concerning the probable mechanism(s) of absorption and the means by which this process may confer physiological advantage is presented.