Distinct mechanisms of splicing regulation in vivo by the Drosophila protein Sex-lethal

The protein Sex-lethal (SXL) controls pre-mRNA splicing of two genes involved in Drosophila sex determination: transformer (tra) and the Sxl gene itself. Previous in vitro results indicated that SXL antagonizes the general splicing factor U2AF⁶⁵ to regulate splicing of tra. In this report, we have used transgenic flies expressing chimeric proteins between SXL and the effector domain of U2AF⁶⁵ to study the mechanisms of splicing regulation by SXL in vivo. Conferring U2AF activity to SXL relieves its inhibitory activity on tra splicing but not on Sxl splicing. Therefore, antagonizing U2AF⁶⁵ can explain tra splicing regulation both in vitro and in vivo, but this mechanism cannot explain splicing regulation of Sxl pre-mRNA. These results are a direct proof that Sxl, the master regulatory gene in sex determination, has multiple and separable activities in the regulation of pre-mRNA splicing.     

Double-stranded-RNA-dependent protein kinase and TAR RNA-binding protein form homo- and heterodimers in vivo.

The yeast two-hybrid system and far-Western protein blot analysis were used to demonstrate dimerization of human double-stranded RNA (dsRNA)-dependent protein kinase (PKR) in vivo and in vitro. A catalytically inactive mutant of PKR with a single amino acid substitution (K296R) was found to dimerize in vivo, and a mutant with a deletion of the catalytic domain of PKR retained the ability to dimerize. In contrast, deletion of the two dsRNA-binding motifs in the N-terminal regulatory domain of PKR abolished dimerization. In vitro dimerization of the dsRNA-binding domain required the presence of dsRNA. These results suggest that the binding of dsRNA by PKR is necessary for dimerization. The mammalian dsRNA-binding protein TRBP, originally identified on the basis of its ability to bind the transactivation region (TAR) of human immunodeficiency virus RNA, also dimerized with itself and with PKR in the yeast assay. Taken together, these results suggest that complexes consisting of different combinations of dsRNA-binding proteins may exist in vivo. Such complexes could mediate differential effects on gene expression and control of cell growth.     

Influences on Food Selection of Women on High- and Low-Fat Diets

There is evidence that long-term maintenance of a low-fat diet reduces preference for high-fat foods. Sensory evaluation of the taste of fat, and preference for high and low-fat foods were studied in a group of former participants in a randomized dietary intervention trial aimed at lowering fat consumption. Intervention subjects consuming less than 25% of daily calories as fat and control subjects consuming more than 35% of daily calories as fat agreed to be in a "taste perception" study. In Study 1, subjects tasted 20 dairy solutions containing different levels of fat and sugar. Subjects rated the perceived intensity of fat taste, and of liking, for each of the solutions. In Study 2, subjects were asked to taste and rate 4 high-fat and 4 low-fat snack foods, and were then allowed to freely consume these foods in a preference test. Intervention and control subjects were similar in their sensory evaluation of the taste of fat in Study 1. In Study 2, intervention subjects reported a reduced hedonic rating of the taste of high-fat snack foods compared to control subjects, yet intervention subjects consumed the same amount of high-fat snack foods as control subjects. We conclude that a successful outcome in a dietary intervention may be due to social and cognitive factors, in addition to potential changes in hedonic response to fat.     

Depressive effect of isoflurane anesthesia on motor evoked potentials

The influence of the volatile anesthetic isoflurane (Forane) on motor evoked potentials was examined in rats. To record motor evoked potentials, single-shock electrical stimulation was delivered to the forelimb representation of the motor cortex. This resulted in elicitation of a compound muscle action potential from the contralateral extensor muscles. The effect of isoflurane was examined at various concentrations ranging from 0.2 to 1.5%. With increasing concentrations of isoflurane there was a progressive increase in onset latency of the compound muscle action potential and a decrease in peak-to-peak amplitude and duration. Latencies were significantly increased over baseline values for concentrations of isoflurane from 0.5 to 1.5% (P values were 0.001 to 0.007). For the amplitude and the duration, responses at 0.5 to 1.5% isoflurane were significantly lower than baseline (P values were 0.001 to 0.007). We conclude that isoflurane anesthesia significantly changes the muscle response evoked by motor cortex stimulation in experimental animals.     

Chlormethiazole, dizocilpine and haloperidol prevent the degeneration of serotonergic nerve terminals induced by administration of MDMA (‘Ecstasy’) to rats

An investigation has been made into the effect of 3,4-methylenedioxymethamphetamine (MDMA or ‘Ecstasy’) administration on the concentration of 5-hydroxytryptamine (5-HT), uptake of [³H]5-HT and [³H]paroxetine binding in rat cerebral cortex tissue. Four days after 2 injections of MDMA (20 mg/kg i.p., 6 hr apart) the concentrations of 5-HT and its metabolite 5-HIAA were reduced by 60%. The binding of [³H]paroxetine to the presynaptic 5-HT transporter was decreased and high affinity uptake of [³H]5-HT was reduced by a similar amount, indicating neurodegeneration of 5-HT terminals. Pretreatment with chlormethiazole (100 mg/kg i.p.), 10 min before each MDMA injection prevented the decrease in both [³H]parotextine binding and uptake of [³H]5-HT. The loss in 5-HT and 5-HIAA content was also attenuated. Pretreatment with dizocilpine (1 mg/kg i.p.) or haloperidol (2 mg/kg i.p.) also prevented the MDMA-induced loss of [³H]paroxetine binding and attenuated the loss of 5-HT and 5-HIAA content. All three compounds also decreased the degree of hyperthermia that follows MDMA administration, although previous studies suggest that the long term neurodegeneration is not associated with the acute hyperthermic response. These data support the findings of others that MDMA injection produces degeneration of 5-HT nerve terminals in the cortex, confirm that chlormethiazole, dizocilpine and haloperidol attenuate MDMA-induced neurotoxic loss of 5-HT and demonstrate for the first time that these compounds prevent the neurodegeneration of 5-HT nerve terminals that follows MDMA administration.     

Phase II trial of piroxantrone in metastatic breast cancer

Summary Thirty-two eligible patients with advanced metastatic breast cancer who had received no more than 1 prior chemotherapy regimen for metastatic disease (16 had received prior doxorubicin) were treated with piroxantrone at a dose of 120 mg/m² intravenously every 21 days. In the twenty-seven patients evaluable for response, two partial responses were seen. Toxicities observed were primarily hematologic with grade 3 or greater granulocytopenia occurring in 34% of the patients. One patient developed symptomatic congestive heart failure at a total cumulative dose of 960 mg/m². We conclude that piroxantrone given at this dose and schedule has minimal activity in patients with metastatic breast cancer.     

Effects of mifepristone on rabbit intraocular pressure in the presence and absence of dexamethasone

Topical dexamethasone was used to elevate rabbit intraocular pressure in order to study the interaction with a steroid antagonist, mifepristone. Dexamethasone did not cause a consistently significant increase in intraocular pressure. Animals treated with mifepristone followed by dexamethasone showed no apparent increase in intraocular pressure after dexamethasone, indeed mifepristone caused a lower intraocular pressure than seen in other groups whether in the presence or absence of dexamethasone. Reductions of intraocular pressure when mifepristone was given after 14 days of dexamethasone administration were not found. No conclusion can be reached regarding any dexamethasone antagonism by mifepristone, except that intraocular pressure tended to be lower even in the presence of dexamethasone.     

Marshall-Smith syndrome: the expanding phenotype.

We report a child of 3 years 9 months with the Marshall-Smith syndrome (MSS), characterised by the typical facial features, developmental delay, and advanced bone age. After the diagnosis was made at 5 months of age, careful observation for respiratory complications and failure to thrive was initiated. By 3 1/2 years of age, although our patient had no life threatening respiratory complications, investigation showed significant upper airway obstruction, which has been successfully treated. Aggressive treatment for failure to thrive has also allowed her to maintain a weight on the 50th centile. The purpose of this report is to suggest that early diagnosis and aggressive management may improve the ultimate prognosis with respect to the respiratory and feeding difficulties seen in this rare syndrome.