Ca2+ calmodulin-dependent protein kinase activity in the ascomycetes Neurospora crassa

DEAE-cellulose column chromatography of Neurospora crassa soluble mycelial extracts leads to the resolution of three major protein kinase activity peaks designated PKI, PKII, and PKIII.PKII activity is stimulated by Ca²⁺ and Neurospora or brain calmodulin. Maximal stimulation was observed at 2 µM-free Ca²⁺ and 1 µg/ml of the modulator. The stimulatory effect of the Ca²⁺-calmodulin complex was blocked by EGTA and by some calmodulin antagonists such as phenothiazine drugs or compound 48/80.PKII phosphorylates different proteins, among which histone II-A at a low concentration and CDPKS, the synthetic peptide specific for Ca²⁺-calmodulin dependent protein kinases, are the best substrates. Some phosphorylation can be detected in the absence of any exogenous acceptor. PKII activity assayed in the presence of histone II-A or in the absence of exogenous phosphate acceptor (autophosphorylation) co-elute in a DEAE-cellulose column at 0.28 M NaCl. As result of the autophosphorylation reaction of the purified enzyme a main phosphorylated component of 70 kDa was resolved by SDS-polyacrylamide gel electrophoresis. It is possible that this component is an active part of this enzyme.     

Segmental release of amino acid neurotransmitters from transcranial stimulation

The present study used microdialysis techniques in an intact rabbit model to measure the release of amino acids within the lumbar spinal cord in response to transcranial electrical stimulation. Dialysis samples from the extracellular space were obtained over a stimulation period of 90 minutes and were examined using high pressure liquid chromatography. Neuronal excitation was verified by recerding corticomotor evoked potentials (CMEPs) from the spinal cord. A significant increase in the release of glycine and taurine compared to sham animals was measured after 90 minutes of transcranial stimulation. Glutamate and aspartate release was not significantly elevated. GABA concentrations were consistently low. CMEP components repeatedly showed adequate activation of descending fiber pathways and segmental interneuron pools during dialysis sampling. Since glycine, and to a lesser extent taurine, have been shown to inhibit motor neuron activity and are closely associated with segmental interneuron pools, suprasegmental modulation of motor activity may be, in part, through these inhibitory amino acid neurotransmitters in the rabbit lumbar spinal cord.     

Candida albicans and three other Candida species contain an elongation factor structurally and functionally analogous to elongation factor 3.

A cell-free poly(U)-dependent translation elongation system from Candida albicans is ATP-dependent due to the presence of an elongation factor 3 (EF3)-like activity. Saccharomyces cerevisiae ribosomes added to a C. albicans postribosomal supernatant (PRS) supported poly(U)-dependent elongation, suggesting that the C. albicans lysate contained a soluble translation factor functionally analogous to the S. cerevisiae translation factor EF-3. The presence of EF-3 in C. albicans was confirmed by Western blotting using an antibody raised against S. cerevisiae EF-3. This antibody was also used to screen a selection of Candida species, all of which possessed EF-3 with molecular mass in the range of 110-130 kDa.     

Nonaspiration fine needle cytology. Application of a new technique to nodular thyroid disease

The basic principle underlying fine needle aspiration (FNA) is the aspiration of cellular material from target masses, often utilizing fairly high suction pressures. The procedure requires a needle and a syringe, advisedly held in a syringe holder, enabling single-handed suction to be exercised. Mastery of the technique is variable, with few operators acquiring consistent skill. A new technique, pioneered in France but essentially unpublicized, eliminates active aspiration, replacing it by the principle of capillary suction of fluid or semifluid material into a thin channel (a fine needle). This nonaspiration sampling method was tested in a consecutive series of 50 solid thyroid nodules. Simultaneously performed conventional FNAs served as controls. Cell samples were cytologically assessed as unsuitable, diagnostic/adequate or diagnostic/superior, without knowledge of the sampling method employed. Diagnostically superior specimens were obtained significantly more frequently by the nonaspiration technique in 36 benign lesions and 13 neoplasms. The method of nonaspiration fine needle cytology ("cytopuncture") is described and illustrated, and the implications for its use in other sites are discussed.     

Cytochemical evaluation of gamma-glutamyl-transpeptidase activity in cervical smears

The cervicovaginal smears of 43 patients attending an outpatient service for early cancer detection were cytochemically studied for the presence of gamma-glutamyl-transpeptidase (GGT) in epithelial cells. This was done in order to evaluate such an enzyme phenotype as a marker for cancer development. The results showed that 70% of the 38 patients with a cytologic diagnosis of "inflammatory" or preneoplastic/neoplastic conditions had GGT-positive cells in their smears. None of the five cytologically normal cases showed any epithelial cells with GGT activity. Although most of the GGT-positive cells were metaplastic, some morphologically normal, dysplastic or neoplastic cells also expressed the enzyme. The data suggest that cytochemically detectable transpeptidase activity appears whenever alterations of the normal epithelial microenvironment occurs, but is not necessarily linked to the carcinogenic process. Therefore, cytochemically GGT-positive cells should not be used as an indicator of neoplastic transformation of the cervical epithelium.     

Holozygosity for sex-linked genes in males of the termiteIncisitermes schwarzi

The termite Incisitermes schwarzi has multiple sex chromosomes that have arisen by repeated translocations between autosomes and previously existing sex chromosomes. Two sex-linked allozyme loci--Acp-1 and Est-3--are holozygous, not hemizygous, in males (the heterogametic sex). Both loci show less than 1% crossing-over between X and Y chromosomes, and alleles of both are in marked disequilibrium with respect to X vs Y linkage. The two loci assort independently in female meiosis, indicating that they lie on different sex chromosomes. But they are tightly linked in male meiosis because of nonrandom assortment of the multiple X and Y chromosomes in males of this species. The findings of holozygosity and strong linkage disequilibrium suggest that differential selection in the two sexes at or near these loci may be responsible for the establishment of the translocations in this species. The existence of active Y-linked alleles also suggests that the translocations may have occurred recently.     

Control of sugar transport in human fibroblasts independent of glucose metabolism or carrier-substrate interaction

Transport regulation by different metabolizable and nonmetabolizable sugars was studied in human fibroblasts. Sugars were classed as glucose-like (D-mannose, 3-0-methyl-D-glucose, thio-D-glucose, and D-allose) and starvation-like (D-galactose, D-fructose, L-glucose, D-xylose, 6-deoxy-D-glucose and 2-deoxy-D-glucose) based on their competence in curbing glucose starvation enhanced transport. No significant correlation existed between the ability of a sugar to curb hexose transport and the KI of that sugar in inhibiting hexose transport. Independence of the transport curb from glucose metabolism was observed since nonmetabolizable analogs of D-glucose when substituted for D-glucose in the culture medium effected glucose [i.e. 3-0-methyl-D-glucose (3-OMG)] and starvation-like (i.e. 6- and 2-deoxy-D-glucose) effects. The KI of inhibition pf 2-deoxy-D-glucose transport for 3-OMG was 8.5 mM, similar to those obtained for 6-deoxyglucose and 2-deoxyglucose on 2-deoxyglycose transport (7.5 and 3.5 mM, respectively) and on 3-0-methylglucose transport (3.5 and 2.5 mM, respectively). An equimolar mixture of D-glucose and 3-OMG (5.55 mM each) was more effective than 11.1 mM D-glucose or 3-OMG alone in curbing hexose transport or reversing hexose starvation induced increases in transport. The effect of 3-OMG may be independent of glucose metabolism but it is possible that 3-OMG structurally mimics a metabolite of glucose that may interact with intracellular regulators of carrier degradation and or expression.     

Blockade of a motor response by cholinergic stimulation of the toad midbrain tegmentum

Application of an acid solution to the dorsal skin of conscious toads having intact nervous system induces a scratching reflex and escape movements, as well as autonomic alterations (hypertension and tachycardia) that are part of the defense response. The motor components of this response are abolished or reduced by microinjection of 60, 30, 15 or 7.5 ng carbachol into the midbrain tegmentum. The cardiovascular components, however, continue to be present, although their amplitude is reduced. The depression of the motor response is statistically significant up to 15 minutes for the 60 ng dose, up to 10 minutes for the 15 and 30 ng doses, and only up to 5 minutes for the 7.5 ng dose. The data suggest that the midbrain tegmentum may modulate the reflex motor response triggered by a noxious stimulus and also participate in the organization of the escape movements. The importance of cholinergic agents in this modulation is discussed. The persistence of the cardiovascular component of the response shows the importance of this parameter as an indicator of alert situations.     

Potato Lectin: A Cell-Wall Glycoprotein

The activity and the amount of potato lectin were measured inpotato tuber slices (Solanum tuberosum cv. Huinkul) aeratedfor 48 h. Lectin was found in a soluble form, liberated to themedium and associated with insoluble structures. Polyacrylamidegel electrophoresis in denaturating conditions and immunologicaltechniques indicated that the lectins associated to cell wall,soluble or liberated to the medium, were identical. The cell-wallfraction was found to contain 65% of total lectin in the tuber.The possible role of potato lectin in tubers was discussed. (Received June 5, 1985; Accepted September 3, 1985)