Localization of Sites in the Tail Domain of the Middle Molecular Mass Neurofilament Subunit Phosphorylated by a Neurofilament-Associated Kinase and by Casein Kinase I

Abstract: We have shown previously that a neurofilament (NF)-associated kinase (NFAK) extracted from chicken NF preparations phosphorylates selectively the middle molecular mass NF subunit (NF-M). Here we show that the major kinase activity in NFAK is indistinguishable from enzymes of the casein kinase I (CKI) family based on the following criteria: (1) inhibition of NFAK phosphorylation by the selective CKI inhibitor CKI-7, (2) the similarity in substrate specificity of NFAK and authentic CKI, (3) the correspondence of two-dimensional phosphopeptide maps of NF-M phosphorylated in vitro by NFAK with those generated by CKI under similar conditions, and (4) immunological cross-reactivity of NFAK with an antibody raised against CKI. We have also identified Ser⁵⁰², Ser⁵²⁸, and Ser⁵³⁶ as phosphorylation sites by NFAK/CKI in vitro, each of which is also phosphorylated in vivo. All three serines are found in peptides with CKI phosphorylation consensus sequences, and Ser⁵²⁸ and Ser⁵³⁶ and flanking amino acids are highly conserved in higher vertebrate NF-M sequences. Neither Ser⁵⁰² nor Ser⁵³⁶ has been identified previously as NF-M phosphorylation sites.     

The sequence and organization of the core histone H3 and H4 genes in the early branching amitochondriate protistTrichomonas vaginalis

Among the unicellular protists, several of which are parasitic, some of the most divergent eukaryotic species are found. The evolutionary distances between protists are so large that even slowly evolving proteins like histones are strongly divergent. In this study we isolated cDNA and genomic histone H3 and H4 clones fromTrichomonas vaginalis. Two histone H3 and three histone H4 genes were detected on three genomic clones with one complete H3 and two complete H4 sequences. H3 and H4 genes were divergently transcribed with very short intergenic regions of only 194 bp, which containedT. vaginalis-specific as well as histone-specific putative promoter elements. Southern blot analysis showed that there may be several more histone gene pairs. The two complete histone H4 genes were different on the nucleotide level but encoded the same amino acid sequence. Comparison of the amino acid sequences of theT. vaginalis H3 and H4 histones with sequences from animals, fungi, and plants as well as other protists revealed a significant divergence not only from the sequences in multicellular organisms but especially from the sequences in other protists likeEntamoeba histolytica, Trypanosoma cruzi, andLeishmania infantum.     

Vacuole Partitioning during Meiotic Division in Yeast

We have examined the partitioning of the yeast vacuole during meiotic division. In pulse-chase experiments, vacuoles labeled with the lumenal ade2 fluorophore or the membrane-specific dye FM 4-64 were not inherited by haploid spores. Instead, these fluorescent markers were excluded from spores and trapped between the spore cell walls and the ascus. Serial optical sections using a confocal microscope confirmed that spores did not inherit detectable amounts of fluorescently labeled vacuoles. Moreover, indirect immunofluorescence studies established that an endogenous vacuolar membrane protein, alkaline phosphatase, and a soluable vacuolar protease, carboxypeptidase Y, were also detected outside spores after meiotic division. Spores that did not inherit ade2- or FM 4-64-labeled vacuoles did generate an organelle that could be visualized by subsequent staining with vacuole-specific fluorophores. These data contrast with genetic evidence that a soluble vacuolar protease is inherited by spores. When the partitioning of both types of markers was examined in sporulating cultures, the vacuolar protease activity was inherited by spores while fluorescently labeled vacuoles were largely excluded from spores. Our results indicate that the majority of the diploid vacuole, both soluble contents and membrane-bound components, are excluded from spores formed during meiotic division.     

Reproductive and neonatal outcomes in captive bred baboons (Papio hamadryas)

Abstract: Baboons are widely used in biomedical research. Although it is widely held that Papio hamadryas breed well in captivity, each established colony has a different reproductive success often hypothesised to be due to husbandry practices. The National Baboon Colony in Australia is a unique colony that houses Papio hamadryas to mimic that structure seen in the wild. In this article; we have analysed their reproductive parameters and neonatal outcomes. The success of the colony husbandry practices was demonstrated by lack of maternal mortality, low foetal morbidity, and known maternal and paternal linage.     

Lactococcus lactis and stress

It is now generally recognized that cell growth conditions in nature are often suboptimal compared to controlled conditions provided in the laboratory. Natural stresses like starvation and acidity are generated by cell growth itself. Other stresses like temperature or osmotic shock, or oxygen, are imposed by the environment. It is now clear that defense mechanisms to withstand different stresses must be present in all organisms. The exploration of stress responses in lactic acid bacteria has just begun. Several stress response genes have been revealed through homologies with known genes in other organisms. While stress response genes appear to be highly conserved, however, their regulation may not be. Thus, search of the regulation of stress response in lactic acid bacteria may reveal new regulatory circuits. The first part of this report addresses the available information on stress response in Lactococcus lactis.Acid stress response may be particularly important in lactic acid bacteria, whose growth and transition to stationary phase is accompanied by the production of lactic acid, which results in acidification of the media, arrest of cell multiplication, and possible cell death. The second part of this report will focus on progress made in acid stress response, particularly in L. lactis and on factors which may affect its regulation. Acid tolerance is presently under study in L. lactis. Our results with strain MG1363 show that it survives a lethal challenge at pH 4.0 if adapted briefly (5 to 15 minutes) at a pH between 4.5 and 6.5. Adaptation requires protein synthesis, indicating that acid conditions induce expression of newly synthesized genes. These results show that L. lactis possesses an inducible response to acid stress in exponential phase.To identify possible regulatory genes involved in acid stress response, we determined low pH conditions in which MG1363 is unable to grow, and selected at 37°C for transposition insertional mutants which were able to survive. About thirty mutants resistant to low pH conditions were characterized. The interrupted genes were identified by sequence homology with known genes. One insertion interrupts ahrC, the putative regulator of arginine metabolism; possibly, increased arginine catabolism in the mutant produces metabolites which increase the pH. Several other mutations putatively map at some step in the pathway of (p)ppGpp synthesis. Our results suggest that the stringent response pathway, which is involved in starvation and stationary phase survival, may also be implicated in acid pH tolerance.     

The effect of variation in parameters of the male calling song of the katydid,Amblycorypha parvipennis (Orthoptera: Tettigoniidae), on female phonotaxis and phonoresponse

After being placed equidistant from playbacks of two computer-generated male calls, females of the katydidAmblycorypha parvipennis preferentially moved toward call phrases that were louder, longer, and leading (initiated without being overlapped by other call phrase endings). Because earlier tests with calling males had indicated that male mating success was related to these call parameters, we suggest that mating success is partly a result of female choice. Females also preferentially responded to the initial, rather than the latter, half of male phrases. Results of other tests suggested that females were responding to increased phonatome rate characteristic of the first half of the phrase. Males may compete to lead in order to prevent jamming of initial phrase information. Females also preferentially phonoresponded (ticked) in response to longer phrases. In earlier studies of male mating success and female phonotaxis using live males, male weight, sound level, and leading were intercorrelated; however, none of these parameters were correlated with phrase length. We suggest that females may respond to different call parameters under different environmental conditions.     

The nervous system of Tricladida. I. Neuroanatomy ofProcerodes littoralis (Maricola,Procerodidae): An immunocytochemical study

The organization of the nervous system ofProcerodes littoralis (Tricladida, Maricola, Procerodidae) was studied by immunocytochemistry, using antibodies to authentic flatworm neuropeptide F (NPF) (Moniezia expansa). Compared to earlier investigations of the neuroanatomy of tricladid flatworms, the pattern of NPF immunoreactivity inProcerodes littoralis reveals differences in the following respects: 1. Shape and structure of the brain. 2. Number and composition of longitudinal nerve cords. 3. Shape of branches of, and transverse connections between, main ventral nerve cords. 4. Composition of the pharyngeal nervous system. The rich innervation by NPF immunoreactive (IR) fibres and cells of the subepithelial muscle layer, the pharynx musculature and the musculature of the male copulatory apparatus indicates a neurotransmitter or neuromodulatory influence on muscular activity.     

A human glial hybrid cell line differentially expressing genes subserving oligodendrocyte and astrocyte phenotype

We have developed a series of immortal human-human hybrid cell lines that express phenotypic characteristics of primary oligodendrocytes, by fusing a 6-thioguanine–resistant mutant of the human rhabdomyosarcoma RD with adult human oligodendrocytes by a lectin-enhanced polyethylene glycol procedure. Hybrids were selected in an aminopterin-containing media. In contrast to the tumor parent cells, a hybrid clone M03.13 expressed surface immunoreactivity for galactosyl cerebroside and intracellular immunoreactivity for myelin basic protein (MBP), proteolipid protein (PLP), and glial fibrillary acidic protein (GFAP). Serum deprivation or chronic treatment with a protein kinase C activator 4-β-phorbol 12-myristate 13-acetate (PMA), but not dibutyl cyclic adenosine monophosphate induced coordinate up-regulation or de novo induction of oligodendrocyte phenotypic markers with concomitant down-regulation of GFAP expression. Consistent with immunohistochemical studies, northern blot analysis demonstrated that both MBP and PLP mRNA were up-regulated in MO3.13 cells by PMA treatment. M03.13 cells provide an immortalized clonal model system suitable for study of gene expression subserving oligodendrocyte and astrocyte phenotypes. © 1995 John Wiley & Sons, Inc.