bluuebunny/crossref_metadata_2025 huggingface dataset:
Stages of cell hair construction in Drosophila
Authors: Mitchell Herschel K. , Edens Jean , Petersen Nancy S.
Published: May 2005

The construction of cell hairs (trichomes) on the wings of Drosophila occurs in synchrony on 30,000 cells over a period of about 20 hr. Changes in both morphology and patterns of protein synthesis occur rapidly during this time period. In this report we describe the use of stressinduced (heat shock) abnormalities in morphogenesis to provide further details on the stepwise processes of differentiation within single wing cells. A cartoon summary of the overall process and a discussion of some possible mechanisms is included.
Expression of soybean lectin gene deletions in tobacco
Authors: Lindstrom Jon T. , Vodkin Lila O. , Harding Roy W. , Goeken Robert M.
Published: May 2005

A series of constructs containing the developmentally regulated soybean lectin gene (Le1) were used to transform tobacco plants in order to assess developmental and quantitative regulation conferred by flanking sequences. The largest of the lectin constructs contained approximately 3,00 base pairs (bp) of Le1 5′ flanking region and 1,500 bp of the 3′ flanking region. The smallest construct contained no 5′ flanking region and 194 bp of the 3′ flanking region. ELISA assays of lectin in individual tobacco seeds and Southern blot analyses confirmed that most constructs were inherited as unique insertion events. Maximal expression of Le1 required more than 338 bp of 5′ sequence, indicating that far upstream factors are involved in quantitative control of lectin expression. Lectin expression declined more than 80% between deletions with 1,700 versus 338 bp of 5′ flanking sequence. In contrast, developmental control of lectin expression was maintained by Le1 inserts with only 190 bp of 5′ sequence. The lectin promoter offers a potential means to target high levels of gene expression to the developing seeds of soybean or other dicotyledonous plants.
Butyrate induces expression of transfected human fetal and endogenous mouse embryonic globin genes in GM 979 erythroleukemia cells
Authors: Zhang Jun‐Wu , Raich Natacha , Enver Tariq , Anagnou Nicholas P. , Stamatoyannopoulos George
Published: May 2005

We have analyzed the expression of endogenous murine genes and of transfected human fetal Aγ globin gene in GM 979, a mouse erythroleukemia line which produces adult as well as embryonic globins. Optimal induction of the endogenous murine adult globin genes was obtained with DMSO or HMBA while the ϵy and βh1 embryonic genes were preferentially induced by butyrate. Similarly, the transferred human Aγ globin gene was preferentially induced by butyrate. These results as well as previous observations in vivo or in erythroid cell cultures suggest that butyrate preferentially induces the expression of fetal globin genes.
Direct gene transfer to protoplasts: Fate of the transferred genes
Authors: Saul Michael W. , Potrykus Ingo
Published: May 2005

Direct gene transfer to protoplasts is one of several methods developed for the production of transgenic plants. This method utilizes the efficient uptake of DNA from the surrounding medium by protoplasts (cell wall‐less plant cells). Where a suitable protoplast system exists large numbers of transformant clones can be efficiently produced and often regenerated to normal fertile plants. This review concentrates on the fate of the DNA which is taken up into the protoplasts. Particular emphasis is given to the factors which can influence the integration and form of the transferred DNA, the expression of transferred genes, and the inheritance in further generations of those genes. The information available suggests (1) that DNA is taken up by a large proportion of the cells in a transformation mixture, (2) that this DNA forms complexes sometimes involving carrier DNA, (3) that fewer cells actually take up DNA into the nucleus, and (4) that the complex may be rearranged and/or amplified and then integrated into the genome. If the DNA is arranged in such a way that a gene can be expressed it does so in a normal manner and is stably inherited both mitotically and meiotically.
Cell cycle‐regulated gene expression in transgenic plant cells
Authors: Kawata Takefumi , Nakayama Takuya , Ohtsubo Norihiro , Tabata Tetsuya , Iwabuchi Masaki
Published: May 2005

A majority of histone genes are expressed in the S phase during the cell cycle. Using the gene expression system of transformed sunflower cells into which wheat histone H3 gene was introduced by the Ti‐plasmid gene transfer technique, we determined three cis‐acting control sequences (hexameric, octameric, and nonameric motifs) which seemed to confer the S‐phase‐specific transcription of wheat histone genes. Furthermore, as candidates for regulatory transcription factors, three nuclear DNA‐binding proteins HBP‐la, HBP‐lb, and HBP‐2 that interact with the hexameric and nonameric motifs were identified. The structural analysis of the cDNA of HBP‐la revealed that a nuclear protein has the leucine‐zipper structure and a DNA‐binding motif. The hexameric motif in the H3 gene was also seen in cauliflower mosaic virus 35S (CaMV 35S) promoter and shown to function as a regulatory element of this promoter. The wheat HBP‐1b can interact with the hexameric motif of the CaMV 35S promoter. Much attention has been paid to the significance of the hexameric sequences within the H3 and CaMV 35S promoters and the DNA‐binding proteins HBP‐la and HBP‐lb.
Analysis of coat‐color patterns in aggregation chimeras between BALB/cA and C3H/HeN mice with special reference to migratory patterns and clone number of epidermal melanoblasts
Authors: Tachi Chikashi , Yokoyama Minesuke , Kojima Hiroko
Published: May 2005

Chimeras provide unique opportunities to study interactions between the phenotypically similar but genotypically allogeneic cell populations during embryogenesis in vivo. From the quantitative analysis of coat‐color patterns in C3H/HeN↔BALB/cA chimeras, a model was proposed stating that the aggregability of the C3H/ HeN‐derived melanoblasts in the chimeras was inversely related to the ratio between the mean free path of the epidermal melanoblasts in the normal C3H/He N mouse and that in the chimeras. As a corollary, the possibility was suggested that during the migration of melanoblasts, mechanisms identical with or similar to contact inhibition of movement might operate after collision between the isogeneic, but not between the allogeneic melanoblasts. With regard to the number of melanoblast clones in the trunk region of the mouse, the present series of analyses yielded the value of 24–28 arranged unilaterally; the value closely approximated the number of the somites in that region and provided further support for the proposition made earlier by Tachi [Dev Genet 9:121–154, 1988; “Development of Preimplantation Embryos and Their Environment.” New York: Alan R. Liss, Inc., 1989, pp 263–274].
Heat shock causes the collapse of the intermediate filament cytoskeleton in Drosophila embryos
Authors: Walter Marika F. , Biessmann Harald , Petersen Nancy S.
Published: May 2005

Heat shock has a dramatic effect on the organization of the cytoplasm, causing the intermediate filament cytoskeleton to aggregate of the nucleus. This has previously been shown in cultured Drosophila and mammalian cells. In this paper we analyze the heat lability of the intermediate filament cytoskeleton in early Drosophila embryos by indirect immunofluorescence. At all stages of embryogenesis tested, the intermediatefilamentcytoskeleton, which is maternally provided, is severely disturbed by 30 min heat shock at 37°C. After the nuclei have migrated to the subcortical cytoplasm, it collapses around them. Nuclei in all heat‐shocked embryos are considerably enlarged and become displaced. Embryos before cellular blastoderm stage, in which heat shock protein synthesis is not inducible, are irreversibly arrested in development by heat shock. Embryos at or after cellular blastoderm, which do synthesize heat shock proteins in response to stress, are also immediately arrested in development but continue development when returned to 25°C. We discuss the possibility that cytoplasmic events such as the intermediate filament cytoskeleton rearrangement may be involved in heat shock‐mediated phenocopy induction.
Cell type specific expression of a CaMV 35S‐GUS gene in transgenic soybean plants
Authors: Yang Ning‐Sun , Christou Paul
Published: May 2005

A number of independently derived transgenic soybean plants expressing a chimeric β‐glucuronidase (GUS) gene under the control of the 355 CaMV promoter and a nopaline synthase polyadenylation signal were recovered using direct DNA transfer via electric discharge particle acceleration. Expression of GUS in R, plants was localized using thin tissue sections. Many tissue types expressed GUS at various levels. Pericycle cells in root, parenchyma cells in xylem, and phloem tissues of stem and leaf had high levels of enzyme activity. Procambium, phloem, and cortex cells in root, vascular cambium cells in stem, and the majority of cortex cells in leaf midrib, expressed low or no GUS activity. Intermediate levels of GUS activity were detected in leaf mesophyll cells, certain ground tissue cells in stem and leaf midrib, and in trichome and epidermal guard cells. Thus, we conclude that the 35S CaMV promoter is cell‐type specific and is developmentally regulated in soybean.
Arabidopsis: Sensitivity of growth to high temperature
Authors: Binelli Giorgio , Mascarenhas Joseph P.
Published: May 2005

Arabidopsis thaliana seedlings as measured by an electrolyte leakage assay, have been found to be extremely sensitive to high temperature stress as compared to a high temperature tolerant variety (Tracy) of soybean. Over 50% ion leakage occurred in Arabidopsis leaves during a 15‐minute exposure to 50°C, indicating a heat killing time of less than 15 minutes. In contrast, the heat killing time for soybean at 50°C was over five times longer. When soybean or Arabidopsis seedlings in culture plates were exposed to 37°C for 2 hours and then returned to 23°C, they suffered no apparent short‐term or long‐term damage. Soybean seedlings given a 42°C, treatment for 2 hours also showed no damage. Arabidopsis seedlings after a 42°C treatment for 2 hours showed no apparent immediate damage, but 48 hours after return to 23°C severe damage symptoms were visible and after 96 hours all the seedlings were dead. Both soybean and Arabidopsis seedlings synthesize heat shock proteins (hsps) when exposed to 42°C for 2 hours. The hsps synthesized are of similar molecular weights, although the relative abundances of the different size classes are very different in the two plants. Even though hsps are produced in Arabidopsis seedlings after a 2 hour exposure to 42°C their presence is not sufficient for the seedlings to recover from the effects of rhe heat shock when returned to 23°C. Our results show that Arabidopsis has a heat sensitive genotype. This along with its other characteristics should make it a good model system in which to assay in transgenic plants, the functions of homologous and heterologous genes that might be candidates for determining heat tolerance in plants.
Characterization of IMP‐E3, A gene active during imaginal disc morphogenesis in Drosophila melanogaster
Authors: Moore John T. , Fristrom Dianne , Hammonds Ann S. , Fristrom James W.
Published: May 2005

The steroid hormone 20‐hydroxyecdysone (20‐HE) induces imcainol discs to form adult appendages in Drosophila. We have isolated a set of six ecdysone‐responsive genes that apparently encode disc cell‐surface or secreted proteins. Transcripts from one of these genes, IMP‐E3, accumulate rapidly within 1–2 h in response to hormone. Developmentally,IMP‐E3 transcripts reach maximum levels during the first stages of metamorphosis (white prepupae, WPP) and are primarily limited to imaginal tissues. Transcripts are also present during embryogenesis (0–3 h and 12–18 h). Two different‐sized transcripts (1.2 and 1.4 kb) result from differential polyadenylation, with the larger transcript predominating in WPP. The conceptual IMP‐E3 protein contains a signal peptide, an RGD sequence, and a potential glycosylphosphatidylinositol anchor. We speculate that the protein provides a transient cue important for imaginal disc morphogenesis.
Molecular and developmental analyses of the protein encoded by the Drosophila gene Ectodermal
Authors: Raha Debasish , Nguyen Quan Dong , Garen Alan
Published: May 2005

The Drosophila gene ectodermal (ect, located at 67D8‐10 on chromosome 3) is expressed for a short period at mid‐embryogenesis in all ectodermally derived tissues except the nervous system. During this stage the tissues involved form tubular structures by a process of invagination followed by cell fusion. Here we report the sequence of the ect protein as deduced from the longest ORF (280 codons) of an ect cDNA. The principal molecular features of the ect protein are: (1) a consensus leader sequence for targeting to the rough ER; (2) a central domain containing a remarkably high density of acidic residues arranged in large clusters separated by smaller clusters of hydrophobic residues; (3) a consensus nuclear‐targeting sequence near the C‐terminus; (4) a single tyrosine residue located at a potential tyrosine‐sulfation site. The antibody staining pattern of the ect protein corresponds to the in situ hybridization pattern of the transcript. A possible role for the ect protein in the complex process of fubular formation that occurs in embryonic ectodermal tissues is discussed.
In situ hybridization studies on murine catalase mRNA expression during embryonic development
Authors: Reimer D. L. , Singh S. M.
Published: May 2005

In situ hybridization using nucleic acid probes was used to detect cell‐ and tissue‐specific transcript(s) of embryonic genes during development and differentiation. This highly sensitive technique has the potential to provide valuable information on the regulation of low‐abundance housekeeping genes during development. We have determined the experimental conditions required to detect the catalase message in adult mouse liver. Catalase effects the breakdown of H2O2 to O2 and H2O and offers protection against the toxic effects of oxygen radicals. We used a cloned 550 bp BamHI‐Pstl fragment from a mouse catalase cDNA (pMCT‐1) to generate 35S‐labeled sense and antisense riboprobes. The experimental conditions used were sensitive enough to quantitate the abundance of silver grains generated by the antisense riboprobe on the adult liver, a tissue known to be positive for this message. The hybridization protocol was applied to serial sections of 13‐ and 18‐day‐old mouse embryos. The results suggest that the catalase expression in the liver and brain begins with somite formation and increases with development and differentiation. On the other hand, this message appears to be absent in mesenchyme, particularly in day 13 embryos. The message in positive tissues appears evenly distributed throughout the cell. The observed expression of the catalase message in the adult liver is approximately six times that in the embryonic liver. It is compatible with the enzyme activity results and emphasizes the sensitivity of the in situ hybridization method (over northern blot, etc.) used in this study.
Cell‐cell adhesion and morphogenesis in Dictyostelium discoideum
Authors: Siu Chi‐Hung , Kamboj Rajender K.
Published: May 2005

During development of Dictyostelium discoideum, cells acquire EDTA‐resistant cell‐cell adhesion at the aggregation stage. The EDTA‐resistant cell binding activity is associated with a cell surface glycoprotein of Mr 80,000 (gp80), which mediates cell‐cell binding via ho‐mophilic interaction. Analysis of the structure of gp80 deduced from cDNA sequence reveals the presence of three internally homologous segments in the NH2‐terminal domain, which also contains regions with homology to the neural cell adhesion molecule. Secondary structure predictions show an abundance of β‐structures and very few α‐helices. This is confirmed by circular dichroism measurements. It is likely that the homologous segments are organized into globular structures, extended from the cell surface by a Pro‐rich stalk domain. The cell binding activity of gp80 resides within the first globular repeat of the NH2‐terminal domain and has been mapped to a 51 amino acid region between Val123 and Leu 173. Synthetic oligopeptides corresponding to sequences within this region have been prepared and assayed for their ability to bind to cell surface gp80. Results lead to identification of the homophilic binding site to an octapeptide sequence within this region. Synthetic peptides containing this octapeptide sequence and univalent antibodies directed against this site block the formation of organized cell streams during aggregation. Although cell aggregates are eventually formed, most fail to undergo further development to give rise to slugs and fruiting bodies, indicating that cell‐cell adhesion involving gp80 is an important step in normal morphogenesis.
Regulation of the anterior‐like cell state by Ammonia in Dictyostelium discoideum
Authors: Nathan Feit Ira , Bonner John Tyler , Suthers Hannah B.
Published: May 2005

Ammonia appears to be an important regulatory signal for several aspects of the Dictyostelium life cycle. The postulated role of ammonia in the determination of the prespore pathway in cells of the slug stage has led us to examine the effect of ammonia on the prestalk/prespore ratio of migrating slugs. In the presence of 10−3 M ammonium chloride, the volume of the prestalk region decreases by 40.8%. The kinetics of the process make it unlikely that this is due to a shift in the differentiation pathway. A test of the hypothesis that the decrease in volume of the prestalk region is due to the conversion of prestalk cells to anterior‐like cells shows that the percent of anterior‐like cells in the posterior region increases by the amount predicted by the hypothesis. This suggests that ammonia may be the molecular signal, produced by the tip, that prevents anterior‐like cells from chemotactically migrating to the tip and thereby becoming anterior cells. The effect of enzymatic removal of ammonia from vitally stained migrating slugs is the appearance of a series of dark stripes beginning at the posterior end and progressing forward. We interpret this as a result of progressive removal of anterior‐like cells from tip dominance and essentially as the formation of new potential tips. Indeed, in a few cases one or even two of the stripes separate from the posterior of the cell mass and form small fruiting bodies. We consider the phenomenon of stripe formation further evidence that the tip acts on anterior‐like cells through ammonia.
Position‐Dependent regulation of the prestalk‐prespore pattern in Dictyostelium slugs
Authors: Pogge‐von Strandmann Ralph , Kay Robert R.
Published: May 2005

In classical transplantation experiments, we have attempted to distinguish between cell sorting and position‐dependent mechanisms of pattern formation during the regulation of amputated prespore regions of Dictyostelium slugs. Host and transplanted tissue were distinguished by prior labelling with 14C and 3H, and their distributions in the manipulated slugs were determined by double‐label scintillation counting of serial sections of the slugs. First, we showed that the regenerated prestalk region is formed predominantly from the anterior of a prespore isolate. This disproves extreme cell sorting models, which hold that the new prestalk cells arise at random in the prespore mass, before sorting to the prestalk zone. Second, we showed that a proportion of posterior cells, which are not normally recruited into the new prestalk region, can be recruited into it if they are transplanted into the anterior of the prespore isolate. Recruitment of these cells occurs only during the first hour of regulation. We believe that these experiments prove a position‐dependent mechanism of pattern formation during slug regulation. This in turn implies the existence of localized inductive signals in the regulating fragments. Although cell sorting occurs efficiently when cells are misplaced in the slug, it does not appear to play a major role in reforming the prestalk/prespore pattern.
Processing of neutral N‐linked oligosaccharides during early Dictyostelium discoideum development
Authors: Hobbs Lisa J. , Das O. Prem , Henderson Ellen J.
Published: May 2005

We have used metabolic radiolabeling with oligosaccharide precursors, coupled with subcellular fractionation, to examine the distribution of several classes of asparagine‐linked oligosaccharides during early development. In Dictyostelium, we have observed endoglycosidase H (endo H)‐sensitive structures with sizes corresponding to 10 (Hex10) and 11 (Hex11) hexose residues on the chitobiose core. Only Hex11 was detected as the major structure on fucosylated endo H‐resistant species. All Hex11 species cofractionated with plasma membrane and secreted glycoproteins, whereas Hex10 appeared to be confined to intracellular membrane and soluble glycoproteins. Sulfated species correlated with lysosomal and secreted fractions, and glucose residues were markedly depressed in Hex11 of secreted glycoproteins. Outer branch structural studies have revealed several components of the endo H‐sensitive species. Usingα‐mannosidase and β‐hexosaminidase as diagnostic tools, species elucidated thus far are: a structure with 10 mannoses, a structure with nine mannoses and an intersecting N‐acetylglucosamine, structures with three glucoses and seven or eight mannoses and several larger species with multiple blocks to digestion.
Formation of the Dictyostelium spore coat
Authors: West Christopher M. , Erdos Gregory W.
Published: May 2005

The spore coat forms as a rigid extracellular wall around each spore cell during culmination. Coats purified from germinated spores contain multiple protein species and an approximately equal mass of polysaccharide, consisting mostly of cellulose and a galactose/N‐acetyl‐galactosamine polysaccharide (GPS). All but the cellulose are prepackaged during prespore cell differentiation in a regulated secretory compartment, the prespore vesicle. The morphology of this compartment resembles an anastomosing, tubular network rather than a spherical vesicle. The molecules of the prespore vesicle are not uniformly mixed but are segregated into partially overlapping domains. Although lysosomal enzymes have been found in the prespore vesicle, this compartment does not function as a lysosome because it is not acidic, and a common antigen associated with acid hydrolases is found in another, acidic vesicle population. All the prespore vesicle profiles disappear at the time of appearance of their contents outside of the cell; this constitutes an early stage in spore coat formation, which can be detected both by microscopy and flow cytometry. As an electron‐dense layer, the future outer layer of the coat, condenses, cellulose can be found and is located immediately beneath this outer layer Certain proteins and the GPS become associated with either the outer or inner layers surrounding this middle cellulose layer. Assembly of the inner and outer layers occurs in part from a pool of glycoproteins that is shared between spores, and unincorporated molecules loosely reside in the interspore matrix, a location from which they can be easily washed away. When the glycosylation of several major protein species is disrupted by mutation, the coat is assembled, but differences are found in its porosity and the extractibility of certain proteins. In addition, the retention or loss of proteolytic fragments in the mutants indicates regions of spore coat proteins that are required for association with the coat. Comparative examination of the macrocyst demonstrates that patterns of molecular distributions are not conserved between the macrocyst and spore coats. Thus spore coat assembly is characterized by highly specific intermolecular interactions, leading to saturable associations of individual glycoproteins with specific layers and the exclusion of excess copies to the interspore space.
Multiple genes for cell surface cAMP receptors in Dictyostelium discoideum
Authors: Saxe Charles L. , Johnson Ronald , Devreotes Peter N. , Kimmel Alan R.
Published: May 2005

We have cloned and characterized three genes (CAR1, CAR2, CAR3) encoding potential cell surface, cyclic adenosine 3′:5′ monophosphate (AMP) receptors from Dictyostelium discoideum. The three proteins are predicted to be substantially similar in amino acid sequence throughout most of their transmembrane (TM) and loop domains but are distinctly different in their carboxyl terminal segments. In addition, all three genes possess an intron which interrupts an equivalent codon of TM3.

CAR1 is expressed early in development when the cAMP relay system is being established. As development proceeds multiple size forms of CAR1 RNA are detected which apparently result from differences in their 5′‐untranslated regions. Late in development levels of CAR1 RNA decrease. In contrast, CAR2 encodes a single sized RNA which is expressed only during postaggregative development. CAR3 expression is ∼10% of CAR1 during early development, is maximal during tight aggregate formation but declines thereafter. Only one size class of CAR3 mRNA is detected throughout development.

Because RNA for each of the three genes is present in postaggregative cells, it was of interest to determine the cell type distribution of each RNA. Gene‐specific probes were hybridized to RNAs isolated from cells of Percoll gradient‐enriched prespore and prestalk fractions and relative levels of hybridization compared. CAR1 and CAR3 show approximately the same pattern of accumulation; a 3–4 fold enrichment in prestalk cells. CAR2, however, is highly enriched in prestalk cells, more than 10 fold relative to prespore cells.
Signal transduction pathways involved in the expression of the uridine diphosphoglucose pyrophosphorylase gene of Dictyostelium discoideum
Authors: Haribabu Bodduluri , Pavlovic Jovan , Bodduluri Sobha R. , Doody Jacqueline F. , Ortiz Benjamin D. , Mullings Suzan , Moon Byoung , Dottin Robert P.
Published: May 2005

The uridine diphosphoglucose pyrophosphorylase (UDPGP1) gene of Dictyostelium discoideum is an excellent marker to study the pathways that control the expression of genes during development. We have previously shown that the UDPGP1 gene is regulated by exogenous cAMP acting on cell‐surface cAMP receptors. Various steps in the signal transduction pathway between receptor stimulation and the induction of the gene can now be studied. Induction does not require the synthesis of intracellular cAMP, but does require new protein synthesis. By deletion and transformation with altered genes, two cis‐acting sequences that are required for UDPGP1 expression have been identified. A GC‐rich palindromic sequence located between −410 and −374 is essential for induction of the gene by extracellular cAMP, but not for its basal expression. A sequence element located between −374 and −337 is required for any basal expression of this gene. When the polarity of the palindromic sequence was reversed such that it resembled the H2K enhancer element, the gene could still be induced by exogenous cAMP. Two DNA binding activities were detected in gel mobility shift assays using a fragment containing both of the regulatory sequence elements of UDPGP1 gene. Transformation with a vector that resulted in the synthesis of anti‐sense UDPGP1 RNA led to almost total elimination of the enzyme antigen and no detectable enzyme activity. However, these transformants developed normally, indicating that either UDPGP is not required for development or residual synthesis of UDPGP may be sufficient for normal development.
Evidence for the presence of a growth factor in Dictyostelium discoideum
Authors: Whitbread John A. , Sims Matthew , Katz Eugene R.
Published: May 2005

Polypeptide hormones, recognized for their ability to regulate cell growth and differentiation, have been classified as growth factors. These growth factors have been extensively described in higher eukaryotic organisms and cell lines [Hedin and Westermark, Cell 37:9–20, 1984]. Here we report the identification and partial characterization of a putative growth factor present in vegetative amoebae of the cellular slime mold Dictyostelium discoideum. A mutant was selected and found to be temperature sensitive due to the absence of an extracellular protein suggestive of a growth factor. The putative growth factor (DGF) is a protein resistant to both heat and strong detergent treatment but sensitive to reducing agents. The physiological significance of DGF is as yet unknown. DGF is of interest both in relation to understanding the events which control cell proliferation in Dictyostelium and in its relationship to other known growth factors.
Sequence elements that affect mRNA translational activity in developing Dictyostelium cells
Authors: Steel Laura F. , Jacobson Allan
Published: May 2005

Post‐transcriptional controls, including changes in both mRNA translational activity and stability, play an important role in the regulation of ribosomal protein gene expression in developing Dictyostelium discoideum cells. Previously we have shown that the mechanisms which regulate the translational activity of the r‐protein mRNAs operate at the level of translational initiation and do not involve changes in polyadenylation or capping. By analysing the translational behavior of chimeric and mutant mRNAs in transformed cells, we have now been able to localize the determinants of translational activity of one of the r‐protein mRNAs to the 5′‐untranslated region. Although this and other r‐protein mRNAs differ strikingly from the Dictyostelium consensus in the region of the initiator AUG codon, we find that improving the match to that consensus does not increase the translational activity of the message in developing cells. Current experiments are designed to determine whether translational regulation is mediated by strong interactions with specific inhibitors or by weak interactions with translational initiation factors.
Developmental protein synthesis is required for the transcription of Dictyostelium prespore genes
Authors: Benedict M. A. , Desilver D. A. , Pelletier D. E. , Pentz W. H. , Ratner D. I.
Published: May 2005

It has been established previ‐ ously that the maintenance of expression of pre‐spore‐specific genes of Dictyostelium discoideum is prevented by the translational inhibitor cyclohex‐ imide. The drug had no effect upon the level of transcripts of the other genes examined, prestalk‐specific or cell type‐nonspecific. However, the interpretation of this result is open to question, because of possible nonspecific effects of cyclo‐heximide. We have now characterized the cellular specificity and temporal profiles of mRNA accumu‐ lation of additional Dictyostelium cDNA clones, and have examined other inhibitors of in vivo protein synthesis. Four structurally and mechanistically distinct translational inhibitors each prevented the reaccumulation of prespore transcripts in cyclic AMP‐primed, disaggregated amoebae. These results establish the importance of developmental protein synthesis in the accumulation of prespore gene transcripts. Nuclear run‐on transcription assays were used to learn whether protein synthesis is required primarily for mRNA synthesis or transcript stability. A transcriptional time course first demonstrated that the abundance of these cell‐specific transcripts during development mirrors their rates of synthesis. Significantly, the protein synthesis requirement of the prespore genes examined also occurs at the level of mRNA transcription, implying the existence of one or more developmentally regulated transcriptional activators.
Dictyostelium discoideum gene family contains a long internal amino acid repeat
Authors: Ennis Herbert L. , Giorda Roberto , Ohmachi Tetsuo , Shaw David R. , Blume John E.
Published: May 2005

Two different cDNA clones denoted pT0270‐6 and pT0270‐11 represent two mRNAs that are developmentally regulated during spore germination in Dictyosteliurn discoideum. The respective mRNAs are found only during early germination and are not present in other stages of growth or multicellular development. Four different genomic clones that hybridize to sequences that are common to both of the 270 cDNA clones were isolated from Dictyostelium libraries and se‐ quenced. Two are the genes for the two cDNAs, and the other two represent genes that do not seem to be transcribed. All four genomic se‐ quences possess a very unusual internal feature in the deduced protein sequences composed of a monotonous repeat of the tetrapeptide threonine‐ glutamic acid‐threonine‐proline. The other portions of the proteins have no homology among them‐ selves. The deduced protein corresponding to the 270‐6 gene is very similar to avocado (Persea arnericana) cellulase. Since cellulose in the spore wall has to be digested during spore germination this suggests that this protein may function as an endo‐( 1,4)‐beta‐D‐glucanase during germination.
Kinase activity and genetic characterization of a growth related antigen of Drosophila
Authors: Bedian Vahe , Jungklaus Christine E. , Cardoza Lisa , Kalm Laurence Von
Published: May 2005

The Drosophila developmental antigen recognized by the monoclonal antibody F7D6 is expressed in dividing embryonic and imaginal cells but is lost from all differentiating fissues except electrogenic cells of the nervous system and spontaneously contracting muscles. The 63 kDa antigen is associated with the inner surface of plasma membranes and is expressed in several classes of fumorous mutants of Drosophila. The monoclonal antibody was used for immunoprecip‐itating the antigen for biochemical characterization and for screening expression vector cDNA libraries. Here we report that this oncodevelopmental antigen is a phosphoprotein and a serine‐threonine specific protein kinase. A 1.6 kb cDNA isolated by immunological screening of an ovarian library hybridized to a single band on polytene chromosomes, localizing the gene to 72F on the left arm of the third chromosome. Immunofluorescence assays of deficiency stocks in the region confirmed the location of the gene and identity of the cDNA clone, and mapped the gene between the left breakpoints of Df(3L) st1100.62 and Df(3L) sti7, i.e., between 72F3–7 and 73A1–2. The biochemical and genetic properties indicate that this is a novel growth‐related kinase of Drosophila.
Despite expression in embryonic visceral mesoderm, H2.0 is not essential for Drosophila visceral muscle morphogenesis
Authors: Barad Mark , Erlebacher Adrian , McGinnis William
Published: May 2005

H2.0, a homeobox gene identified by homology to the Sex combs reduced homeobox of Drosophila, is expressed in all the cellular precursors of the visceral musculature. By analogy to the essential function of most other known homeobox genes in determining the fate of cells where they are expressed, we hypothesized that mutation of H2.0 would disrupt gut muscle development. In this paper, we show that a small deletion, which eliminates H2.0, has no detectable effect on normal gut morphogenesis, visceral muscle actin organization, or larval peristalsis.
Regulatory elements near the Drosophila hsp 22 gene required for ecdysterone and heat shock induction
Authors: Rudolph Karen , Morganelli Christine , Berger Edward M.
Published: May 2005

A transient expression assay was used to localize cis‐acting DNA regulatory elements near the Drosophila heat shock protein (hsp) 22 gene, that are involved in heat shock expression and in ecdysterone‐induced expression. The results identify a region between positions −320 and −232 that is essential for ecdysterone control, but not for heat‐induced expression, and a sequence between −199 and −56, which, when deleted, leads to the loss of heat shock induction. To investigate the function of these DNA sequences, transfection‐competition experiments were carried out. The evidence suggests that the DNA regulatory sequences identified by transient expression studies contain binding sites for transacting transcription factors.
Characterization and complementation analysis of sporogenous mutants of Dictyostelium discoideum
Authors: Sadiq May F. , Town Christopher D.
Published: May 2005

Sporogenous mutants of the cellular slime mold Dictyostelium discoideum are defined as mutants which are able to undergo terminal differentiation into spores in monolayer cultures in the presence of millimolar amounts of exogenous cyclic AMP. We describe the morphological development and cellular differentiation of a collection of 12 independently isolated sporogenous mutants of strain V12 M2. All mutants develop more rapidly than do wild‐type at an air‐water interface, display aberrant morphogenesis, and show overt spore and stalk differentiation as soon as 4 hr after starvation. All mutants differentiate in submerged monolayer culture in the presence of cAMP into variable proportions of spores and stalk cells. A number of the mutants also form both stalk cells and spores in submerged culture in the absence of exogenous cAMP. The spores formed by many of the mutants have a greatly reduced viability. Using parasexual genetics, we have found that two of the 12 mutants analyzed are dominant to wild‐type and the remaining ten fall into a minimum of four complementation groups, the overall analysis thus yielding a minimum of four and a maximum of seven complementation groups. Intracellular cAMP levels in vegetative cells are significantly elevated in the two dominant mutants but are similar to wild type in all the other mutants.
Effect of prenatal ethanol exposure on postnatal neural gene expression in the rat
Authors: Naus Christian C. G. , Bechberger John F.
Published: May 2005

To examine the effects of ethanol exposure on neural development, pregnant rats were fed a liquid diet in which 37.5% of the total caloric content was ethanol‐derived. The developmental appearance of the messenger RNAs coding for preprosomatostatin, glial fibrillary acidic protein, and proteolipid protein was examined by Northern blotting of total cellular RNA obtained from forebrain and hindbrain at various times after birth. In general, there was a delay in the developmental pattern of appearance of these mRNAs which was most noticeable at the early postnatal times. These results suggest that the previously reported delay in neural maturation is reflected at the level of the gene expression.
Several testis‐expressed genes in the mouse t‐complex have expression differences between wild‐type and t‐mutant mice
Authors: Ha Haesook , Howard C. Alan , Yeom Young Il , Abe Kuniya , Uehara Hiroshi , Artzt Karen , Bennett Dorothea
Published: May 2005

The t‐complex of the mouse occupies the proximal half of chromosome 17 and cantains genes which have profound effects on spermatogenesis. Mutations of several loci in the t‐complex appear to interact to cause male sterility or transmission ratio distortion (TRD).

By cDNA screening or chromosomal walking we have identified seven genes, which are expressed in the germ cells of testis and map to various regions of the t‐complex. These genes were named t‐complex testis‐expressed (Tctex) genes. An analysis of their expression patterns in testes from +/+, +/t, and t/t mice was done by in situ hybridization and by northern blotting. Six genes begin to be expressed at the pachytene stage: Three of them are more abundant at pachytene stage, while three others are more abundant at postmeiotic stages. One gene is expressed at all the stages of spermatogenesis. Interestingly, four Tctex genes show differences in the amount of transcript between wild‐type and t‐mutant testes. The chromosomal location and expression pattern imply that Tctex genes might be candidate genes for sterility or TRD.
Elevated paraquat resistance can be used as a bioassay for longevity in a genetically based long‐lived strain of Drosophila
Authors: Arking Robert , Buck Steven , Berrios Angela , Dwyer Susan , Baker George T.
Published: May 2005

A long‐lived (L) strain of Drosophila melanogaster, derived from a normal‐lived (R) strain by artificial selection, has a significantly different adult longevity. Previous work has shown that (1) the two strains age in the same manner, (2) the major genes responsible for much of the L strain's extended longevity are located on the 3rd chromosome, and (3) the extended longevity phenotype is significantly modulated by the larval environment. In this report, we investigate the resistance of the L and R strains to the lethal effects of dietary paraquat. We show that, within the limitations of our described chromosomal and environmental manipulations, the extended longevity phenotype always accompanies the phenotype of elevated paraquat resistance. In addition, reversed selection applied to the L strain results in the simultaneous decrease of both life span and paraquat resistance. Thus, the presence or absence of the latter phenotype may be used as a bioassay for the presence or absence of the extended longevity phenotype, without any necessary implication of causality. Use of this bioassay should greatly speed up the genetic analysis of this system by allowing us to identify long‐lived animals at a young age. Finally, we show that the age‐related loss of elevated paraquat resistance in both strains precedes all the other age‐related functional decrements which we have previously noted in this system.
Morphological abnormalities, neonatal mortality, and reproductive abnormalities in mice transgenic for diphtheria toxin genes that are driven by the promoter for adipocyte lipid binding protein
Authors: Homanics Gregg E.
Published: May 2005

Transgenic mice were used in an experiment that was designed to serve as a model of a possible approach to reducing the amount of carcass fat in meat animals. The objective was to reduce the number of adipocytes in transgenic mice thereby restricting the capacity to accumulate lipid. Our approach employed the technique of genetic ablation. The promoter for the adipocyte lipid binding protein gene was used in an attempt to direct expression of diphtheria toxin genes specifically to adipocytes. Three diphtheria toxin genes were used; they encode, respectively, an extremely cytotoxic wild type toxin, a less toxic attenuated toxin, and a nonfunctional toxin. While it was not possible to accurately assess effects of the transgenes on lipid accumulation, several informative observations were noted. A large percentage of transgenic founder mice that harbor either wild type or attenuated toxin genes are morphologically abnormal, die as neonates, or exhibit reproductive abnormalities including sterility or failure to transmit the transgene to offspring. In contrast, mice that harbor the nonfunctional toxin gene or are nontransgenic rarely have these same abnormalities. These results suggest that the trans‐genic mice are expressing the transgenes in cells other than adipocytes and that the aberrant production of functional toxin is responsible for the congenital abnormalities. The production of morphological and reproductive abnormalities in transgenic animals should be useful for investigating normal developmental processes.
Expression of glucose phosphate isomerase in interspecific hybrid (Mus musculus × Mus caroli)
Authors: West John D. , Ansell John D. , Flockhart Jean H.
Published: May 2005

Hybrid Mus musculus × Mus caroli embryos were produced by inseminating M. musculus (C57BL/Ola Ws) females with M. caroli sperm. Control M. caroli embryos developed more rapidly than did control M. musculus embryos and implanted approximately 1 day earlier. At 1 1/2 days, both the hybrid embryos and those of the maternal species (M. musculus) had cleaved to the 2‐cell stage. By 2 1/2 days some of the hybrids were retarded compared to M. musculus, and by 3½ days most were lagging behind. This is consistent with the idea that the rate of development of hybrid embryos declines once it becomes dependent on embryo‐coded gene products.

We have used this difference in rate of preim‐plantation development, between hybrid and M. musculus embryos, to try to determine whether the activation of embryonic Gpi‐1s genes, that encode glucose phosphate isomerase (GPI‐1), is age‐related or stage‐related. In control M. musculus embryos (both mated and Al groups), the GPI‐1AB and GPI‐1A allozyme, indicative of paternal gene expression, were detected in 7 of 9 samples of 3 1/2‐day compacted morula stage embryos and were seen in all 19 samples of 31/2‐day blastocysts. In hybrid embryos, these allozymes were detected 1 day later. They were not detected in any 31/2‐day samples (12 samples of compacted morulae) but were consistently detected at 4½ days (4 samples of blastocysts and 2 samples of uncompacted morulae). Our interpretation of the results is that gene activation in hybrid embryos is stage‐specific, rather than age‐specific, and probably begins around the 8‐cell stage, with detectable levels of enzyme accumulating later. Analysis of GPI‐1 elec‐trophoresis indicated that both the paternal (M. caroli) and maternal (M. musculus) Gpi‐1s alleles were equally expressed in hybrid embryos and that the paternally derived allele was not activated before the maternally derived allele. © 1992 Wiley‐Liss, Inc.
Catalase and superoxide dismutase gene expression and distribution during stem development in maize
Authors: Acevedo Alberto , Scandalios John G.
Published: May 2005

The temporal and spatial patterns of expression of the Cat and Sod genes encoding the multiple catalases and superoxide dismutases in maize have been examined throughout stem development. Three stages of stem development have been defined based upon catalase activity profiles and stem internode elongation. At stage 1, catalase activity is low and internodes remain short; at stage 2, catalase activity dramatically increases and internodes rapidly elongate; and at stage 3, catalase activity decreases to levels intermediate to stage 1 and 2, and internode elongation ceases. Zymogram analysis and immunoassays show that only the CAT‐3 catalase isozyme is present in the stem, even though both Cat1 and Cat3 mRNAs accumulate throughout stem development. Cat2 mRNA is not detectable in the developing stem. In full‐grown stems catalase is localized primarily in the sclerenchyma beneath the epidermis and around the vascular bundles and may possibly play a role in lignification. Unlike catalase, all the superoxide dismutase (SOD) isozymes and transcripts are present in the developing stem. Thus, these two major antioxidant gene‐enzyme systems show differential patterns of expression during stem development in maize. © 1992 Wiley‐Liss, Inc.
Distinct developmental regulatory mechanisms for two members of the aldolase gene family
Authors: Maine Ann B. , Ciejek‐Baez Elena
Published: May 2005

The aldolase isozyme family is composed of three members, A, B, and C, which are encoded by separate genes. The proteins are expressed in a tissue‐restricted manner during development and in the adult. To elucidate the regulation of aldolose mRNA in the mouse liver, we analyzed its expression by a number of methods including Northern blot, RNA dot blot, and nuclear run‐on assays. Our experiments demonstrate that the expression of aldolase A in the liver is primarily regulated by post‐transcriptional control. In contrast, we found that changes in the level of aldolase B mRNA are due to changes in the rate of initiation of transcription. In addition, we examined the regulation of aldolase expression in the adult kidney. We found that although the kidney has eight times more aldolase B than the live, the rate of initiation of transcription is similar in both tissues. Also, the rate of initiation of transcription of aldolase A is the same in the adult kidney and liver although there is 40 times more steady state aldolase A mRNA in the kidney than in the liver. © 1992 Wiley‐Liss, Inc.