Distribution of the myosin I-like ninaC proteins in the Drosophila retina and ultrastructural analysis of mutant phenotypes

1992 ◽  
Vol 101 (1) ◽  
pp. 247-254 ◽  
Author(s):  
J.L. Hicks ◽  
D.S. Williams
Keyword(s):  
Compound Eye ◽  
Photoreceptor Cells ◽  
Amino Acid Sequences ◽  
Ultrastructural Analysis ◽  
Multivesicular Bodies ◽  
Filamentous Actin ◽  
Electron Microscopic ◽  
Myosin I ◽  
Specific Proteins ◽  
Mutant Phenotypes

The Drosophila ninaC gene encodes for two head-specific proteins of 132 kDa and 174 kDa. Their predicted amino acid sequences indicate that they may have myosin I and kinase properties. We have: (1) determined the cellular and subcellular distributions of the ninaC proteins in the Drosophila retina by electron microscopic immunocytochemistry with an antibody specific for epitopes shared by both proteins; (2) characterized the ultrastructure of the mutant phenotype. The proteins were detected only in the photoreceptor cells, but were detected in all classes of the compound eye photoreceptors. Within the photoreceptors, they were found in the rhabdomeral microvilli and the cytoplasm adjacent to the rhabdomeres. This distribution coincides with that shown previously for actin filaments. Immunolabelling of tissue from the ninaC P221 mutant, which lacks the 174 kDa protein, and two mutants whose rhabdomeres degenerate, suggests that the 132 kDa protein is present primarily in the cytoplasm adjacent to the rhabdomeres, and that the 174 kDa protein is concentrated in the rhabdomeres. Our ultrastructural analysis showed that the axial cytoskeleton of the rhabdomeral microvilli (which contains filamentous actin) was absent in both the null and P221 mutants. In the photoreceptor cell cytoplasm, the number of multivesicular bodies in the null mutant, but not the P221 mutant, was 3-fold greater in comparison with wild-type.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Differential localizations of and requirements for the two Drosophila ninaC kinase/myosins in photoreceptor cells.

1992 ◽  
Vol 116 (3) ◽  
pp. 683-693 ◽  
Author(s):  
J A Porter ◽  
J L Hicks ◽  
D S Williams ◽  
C Montell
Keyword(s):  
Retinal Degeneration ◽  
Photoreceptor Cells ◽  
Kinase Domain ◽  
Electron Microscopic Level ◽  
Head Region ◽  
Electron Microscopic ◽  
Myosin I ◽  
Microvillar Structure ◽  
The Difference ◽  
The Individual

The ninaC gene encodes two retinal specific proteins (p132 and p174) consisting of a protein kinase domain joined to a domain homologous to the head region of the myosin heavy chain. The putative myosin domain of p174 is linked at the COOH-terminus to a tail which has some similarities to myosin-I tails. In the current report, we demonstrate that the ninaC mutation results in light- and age-dependent retinal degeneration. We also show that ninaC flies display an electrophysiological phenotype before any discernible retinal degeneration indicating that the electrophysiological defect is the primary effect of the mutation. This suggests that ninaC has a role in phototransduction and that the retinal degeneration is a secondary effect resulting from the defect in phototransduction. To examine the requirements for the individual ninaC isoforms, mutant alleles were generated which express only p132 or p174. Elimination of p174 resulted in a ninaC phenotype as strong as the null allele; however, elimination of p132 had little if any effect. As a first step in investigating the basis for the difference in requirements for p174 and p132 we performed immuno-localization at the electron microscopic level and found that the two isoforms display different subcellular distributions in the photoreceptor cells. The p132 protein is restricted primarily to the cytoplasm and p174 to the rhabdomeres, the microvillar structure which is the site of action of many of the steps in phototransduction. This suggests that the p174 myosin-I type tail is the domain responsible for association with the rhabdomeres and that the substrate for the p174 putative kinase may be a rhabdomeric protein important in photo-transduction.

scholarly journals The Drosophila bifocal gene encodes a novel protein which colocalizes with actin and is necessary for photoreceptor morphogenesis.

1997 ◽  
Vol 17 (9) ◽  
pp. 5521-5529 ◽  
Author(s):  
S M Bahri ◽  
X Yang ◽  
W Chia
Keyword(s):  
Compound Eye ◽  
Photoreceptor Cells ◽  
P Element ◽  
Apical Surface ◽  
Loss Of Function ◽  
Filamentous Actin ◽  
Light Gathering ◽  
Morphological Defects ◽  
Microvillar Structure ◽  
Eye Imaginal Disc

Photoreceptor cells of the Drosophila compound eye begin to develop specialized membrane foldings at the apical surface in midpupation. The microvillar structure ultimately forms the rhabdomere, an actin-rich light-gathering organelle with a characteristic shape and morphology. In a P-element transposition screen, we isolated mutations in a gene, bifocal (bif), which is required for the development of normal rhabdomeres. The morphological defects seen in bif mutant animals, in which the distinct contact domains established by the newly formed rhabdomeres are abnormal, first become apparent during midpupal development. The later defects seen in the mutant adult R cells are more dramatic, with the rhabdomeres enlarged, elongated, and frequently split. bif encodes a novel putative protein of 1063 amino acids which is expressed in the embryo and the larval eye imaginal disc in a pattern identical to that of F actin. During pupal development, Bif localizes to the base of the filamentous actin associated with the forming rhabdomeres along one side of the differentiating R cells. On the basis of its subcellular localization and loss-of-function phenotype, we discuss possible roles of Bif in photoreceptor morphogenesis.

Actin and α-tubulin immuno-EM labelings reveal differences in intra versus interphotoreceptor matrix

1990 ◽  
Vol 48 (3) ◽  
pp. 466-467
Author(s):  
Matti Järvilehto ◽  
Riitta Harjula
Keyword(s):  
Compound Eye ◽  
Frozen Section ◽  
Smooth Muscle Actin ◽  
Photoreceptor Cells ◽  
Immune Serum ◽  
Cell Organelles ◽  
Calliphora Erythrocephala ◽  
Optical Axes ◽  
Interphotoreceptor Matrix ◽  
Similar Kind

The photoreceptor cells in the compound eyes of higher diptera are clustered in groups (ommatidia) of eight receptor cells. The cells from six adjacent ommatidia are organized into optical units, neuro-ommatia sharing the same visual field. In those ommatidia the optical axes of the photopigment containing structures (rhabdomeres) are parallel. The rhabdomeres of the photoreceptor cells are separated from each other by an interstitial i.e innerommatidial space (IOS). In the photoreceptor cell body, besides of the normal cell organelles, a cellular matrix is a structurally apparent component. Similar kind of reticular formation is also found in the IOS containing some unidentified filamentary substance, of which composition and functional significance for optical properties of vision is the aim of this report.The prefixed (2% PA + 0.2% GA in 0.1-n phosphate buffer, pH 7.4, for 1h), frozen section blocks of the compound eye of the blowfly (Calliphora erythrocephala) were prepared by immuno-cryo-techniques. The ultrathin cryosections were incubated with antibodies of monoclonal α-tubulin and polyclonal smooth muscle actin. Control labelings of excess of antigen, non-immune serum and non-present antibody were perforated.

scholarly journals Identification, characterization and cloning of myr 1, a mammalian myosin-I.

1993 ◽  
Vol 120 (6) ◽  
pp. 1393-1403 ◽  
Author(s):  
C Ruppert ◽  
R Kroschewski ◽  
M Bähler
Keyword(s):  
Amino Acid ◽  
Light Chain ◽  
Brush Border ◽  
Binding Sites ◽  
Neuronal Development ◽  
Amino Acid Sequences ◽  
Dependent Manner ◽  
Tail Domain ◽  
Myosin I ◽  
Splice Forms

We have identified, characterized and cloned a novel mammalian myosin-I motor-molecule, called myr 1 (myosin-I from rat). Myr 1 exists in three alternative splice forms: myr 1a, myr 1b, and myr 1c. These splice forms differ in their numbers of putative calmodulin/light chain binding sites. Myr 1a-c were selectively released by ATP, bound in a nucleotide-dependent manner to F-actin and exhibited amino acid sequences characteristic of myosin-I motor domains. In addition to the motor domain, they contained a regulatory domain with up to six putative calmodulin/light chain binding sites and a tail domain. The tail domain exhibited 47% amino acid sequence identity to the brush border myosin-I tail domain, demonstrating that myr 1 is related to the only other mammalian myosin-I motor molecule that has been characterized so far. In contrast to brush border myosin-I which is expressed in mature enterocytes, myr 1 splice forms were differentially expressed in all tested tissues. Therefore, myr 1 is the first mammalian myosin-I motor molecule with a widespread tissue distribution in neonatal and adult tissues. The myr 1a splice form was preferentially expressed in neuronal tissues. Its expression was developmentally regulated during rat forebrain ontogeny and subcellular fractionation revealed an enrichment in purified growth cone particles, data consistent with a role for myr 1a in neuronal development.

scholarly journals Genetic Characterization and Immunogenicity of Coli Surface Antigen 4 from Enterotoxigenic Escherichia coli when It Is Expressed in a Shigella Live-Vector Strain

2003 ◽  
Vol 71 (3) ◽  
pp. 1352-1360 ◽  
Author(s):  
Zeev Altboum ◽  
Myron M. Levine ◽  
James E. Galen ◽  
Eileen M. Barry
Keyword(s):  
Escherichia Coli ◽  
Shigella Flexneri ◽  
Regulatory Protein ◽  
Electron Microscopic Examination ◽  
Amino Acid Sequences ◽  
Enterotoxigenic Escherichia Coli ◽  
Electron Microscopic ◽  
Bacterial Surface ◽  
Fimbrial Subunit ◽  
Mucosal Antibody

ABSTRACT The genes that encode the enterotoxigenic Escherichia coli (ETEC) CS4 fimbriae, csaA, -B, -C, -E, and -D′, were isolated from strain E11881A. The csa operon encodes a 17-kDa major fimbrial subunit (CsaB), a 40-kDa tip-associated protein (CsaE), a 27-kDa chaperone-like protein (CsaA), a 97-kDa usher-like protein (CsaC), and a deleted regulatory protein (CsaD′). The predicted amino acid sequences of the CS4 proteins are highly homologous to structural and assembly proteins of other ETEC fimbriae, including CS1 and CS2, and to CFA/I in particular. The csaA, -B, -C, -E operon was cloned on a stabilized plasmid downstream from an osomotically regulated ompC promoter. pGA2-CS4 directs production of CS4 fimbriae in both E. coli DH5α and Shigella flexneri 2a vaccine strain CVD 1204, as detected by Western blot analysis and bacterial agglutination with anti-CS4 immune sera. Electron-microscopic examination of Shigella expressing CS4 confirmed the presence of fimbriae on the bacterial surface. Guinea pigs immunized with CVD 1204(pGA2-CS4) showed serum and mucosal antibody responses to both the Shigella vector and the ETEC fimbria CS4. Among the seven most prevalent fimbrial antigens of human ETEC, CS4 is the last to be cloned and sequenced. These findings pave the way for CS4 to be included in multivalent ETEC vaccines, including an attenuated Shigella live-vector-based ETEC vaccine.

scholarly journals First Report of Malva vein clearing virus Naturally Occurring in Hollyhock in Germany

Plant Disease ◽  
2010 ◽  
Vol 94 (2) ◽  
pp. 276-276 ◽  
Author(s):  
W. Menzel ◽  
S. Winter ◽  
K. R. Richert-Pöggeler
Keyword(s):  
Host Range ◽  
Direct Sequencing ◽  
Amino Acid Sequences ◽  
Rt Pcr ◽  
Electron Microscopic ◽  
First Report ◽  
Vein Clearing ◽  
Naturally Occurring ◽  
Potyvirus Species ◽  
Cloned Cdna

Hollyhocks are popular garden plants and selected cultivars of Alcea rosea (family Malvaceae) are widespread in Germany. In spring 2009, dozens of A. rosea plants displaying strong vein clearing and veinal yellowing symptoms were found in private gardens in Hannover, Lower Saxony. Electron microscopic examinations of negatively stained adsorption preparations of five randomly selected samples of symptomatic plants or their offshoots revealed flexuous filamentous particles resembling those of potyviruses. Sap extracts also reacted strongly positive in an antigen coated plate (ACP)-ELISA with the broad-spectrum potyvirus antiserum AS-0573/I (DSMZ, Braunschweig, Germany). RNA extracts (RNeasy Kit, Qiagen, Valencia, CA) of the above mentioned leaf samples were used as templates in reverse transcription (RT)-PCR assays with potyvirus specific primers (2) that have been shown to amplify the 3′ terminus of the genome of many potyvirus species. For extracts from symptomatic samples, this resulted in a consistent amplification of an ~1.6-kbp fragment, whereas no products were obtained from RNA extracts of asymptomatic plants. From one positive sample, the amplified fragment was cloned and one clone was partially sequenced. The nucleotide (nt) and amino acid sequences showed the highest identities (81 to 83% and 87 to 90%, respectively) to GenBank sequences FJ539084, FM212972, EU884405, and FJ561293 of the potyvirus Malva vein clearing virus (MVCM). On the basis of these identity values and according to the species demarcation criteria in the genus Potyvirus, the virus can be regarded as a German isolate of the recently sequenced MVCV (3,4). Direct sequencing of the 5′-end of the amplified RT-PCR fragment revealed sequences of only one potyvirus species. The virus isolate has been submitted to the DSMZ Plant Virus Collection (Braunschweig, Germany) under accession PV-0963 and the sequence obtained from the cloned cDNA is deposited in GenBank (GQ856544). In addition, sap from affected leaves was mechanically inoculated onto sets of herbaceous indicator plants (Chenopodium quinoa, C. foliosum, C. murale, C. amaranticolor, Datura stramonium, Nicotiana benthamiana, N. hesperis, Petunia hybrida, and Solanum lycopersicum) of which only C. quinoa plants became infected. Symptoms of weak chlorosis along and beside veins of inoculated leaves, but not systemic leaves, became visible 2 weeks postinoculation. Symptomatic leaves contained flexuous filamentous particles and ACP-ELISA and RT-PCR confirmed virus presence. The partially sequenced amplicon showed 99% nt identity to the sequence from the cloned cDNA. To our knowledge, this is the first report of a MVCV isolate naturally occurring in A. rosea and C. quinoa is the first host identified that does not belong to the plant family Malvaceae. In contrast, the MVCV isolate used in the host range study of Lunello et al. (4) did not infect A. rosea and C. quinoa, confirming previous host range descriptions by Brunt et al. (1). Since MVCV infections of hollyhocks seem to cause only leaf symptoms and do not noticeably affect growth or flowering of the plants, this will hopefully not impair the usability of this popular garden plant. References: (1) A. A. Brunt et al. Descriptions and Lists from the VIDE Database. Online publication. Version: 16th January, 1997. (2) J. Chen et al. Arch. Virol. 146:757, 2001. (3) A. Hein Phytopathol. Z. 28:205, 1957. (4) P. Lunello et al. Virus Res. 140:91, 2009.

Targeting of the myosin-I myr 3 to intercellular adherens type junctions induced by dominant active Cdc42 in HeLa cells

1998 ◽  
Vol 111 (18) ◽  
pp. 2779-2788 ◽  
Author(s):  
H.E. Stoffler ◽  
U. Honnert ◽  
C.A. Bauer ◽  
D. Hofer ◽  
H. Schwarz ◽  
...  
Keyword(s):  
Hela Cells ◽  
Adherens Junction ◽  
Intercellular Junctions ◽  
Distinct Type ◽  
Beta Catenin ◽  
Tail Domain ◽  
Filamentous Actin ◽  
Myosin I ◽  
Myosin Motor Domain ◽  
Constitutively Active

Myr 3, a member of the myosin-I family from rat, is shown in this study to be localized at adherens-type intercellular junctions in epithelial and nonepithelial tissues. Formation of intercellular junctions and the accompanying recruitment of myr 3 to these junctions involves signaling by the Rho subfamily of small GTP-binding proteins. This conclusion is based on studies with HtTA-1 HeLa cells that were induced by overexpression of constitutively active Cdc42Hs to form typical adherens-type intercellular junctions enriched in cadherins (N-cadherin), beta-catenin, filamentous actin and myr 3. Recruitement of myr 3 to Cdc42-induced adherens junctions in HeLa cells was dependent on a short region of the tail domain and a functional myosin motor domain, but was independent of its myosin-I tail homology and SH3 regions. Overexpression of constitutively active Rac1 induced a distinct type of adherens junction in HeLa cells that was characterized by elaborate intercellular interdigitations enriched in N-cadherin, beta-catenin and F-actin. Myr 3 was often present, but not specifically enriched in the intercellular junctions induced by constitutively active Rac1.

Cell death and the development of limb form and skeletal pattern in normal and wingless (ws) chick embryos

Development ◽  
1973 ◽  
Vol 30 (3) ◽  
pp. 753-772
Author(s):  
J. R. Hinchliffe ◽  
D. A. Ede
Keyword(s):  
Cell Death ◽  
Phenotypic Expression ◽  
Mesenchymal Cell ◽  
Limb Bud ◽  
Death Process ◽  
Electron Microscopic ◽  
Wide Range ◽  
Skeletal Pattern ◽  
Autoradiographic Analysis ◽  
Mutant Phenotypes

The wingless condition resulting from the action of the sex-linked wingless (ws) gene arises from the precocious appearance of cell death in the anterior necrotic zone (ANZ) of the forelimb-bud at stage 19 (3 days) and its progressive extension beyond its normal area during stages 20–23. A similar though less pronounced effect occurs in the hindlimb-bud. Although some wingless hindlimb-buds are normal, others are affected by the precocious appearance of cell death in the ANZ. The ws wingless mutant resembles the different wingless mutant investigated by Zwilling (1956) in that the apical ectodermal ridge (AER) is absent in most ws wing-buds. AER absence could be due to ws mesenchymal cell death interfering with the production of apical ectodermal maintenance factor (AEMF), which Zwilling claims is necessary to maintain the AER which plays an essential role in inducing limb outgrowth. Wingless mutant phenotypes range from birds with rudimentary wings and normal legs through a modal type with forelimbs absent and hindlimbs normal to wingless and legless forms showing a high degree of expressivity. Individual wingless embryos vary in the degree to which the precocious ANZ appearing at 3 days is extended into the limb-bud and the wide range of wingless phenotypic expression is attributed to this variation. Electron microscopic and histochemical analysis of the cell death process in wingless wing-buds revealed the presence of both isolated dead cells and macrophages, which contained intense acid phosphatase activity. These findings are interpreted as showing that isolated dead cells are ingested by neighbouring mesenchymal cells which thus become transformed into macrophages, first ingesting and then digesting further dead cells. A study was made of the origin of the anomalous hindlimb condition, including absence or reduction of the tibia and digits, characteristic of severely affected wingless embryos. Autoradiographic analysis of the pattern of 35SO4 uptake revealed that at stage 24/5 (4½ days) wingless hindlimb-buds which were smaller than normal had a normal prospective fibula region, but that the prospective tibia region was small or absent. Thus the effect of a precocious hindlimb ANZ at stages 19–22 is to reduce or delete the pre-axial prospective tibia at stage 24/5.

The accuracy of the patterns of connexions of the first- and second-order neurons of the visual system of Calliphora

1970 ◽  
Vol 175 (1038) ◽  
pp. 69-82 ◽  
Keyword(s):  
Single Neuron ◽  
Compound Eye ◽  
Photoreceptor Cells ◽  
Mirror Image ◽  
Second Order ◽  
Visual Axis ◽  
First Order ◽  
Definite Pattern ◽  
Small Bundle ◽  
Do So

The axons of the primary photoreceptor cells of the compound eye of the fly interweave in a complex but definite pattern before they terminate upon the second-order neurons. Of approximately 650 short retinula axons from behind 120 facets of the eye none terminated at an incorrect lamina cartridge. Six, seven, or eight first-order terminals upon one pair of second-order cells are arranged in a rotational sequence that is related to the positions of the retinula cells within the ommatidia. Errors in location of the terminal among its neighbours occurred only ten times. The asymmetry of the receptor pattern in the dorsal half of the eye has a mirror image in the ventral half. Along the equator of the eye is a plane of symmetry which many axons necessarily cross in maintaining the appropriate connexions of their receptors. Axons which cross this plane of symmetry have somehow found their appropriate second-order cells, although to do so they must have grown through a milieu which is the mirror image of that in their own half of the eye. Each pair of second-order axons proceeding from the lamina forms a small bundle with the axons of the two long retinula cells that have the same visual axis. Between the lamina and the medulla is a chiasma (with the crossing in the horizontal plane) through which bundles from the lamina pass to project in exactly reverse order upon the medulla. No errors of projection have been found at the single neuron level in this chiasma.

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