scholarly journals The “8-kD” Cytoplasmic Dynein Light Chain Is Required for Nuclear Migration and for Dynein Heavy Chain Localization in Aspergillus nidulans

1998 ◽  
Vol 143 (5) ◽  
pp. 1239-1247 ◽  
Author(s):  
Susan M. Beckwith ◽  
Christian H. Roghi ◽  
Bo Liu ◽  
N. Ronald Morris
Keyword(s):  
Aspergillus Nidulans ◽  
Heavy Chain ◽  
Light Chain ◽  
Nuclear Migration ◽  
Cytoplasmic Dynein ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Dynein Light Chain ◽  
Myosin V ◽  
Dynein Heavy Chain

The heavy chain of cytoplasmic dynein is required for nuclear migration in Aspergillus nidulans and other fungi. Here we report on a new gene required for nuclear migration, nudG, which encodes a homologue of the “8-kD” cytoplasmic dynein light chain (CDLC). We demonstrate that the temperature sensitive nudG8 mutation inhibits nuclear migration and growth at restrictive temperature. This mutation also inhibits asexual and sexual sporulation, decreases the intracellular concentration of the nudG CDLC protein and causes the cytoplasmic dynein heavy chain to be absent from the mycelial tip, where it is normally located in wild-type mycelia. Coimmunoprecipitation experiments with antibodies against the cytoplasmic dynein heavy chain (CDHC) and the nudG CDLC demonstrated that some fraction of the cytoplasmic dynein light chain is in a protein complex with the CDHC. Sucrose gradient sedimentation analysis, however, showed that not all of the NUDG protein is complexed with the heavy chain. A double mutant carrying a cytoplasmic dynein heavy chain deletion plus a temperature-sensitive nudG mutation grew no more slowly at restrictive temperature than a strain with only the CDHC deletion. This result demonstrates that the effect of the nudG mutation on nuclear migration and growth is mediated through an interaction with the CDHC rather than with some other molecule (e.g., myosin-V) with which the 8-kD CDLC might theoretically interact.

scholarly journals Extragenic suppressors of nudC3, a mutation that blocks nuclear migration in Aspergillus nidulans.

Genetics ◽  
1995 ◽  
Vol 141 (2) ◽  
pp. 453-464 ◽  
Author(s):  
Y H Chiu ◽  
N R Morris
Keyword(s):  
Aspergillus Nidulans ◽  
Normal Level ◽  
Growth And Development ◽  
Nuclear Migration ◽  
Cytoplasmic Dynein ◽  
Intracellular Concentration ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Extra Copy ◽  
Extragenic Suppressors

Abstract Nuclear migration plays an important role in the growth and development of many organisms including the filamentous fungus Aspergillus nidulans. We have cloned three genes from A. nidulans, nudA, nudC, and nudF, in which mutations affect nuclear migration. The nudA gene encodes the heavy chain of cytoplasmic dynein. The nudC gene encodes a 22-kD protein. The nudF gene was identified as an extra copy suppressor of the temperature sensitive (ts-) nudC3 mutation. The nudC3 mutation substantially decreases the intracellular concentration of the nudF protein at restrictive temperature. This is restored toward the normal level by an extra copy of nudF. To identify other genes whose products interact directly or indirectly with the NUDC protein, we have isolated a set of extragenic suppressors showed them to represent nine different genes, designated sncA-sncI (for suppressor of nudC). sncA-sncH were either dominant or semidominant in diploids homozygous for nudC3 and heterozygous for the snc mutations. All of the suppressors reversed the ts- phenotype of nudC3 by restoring the intracellular concentration of the NUDF protein.

scholarly journals An alpha tubulin mutation suppresses nuclear migration mutations in Aspergillus nidulans.

Genetics ◽  
1995 ◽  
Vol 141 (4) ◽  
pp. 1287-1298 ◽  
Author(s):  
D A Willins ◽  
X Xiang ◽  
N R Morris
Keyword(s):  
Aspergillus Nidulans ◽  
Heavy Chain ◽  
Nuclear Migration ◽  
Cytoplasmic Dynein ◽  
Beta Subunit ◽  
Alpha Tubulin ◽  
Dynein Heavy Chain ◽  
Suppressor Mutations ◽  
Dynein Motor ◽  
Low Concentrations

Abstract Microtubules and cytoplasmic dynein, a microtubule-dependent motor, are required for nuclei to move along the hyphae of filamentous fungi. Nuclear migration in Aspergillus nidulans is blocked by heat-sensitive (hs-) mutations in the nudA gene, which encodes dynein heavy chain, and the nudF gene, which encodes a G protein beta-subunit-like protein. Hs- mutations in the nudC and nudG genes also prevent nuclear migration. We have isolated extragenic suppressor mutations that reverse the hs- phenotypes caused by these mutations. Here we show that one nudF suppressor also suppresses hs- mutations in nudA, nudC, and nudG and deletions in nudA and nudF. This suppressor mutation is in the tubA alpha tubulin gene, and its characteristics suggest that it destabilizes microtubules. The mutation alters microtubule staining and confers sensitivity to cold and benomyl, two treatments that destabilize microtubules. Treatment with low concentrations of benomyl also suppresses the hs- nudA, nudC, nudF, and nudG mutations and the nudA and nudF deletions. Suppression of the hs- nudA mutation and the nudA deletion is especially interesting because these strains lack active dynein heavy chain. Together, these results suggest that microtubule destabilization allows nuclei to migrate even in the absence of cytoplasmic dynein motor function.

scholarly journals Functional Analysis of Cytoplasmic Dynein Heavy Chain in Caenorhabditis elegans with Fast-acting Temperature-sensitive Mutations

2005 ◽  
Vol 16 (3) ◽  
pp. 1200-1212 ◽  
Author(s):  
Diane J. Schmidt ◽  
Debra J. Rose ◽  
William M. Saxton ◽  
Susan Strome
Keyword(s):  
Caenorhabditis Elegans ◽  
Heavy Chain ◽  
Metaphase Plate ◽  
Cytoplasmic Dynein ◽  
Model Systems ◽  
Temperature Sensitive ◽  
Permissive Temperature ◽  
Timely Initiation ◽  
Dynein Heavy Chain ◽  
Dominant Temperature Sensitive

Cytoplasmic dynein, a minus-end–directed microtubule motor, has been implicated in many cellular and developmental processes. Identification of specific cellular processes that rely directly on dynein would be facilitated by a means to induce specific and rapid inhibition of its function. We have identified conditional variants of a Caenorhabditis elegans dynein heavy chain (DHC-1) that lose function within a minute of a modest temperature upshift. Mutant embryos generated at elevated temperature show defects in centrosome separation, pronuclear migration, rotation of the centrosome/nucleus complex, bipolar spindle assembly, anaphase chromosome segregation, and cytokinesis. Our analyses of mutant embryos generated at permissive temperature and then upshifted quickly just before events of interest indicate that DHC-1 is required specifically for rotation of the centrosome/nucleus complex, for chromosome congression to a well ordered metaphase plate, and for timely initiation of anaphase. Our results do not support the view that DHC-1 is required for anaphase B separation of spindle poles and chromosomes. A P-loop mutation identified in two independent dominant temperature-sensitive alleles of dhc-1, when engineered into the DHC1 gene of Saccharomyces cerevisiae, conferred a dominant temperature-sensitive dynein loss-of-function phenotype. This suggests that temperature-sensitive mutations can be created for time-resolved function analyses of dyneins and perhaps other P-loop proteins in a variety of model systems.

scholarly journals Extragenic suppressors of a dynein mutation that blocks nuclear migration in Aspergillus nidulans.

Genetics ◽  
1995 ◽  
Vol 139 (3) ◽  
pp. 1223-1232 ◽  
Author(s):  
G H Goldman ◽  
N R Morris
Keyword(s):  
Aspergillus Nidulans ◽  
Heavy Chain ◽  
Nuclear Migration ◽  
Cytoplasmic Dynein ◽  
Complementation Group ◽  
Large Molecular Weight ◽  
Weight Protein ◽  
Partial Suppression ◽  
Extragenic Suppressors

Abstract Cytoplasmic dynein is a large molecular weight protein complex that functions as a microtubule-dependent, negative, end-directed "motor." Mutations in nudA, which encodes the heavy chain of cytoplasmic dynein, inhibit nuclear migration in Aspergillus nidulans. This paper describes the selection and characterization of extragenic suppressors of the nudA1 mutation preparatory to the identification of other proteins that interact directly or indirectly with the cytoplasmic dynein heavy chain. To facilitate future cloning of the suppressor genes, we have searched particularly for extragenic suppressor mutations that also convey a selectable phenotype, such as cold or dimethyl sulfoxide sensitivity. Genetic analysis of 16 revertants has defined at least five extragenic suppressors of nudA1 (snaA-E). All the sna mutations except one were recessive in diploids homozygous for nudA1 and heterozygous for sna mutations. To characterize the nuclear migration phenotype in the sna mutants, conidia of one representative of each complementation group were germinated, fixed and nuclei stained. The sna mutants display partial suppression of the nudA1 nuclear migration defect. Although conidiophores were produced in the sna mutants, they failed to develop normally and to produce spores. Examination of the nudA1,sna conidiophores under the microscope showed that nuclear migration into the metulae and phialides was defective.

scholarly journals Roles of NUDE and NUDF Proteins of Aspergillus nidulans: Insights from Intracellular Localization and Overexpression Effects

2003 ◽  
Vol 14 (3) ◽  
pp. 871-888 ◽  
Author(s):  
Vladimir P. Efimov
Keyword(s):  
Green Fluorescent Protein ◽  
Aspergillus Nidulans ◽  
Heavy Chain ◽  
Fluorescent Protein ◽  
Specific Interaction ◽  
Cytoplasmic Dynein ◽  
Cell Cortex ◽  
Dynein Heavy Chain ◽  
Terminal Domain ◽  
Green Fluorescent

The NUDF protein of the filamentous fungus Aspergillus nidulans functions in the cytoplasmic dynein pathway. It binds several proteins, including the NUDE protein. Green fluorescent protein-tagged NUDF and NUDA (dynein heavy chain) localize to linearly moving dashes (“comets”) that coincide with microtubule ends. Herein, deletion of the nudE gene did not eliminate the comets of NUDF and NUDA, but affected the behavior of NUDA. Comets were also observed with the green fluorescent protein-tagged NUDE and its nonfunctional C-terminal domain. In addition, overexpressed NUDA and NUDE accumulated in specks that were either immobile or bounced randomly. Neither comets nor specks were observed with the functional N-terminal domain of NUDE, indicating that these structures are not essential for NUDE function. Furthermore, NUDF overproduction totally suppressed deletion of the nudEgene. This implies that the function of NUDE is secondary to that of NUDF. Unexpectedly, NUDF overproduction inhibited one conditionalnudA mutant and all tested apsA mutants. An allele-specific interaction between the nudF andnudA genes is consistent with a direct interaction between NUDF and dynein heavy chain. Because APSA and its yeast homolog Num1p are cortical proteins, an interaction between thenudF and apsA genes suggests a role for NUDF at the cell cortex.

scholarly journals Nuclear migration in a nud mutant of Aspergillus nidulans is inhibited in the presence of a quantitatively normal population of cytoplasmic microtubules.

1988 ◽  
Vol 106 (3) ◽  
pp. 773-778 ◽  
Author(s):  
S L Meyer ◽  
S G Kaminskyj ◽  
I B Heath
Keyword(s):  
Aspergillus Nidulans ◽  
Germ Tube ◽  
Normal Population ◽  
Nuclear Migration ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Permissive Temperature ◽  
Significant Temperature ◽  
Cytoplasmic Microtubules ◽  
Peripheral Cytoplasm

Nuclear migration was studied in germinating conidia of a temperature-sensitive mutant of the fungus Aspergillus nidulans. At the restrictive temperature motility was demonstrably impaired because significantly fewer nuclei migrated into the germ tube relative to a population of similarly sized germlings grown at the permissive temperature. Further comparison of these populations showed that the mutant was leaky in that an increasing number of nuclei migrated as the total nuclear content increased in each germling. The restrictive temperature also induced elevated mitotic asynchrony and increased numbers of nuclei per germling. Serial section-based reconstruction of the microtubules in a freeze-substituted germling showed that they were not attached to the nucleus-associated organelles, were approximately parallel to the long axis of the germ tube, and seemed to be randomly distributed between the central and peripheral cytoplasm. Five germlings from each temperature were selected for quantitative analysis of cytoplasmic microtubules. All 10 germlings had typical nuclear migration phenotypes. No significant temperature-related difference in microtubule density was found. We conclude that inhibition of nuclear migration in the mutant is the effect of some defect other than the failure of cytoplasmic microtubules to assemble to their normal population density. We also suggest that nuclear motility is not dependent on mitosis-related microtubules.

Sign in / Sign up

Export Citation Format

Share Document