Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.1 (protein kinase)
 81,284 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Interactions between adhesion molecules, agglutinins, on the surfaces of the flagella of mt+ and mt- gametes in Chlamydomonas rapidly generate a sexual signal, mediated by cAMP, that prepares the cells for fusion to form a zygote. The mechanism that couples agglutinin interactions to increased cellular levels of cAMP is unknown. In previous studies on the adenylyl cyclase in flagella of a single mating type (i.e., non-adhering flagella) we presented evidence that the gametic form of the enzyme, but not the vegetative form, was regulated by phosphorylation and dephosphorylation (Zhang, Y., E. M. Ross, and W. J. Snell. 1991. J. Biol. Chem. 266:22954-22959; Zhang, Y., and W. J. Snell. 1993. J. Biol. Chem. 268:1786-1791). In the present report we describe studies on regulation of flagellar adenylyl cyclase during adhesion in a cell-free system. The results show that the activity of gametic flagellar adenylyl cyclase is regulated by adhesion in vitro between flagella isolated from mt+ and mt- gametes. After mixing mt+ and mt- flagella together for 15 s in vitro, adenylyl cyclase activity was increased two- to threefold compared to that of the non-mixed (non-adhering), control flagella. This indicates that the regulation of gametic flagellar adenylyl cyclase during the early steps of fertilization is not mediated by signals from the cell body, but is a direct and primary response to interactions between mt+ and mt- agglutinins. By use of this in vitro assay, we discovered that 50 nM staurosporine (a protein kinase inhibitor) blocked adhesion-induced activation of adenylyl cyclase in vitro, while it had no effect on adenylyl cyclase activity of non-adhering gametic flagella. This same low concentration of staurosporine also inhibited adhesion-induced increases in vivo in cellular cAMP and blocked subsequent cellular responses to adhesion. Taken together, our results indicate that flagellar adenylyl cyclase in Chlamydomonas gametes is coupled to interactions between mt+ and mt- agglutinins by a staurosporine-sensitive activity, probably a protein kinase.
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PMID:Flagellar adhesion-dependent regulation of Chlamydomonas adenylyl cyclase in vitro: a possible role for protein kinases in sexual signaling. 817 84

Fertilization in Chlamydomonas is initiated by adhesive interactions between gametes of opposite mating types through flagellar glycoproteins called agglutinins. Interactions between these cell adhesion molecules signal for the activation of adenylyl cyclase through an interplay of protein kinases and ultimately result in formation of a diploid zygote. One of the early events during adhesion-induced signal transduction is the rapid inactivation of a flagellar protein kinase that phosphorylates a 48-kDa protein in the flagella. We report the biochemical and molecular characterization of the 48-kDa protein. Experiments using a bacterially expressed fusion protein show that the 48-kDa protein is capable of autophosphorylation on serine and tyrosine and phosphorylation of bovine beta-casein on serine, confirming that the 48-kDa protein itself has protein kinase activity. This protein kinase exhibits limited homology to members of the eukaryotic protein kinase superfamily and may be an important element in a signaling pathway in fertilization.
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PMID:Molecular cloning of a protein kinase whose phosphorylation is regulated by genetic adhesion during Chlamydomonas fertilization. 855 45

The regulation of mitosis in higher plant cells has been investigated by microinjecting protein kinase from the metaphase-arresting (met1) mutant of Chlamydomonas. Biochemical characterization of this enzyme complex confirms the presence of a p34cdc2/cyclin B-like kinase. The enzyme was injected into living stamen hair cells of Tradescantia virginiana in which microtubules (MTs) were visualized using fluorescent analogue cytochemistry and confocal laser scanning microscopy. Microinjection of this p34cdc2/cyclin B-like kinase caused rapid disassembly of the preprophase band of MTs but not of interphase-cortical, spindle or phragmoplast MTs. Effects of the enzyme on the cytomorphology of live prophase cells were also monitored using video microscopy. We found that injection of this enzyme accelerated chromatin condensation and nuclear envelope breakdown. This indicates the presence and function in plants of an enzyme that can initiate nuclear division similar to the maturation or mitosis promoting factor (MPF) of animal cells. These studies provide the first direct evidence that the mitotically-active form of plant MPF can drive disassembly of preprophase band MTs, chromosome condensation and initiation of mitosis in plant cells.
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PMID:Plant mitosis promoting factor disassembles the microtubule preprophase band and accelerates prophase progression in Tradescantia. 866 51

The following is a summary of physiological and pharmacological studies of the regulation of dynein-driven microtubule sliding in Chlamydomonas flagella. The experimental basis for the study is described, and data indicating that an axonemal cAMP-dependent protein kinase can regulate inner arm dynein activity are reviewed. In addition, preliminary data are summarized indicating that an axonemal type 1 phosphatase can also regulate dynein-drive microtubule sliding velocity. It is predicted that the protein kinase, phosphatase, and an inner dynein arm component form a regulatory complex in the axoneme.
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PMID:Regulation of dynein-driven microtubule sliding by an axonemal kinase and phosphatase in Chlamydomonas flagella. 868 89

Physical connections between higher plant cell walls and the plasma membrane have been identified visually, but the molecules involved in the contact are unknown. We describe here an Arabidopsis thaliana protein kinase, designated Wak1 for wall-associated kinase, whose predicted extracytoplasmic domain contains several epidermal growth factor repeats and identity with a viral movement protein. Wak1 fractionates with insoluble material when plant tissue is ground in a variety of buffers and detergents, suggesting a tight association with the plant extracellular matrix. Immunocytochemistry confirms that Wak1 is associated with the cell wall. Enzymatic digestion of the cell wall allows the release of Wak1 from the insoluble cell wall fraction, and protease experiments indicate that Wak1 likely has a cytoplasmic kinase domain, and the EGF containing domain is extracellular. Wak1 is found in all vegetative tissues of Arabidopsis, and has relatives in other angiosperms, but not Chlamydomonas. We suggest that Wak1 is a good candidate for a physical continuum between the cell wall and the cytoplasm, and since the kinase is cytoplasmic, it also has the potential to mediate signals to the cytoplasm from the cell wall.
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PMID:A cell wall-associated, receptor-like protein kinase. 870 86

Within seconds after the flagella of mt+ and mt- Chlamydomonas gametes adhere during fertilization, their flagellar adenylyl cyclase is activated several fold and preparation for cell fusion is initiated. Our previous studies indicated that early events in this pathway, including control of adenylyl cyclase, are regulated by phosphorylation and dephosphorylation. Here, we describe a soluble, flagellar protein kinase activity that is regulated by flagellar adhesion. A 48-kDa, soluble flagellar protein was consistently phosphorylated in an in vitro assay in flagella isolated from nonadhering mt+ and mt- gametes, but not in flagella isolated from mt+ and mt- gametes that had been adhering for 1 min. Although the 48-kDa protein was present in the flagella isolated from adhering gametes, we demonstrate that its protein kinase was inactivated by flagellar adhesion. Immunoblot analysis and inhibitor studies indicate that the 48-kDa protein in nonadhering gametes is phosphorylated by a protein tyrosine kinase. In vivo experiments showing that the protein tyrosine phosphatase inhibitor sodium orthovanadate inhibits fertilization suggest that protein dephosphorylation may be required for signal transduction. The 48-kDa protein and its protein kinase may be among the first elements of a novel signalling pathway that couples interaction of flagellar adhesion molecules to gamete activation.
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PMID:Cell adhesion-dependent inactivation of a soluble protein kinase during fertilization in Chlamydomonas. 873 96

Fertilization in the biflagellated eukaryote, Chlamydomonas, is initiated by flagellar adhesion between gametes of opposite mating types. An early event in the signal transduction pathway induced by these cell-cell interactions is the rapid inactivation of a flagellar protein kinase that phosphorylates a 48 kDa flagellar protein. Molecular cloning and characterization indicated that the 48 kDa substrate, termed SksC, itself is a novel protein kinase. Here, we have determined that its transcript levels were unchanged during prolonged flagellar adhesion. Moreover, resynthesis of new flagellar proteins following deflagellation was not accompanied by increases in transcript levels of SksC, suggesting that expression of this soluble protein kinase might not be restricted to flagella. Immunoblot analysis indicated that expression of SksC was ubiquitous: this soluble protein was found in both flagella and cell bodies and was expressed throughout the cell cycle and gametogenesis. Immunoprecipitation experiments indicated that SksC was phosphorylated in both flagella and cell bodies. Thus, in addition to its potential role in fertilization, this novel protein kinase may play a role in other signaling events in Chlamydomonas.
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PMID:SksC, a fertilization-related protein kinase in Chlamydomonas, is expressed throughout the cell cycle and gametogenesis, and a phosphorylated form is present in both flagella and cell bodies. 891 34

Calcium-stimulated protein kinase activity in the flagella of the green alga Chlamydomonas moewusii (Gerloff) was characterised. Using SDS-PAGE and an on-blot phosphorylation assay, a 65-kDa protein was identified as the major calcium-stimulated protein kinase. Its activity was directly stimulated by calcium, a characteristic of the calmodulin-like domain protein kinases (CDPKs). Monoclonal antibodies raised against the CDPK alpha from soybean cross-reacted with the 65-kDa protein in the flagella, and also with other proteins in the flagellum and cell body. The same monoclonal antibodies were used to screen a C. moewusii cDNA expression library in order to isolate CDPK cDNAs from C. moewusii. The CCK1 cDNA encodes a protein with a kinase and calmodulin-like domain linked by a junction domain typical of CDPKs. From Southern analyses, evidence was obtained for a CDPK gene family in C. moewusii and C. reinhardtii.
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PMID:Characterisation and cloning of a calmodulin-like domain protein kinase from Chlamydomonas moewusii (Gerloff). 917 53

The length of eukaryotic cilia and flagella depends on the cell cycle-regulated assembly and disassembly of at least 9 doublet and 2 central microtubules, their associated proteins, and the surrounding membrane. In light-synchronized Chlamydomonas cells, flagella assembled to 10-14 microm in length near the beginning of the light period and they disassembled prior to cell division, during the dark period. Flagella on light-synchronized pf18 Chlamydomonas mutants grew to 10-12 microm near the beginning of the light period but shortened by 50% or more by the end of the light period. Flagellar length was cell-cycle regulated: when flagella were amputated at various times during the light period, new flagella regenerated to the lengths of control cells at that time of the light cycle. The later in the cycle pf18 cells were deflagellated, the shorter were the regenerated flagella. Flagellar shortening was not affected, in either pf18 or wild-type (wt) cells, by inhibitors of protein synthesis or of microtubule assembly, so flagellar length cannot depend on protein turnover. Shortening in pf18 was attenuated by Li+, which stimulated flagellar growth in wt cells, by red light, by protein kinase inhibitors, and by the Ca2+ channel blockers La3+ and Cd2+. Shortening was increased by cAMP, Na+, K+, and EGTA. Ca2+-CAM blockers did not affect pf18 shortening but they increased shortening in wt and fa1 cells. We propose that flagellar length is regulated by a signal transduction pathway that is sensitive to Ca2+ levels and red light.
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PMID:Regulation of flagellar length in Chlamydomonas. 963 11

During fertilization in Chlamydomonas, flagellar adhesion between mt(+) and mt(-) gametes induces a cAMP-dependent signal transduction pathway that prepares the gametes for cell fusion and zygote formation. Previously, our laboratory identified a homeodomain protein (GSP1) whose expression was restricted to the cell bodies of mt(+) gametes and whose transcript level was up-regulated during flagellar adhesion. In this report, we describe a new form of GSP1 that appears early during gamete interactions. Immunoblot analysis showed that in addition to the 120-kDa form of GSP1 normally present in mt(+) gametes, a 122-kDa form was detected when the cells were mixed with mt(-) gametes. The more slowly migrating form of GSP1 was detectable within minutes after gametes were mixed together, and its appearance did not require new protein synthesis. Thus, the 122-kDa form represents a post-translational modification of the pre-existing 120-kDa form of GSP1. Moreover, conversion to the 122-kDa form did not require cell fusion. Although the 120-kDa form was expressed 10 h after vegetative cells were transferred to gametic induction medium, the 122-kDa form was detected only after mt(+) gametes were induced to undergo the sexual signaling that accompanies fertilization. Incubation of mt(+) gametes with dibutyryl cAMP led to the appearance of the 122-kDa form of GSP1, and the cyclic nucleotide-dependent protein kinase inhibitor H-8 inhibited the adhesion-induced conversion. Incubation of GSP1 immunoprecipitated from signaling mt(+) gametes with alkaline phosphatase showed that the conversion was due to phosphorylation. The results indicate that flagellar adhesion induces a rapid, cAMP-dependent phosphorylation of the homeodomain protein GSP1 early during fertilization in Chlamydomonas.
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PMID:Flagellar adhesion between mt(+) and mt(-) Chlamydomonas gametes regulates phosphorylation of the mt(+)-specific homeodomain protein GSP1. 1056 16


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