• Applied Biological Chemistry
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• v.39 no.1
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• pp.25-31
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• 1996

Transgenic tobacco plants expressing calmodulin derivative($lys{\rightarrow}ile$ 115 calmodulin) and hygromycin resistance genes or hygromycin resistance gene alone(control) were generated by Agrobacterium-mediated DNA transfer. Seeds obtained from the transgenic plants($F_o$) were tested for resistance to hygromycin and the expected 3 : 1 ratio was observed. The expression of calmodulin derivative in the tobacco plants was identified by a combined method of Western blot and Chemiluminescence. The effects of surface sterilizers on the germiation of seeds from transgenic tobacco plants were tested in Murashige and Skoog agar medium. Seeds obtained from transgenic tobacoo plants expressing the calmodulin derivative showed no fungi contamination with normal germination by treating with sterilized water alone or sodium hypochlorite(2% effective chlorine). In contrast, seeds from the control transgenic tobacco plants showed severe contamination with fungi by treating with sterilized water alone and showed no contamination with normal germination by treating with sodium hypochlorite(2% chlorine). The effects of calcium concentration on the germination of seeds from transgenic tobacco plants were tested in Murashige and Skoog agar medium. Seeds obtained from transgenic tobacco plants expressing the calmodulin derivative showed better germination frequency than that of the control transgenic tobacco seeds in the medium containing 30 mM $CaCl_2$. The data raise the possibility that the expression of calmodulin derivative gene in tobacco plants could increase adaptability of the seeds to environmental stresses.

• Applied Biological Chemistry
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• v.43 no.4
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• pp.231-235
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• 2000

The changes of calmodulin levels, calmodulin-binding proteins, and $Ca^{2+}$/calmodulin-dependent glutamate decarboxylase during the growth of tobacco suspension cells were investigated. Tobacco cells exhibited a typical growth curve, including an exponential growth phase between 3 and 5 days after inoculation, and an apparent stationary phase occurring after 5 day. Although slight changes were observed from sample to sample, calmodulin protein levels remained similar during the phases of culture growth. Several $Ca^{2+}-dependent$ calmodulin-binding proteins including 56, 46, 36, and 32-kDa proteins were detected in tobacco cell extracts. The 56-kDa protein was identified as glutamate decarboxylase by Western-blot analysis using an anti-GAD monoclonal antibody. The levels of GAD protein and the specific activity of GAD enzyme were highest during the middle exponential phase of the culture growth cycle. These data suggest that $Ca^{2+}$/calmodulin-dependent glutamate decarboxylase is modulated during the growth of tobacco suspension cells.

• YAKHAK HOEJI
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• v.41 no.6
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• pp.773-781
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• 1997

The activity of $Ca^{2+}$calmodulin (CaM)-dependent protein kinase Ia (CaM kinase Ia) is shown to be regulated through direct phosphorylation by CaM kinase I kinase (CaMK IK). In the present study, three distinct CaMKIK peaks were separated from Q-Sepharose colunm chromatography of pig brain homogenate using a Waters 650 Protein Purification System. The purified CaMKIK from the major peak potently and rapidly enhanced CaM kinase Ia activity, reaching a maximal stimulation within 2min at the concentrations of 12-15nM. The activated state of CaM kinase Ia is characterized by a markedly enhanced $V_{max}4 as well as significantly decreased $K_m\;and\;K_a$ values toward peptide substrate and CaM, respectively. These observations suggest the activation process of CaM kinase Ia. The phosphorylation of CaM kinase Ia by CaMKIK may induce its conformational change responsible for the alterations in the kinetic properties, which ultimately leads to the rapid enzyme activation.

• Journal of Plant Biology
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• v.38 no.2
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• pp.195-201
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• 1995

The present study was made of the effects of abscisic acid(ABA) and calcium ions on dark respiration in protoplasts isolated from the basal intercalary meristematic tissues of oat (Avena sativa L.) seedlings. The influences of calcium channel blockers diitiazem(DTZ), verapamil(VPM), and $LaCl_2$ and the calmodulin antagonist trifluoperazine(TFP) on protoplast respiration activities were also investigated in order to evaluate the possible involvement of calcium channels and calmodulin during the dark respiration. The ABA only caused an 21% inhibition of protoplast respiration at $10^{-6}\;M$, but the extent of inhibition was very low by calcium treatments in the absence of ABA. In the presence of $10^{-6}\;M$ ABA, however, this inhibition of respiration increased by the increment of calcium ions concentrations. Treatments of DTZ and VPM were all found to restore the calcium-dependent inhibition of protoplast respiration by ABA and it was the same in thc $LaCl_2$ treatment except at $10^{-4}\;M$. At concentration from $10^{-6}\;M\;to\;10^{-4}\;M$, TFP also restored an inhibition of respiration. These results support the possibility that ABA increases plasmalemma permeability to calcium ions which might then bind to calmodulin to regulate oat protoplast dark respiration.ration.

• Journal of Life Science
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• v.21 no.5
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• pp.671-677
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• 2011

We previously reported the isolation and characterization of a gene, SCA1 (for soybean $Ca^{2+}$-ATPase 1), encoding a calmodulin-regulated $Ca^{2+}$-ATPase that is located in the plasma membrane in soybean. Here, a $Ca^{2+}$-ATPase designated as SCA2 was isolated from soybean. The two $Ca^{2+}$-ATPases, SCA1 and SCA2, share a remarkably high degree of similarity (78%). Ten transmemebrane domains were predicted by hydropathy analysis. Using gel overlay assays, CaM was found to bind to SCA2 in a $Ca^{2+}$-dependent manner. Southern blot analysis revealed the presence of two copies of the $Ca^{2+}$-ATPase gene in the soybean genome. An N-terminal truncation mutant that deletes sequence through the putative calmodulin binding site was able to complement a yeast mutant (K616) that was deficient in two endogenous $Ca^{2+}$ pumps. Our results indicate that SCA2 is structurally highly conserved with type IIB $Ca^{2+}$ pumps in plants.

• Journal of Plant Biotechnology
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• v.47 no.2
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• pp.157-163
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• 2020

Calmodulin (CaM) mediates cellular Ca²⁺ signals in the defense responses of plants. We previously reported that GmCaM-4 and 5 are involved in salicylic acid-independent activation of disease resistance responses in soybean (Glycine max). Here, we generated a GmCaM-4 cDNA construct under the control of the cauliflower mosaic virus (CaMV) 35S promoter and transformed this construct into potato (Solanum tuberosum L.). The constitutive over-expression of GmCaM-4 in potato induced high-level expression of pathogenesis-related (PR) genes, such as PR-2, PR-3, PR-5, phenylalanine ammonia-lyase (PAL), and proteinase inhibitorII (pinII). In addition, the transgenic potato plants exhibited enhanced resistance against a bacterial pathogen, Erwinia carotovora ssp. Carotovora (ECC), that causes soft rot disease and showed spontaneous lesion phenotypes on their leaves. These results strongly suggest that a CaM protein in soybean, GmCaM-4, plays an important role in the response of potato plants to pathogen defense signaling.

• Journal of KIISE:Software and Applications
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• v.35 no.8
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• pp.501-511
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• 2008

Probe design is one of the essential tasks in successful DNA microarray experiments. The requirements for probes vary as the purpose or type of microarray experiments. In general, most previous works use the simple filtering approach with the fixed threshold value for each requirement. Here, we formulate the probe design as a multiobjective optimization problem with the two objectives and solve it using ${\epsilon}$-multiobjective evolutionary algorithm. The suggested approach was applied in designing probes for 19 types of Human Papillomavirus and 52 genes in Arabidopsis Calmodulin multigene family and successfully produced more target specific probes compared to well known probe design tools such as OligoArray and OligoWiz.

• Journal of Genetic Medicine
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• v.6 no.1
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• pp.81-86
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• 2009

Multiple epiphyseal dysplasia (MED) is a clinically and genetically heterogeneous chondroplasia, characterized by delayed development of the ossification centers and, deformities of the extremities that involve only the epiphysis and result in mild short stature. Mutations in the cartilage oligomeric matrix protein (COMP) gene are most commonly found, and most of the mutations are located in the calmodulin-like repeats and the C-terminal domain. We report a Korean kindred of 12 family members with MED in four generations who were found to have a novel mutation in the COMP gene. A pedigree showed early onset osteoarthritis requiring arthroplasty that was an autosomal dominant inherited trait. Radiological examinations demonstrated the presence of osteochondral defects in the medial femoral condyles, and the knee and hip joints showed variable degrees of precocious degenerative changes. Mutation analysis of the COMP gene in the proband and five other affected family members identified a novel missense mutation, c.1280G>C (p.Gly427Ala) in exon 12, which was not found in three unaffected family members. Direct sequencing of the COMP gene may yield pathogenic mutations in dominantly inherited MED cases, and may provide opportunities of carrier detection among high-risk family members, leading to genetic counseling for early diagnosis and intervention before the onset of complications.

• Journal of Plant Biotechnology
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• v.38 no.1
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• pp.62-68
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• 2011

Calmodulin (CaM), a $Ca^{2+}$ binding protein in eukaryotes, mediates cellular $Ca^{2+}$ signals in response to a variety of biotic and abiotic external stimuli. The $Ca^{2+}$-bound CaM transduces signals by modulating the activities of numerous CaM-binding proteins. As a CaM binding protein, AtCBP63 ($\b{A}$rabidopsis thaliana $\b{C}$aM-binding protein $\underline{63}$ kD) has been known to be positively involved in plant defense signaling pathway. To investigate the pathogen resistance function of AtCBP63 in potato, we constructed transgenic potato (Solanum tuberosum L.) plants constitutively overexpressing AtCBP63 under the control of cauliflower mosaic virus (CaMV) 35S promoter. The overexpression of the AtCBP63 in potato plants resulted in the high level induction of pathogenesis-related (PR) genes such as PR-2, PR-3 and PR-5. In addition, the AtCBP63 transgenic potato showed significantly enhanced resistance against a pathogen causing bacterial soft rot, Erwinia carotovora ssp. Carotovora (ECC). These results suggest that a CaM binding protein from Arabidopsis, AtCBP63, plays a positive role in pathogen resistance in potato.

• Analytical Science and Technology
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• v.25 no.5
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• pp.333-338
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• 2012

IQ motif-containing GTPase-activating proteins (IQGAPs), which are well-known $Ca^{2+}$-independent calmodulin (CaM) binding proteins, are involved in various cellular functions such as cell proliferation, carcinogenesis and cell migration. The IQGAP3 similar to IQGAP1 has four repeated IQ motifs, which are crucial for CaM binding. It has been recently shown that all four IQ motifs of the IQGAP1 could bind to CaM, while not clear the binding of four IQ motifs of the IQGAP3. In this study, we examined the binding between CaM and each IQ motif of IQGAP3. As a result, we found that IQ2 and IQ3, but not IQ1 and IQ4, have a $Ca^{2+}$-independent CaM binding activity. We also found that IQ(3.5-4.4) on the IQGAP3 has $Ca^{2+}$-dependent CaM binding activity as similar with that of IQGAP1. This finding indicates that IQ motifs of the IQGAP3 plays a dynamic role via different interaction of IQ motifs with $Ca^{2+}$/CaM or apoCaM.