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論文

DNA gyrase of ${it Deinococcus radiodurans}$ is characterized as Type II bacterial topoisomerase and its activity is differentially regulated by PprA in vitro

Kota, S.*; Rajpurohit, Y. S.*; Charaka, V. K.*; 佐藤 勝也; 鳴海 一成*; Misra, H, S.*

Extremophiles, 20(2), p.195 - 205, 2016/03

 被引用回数:6 パーセンタイル:47.77(Biochemistry & Molecular Biology)

The multipartite genome of ${it Deinococcus radiodurans}$ forms toroidal structure. It encodes topoisomerase IB and both the subunits of DNA gyrase (DrGyr) while lacks other bacterial topoisomerases. Recently, PprA a pleiotropic protein involved in radiation resistance in ${it D. radiodurans}$ has been suggested for having roles in cell division and genome maintenance. In vivo interaction of PprA with topoisomerases has also been shown. DrGyr constituted from recombinant gyrase A and gyrase B subunits showed decatenation, relaxation and supercoiling activities. Wild type PprA stimulated DNA relaxation activity while inhibited supercoiling activity of DrGyr. Thus, we showed that DrGyr confers all three activities of bacterial type IIA family DNA topoisomerases, which are differentially regulated by PprA, highlighting the significant role of PprA in DrGyr activity regulation and genome maintenance in ${it D. radiodurans}$.

論文

Screening of cesium-accumulating mutant of radioresistant bacterium ${it Deinococcus radiodurans}$ by ion beam breeding technology

佐藤 勝也; 上田 涼史郎*; 長谷 純宏; 鳴海 一成*; 大野 豊

JAEA-Review 2015-022, JAEA Takasaki Annual Report 2014, P. 100, 2016/02

Bioremediation uses biological organisms to solve an environmental problem. However, microorganisms are sometimes not enough effective to achieve efficient recovery. Therefore, enhancing the functionality of microorganisms is needed to promote the active use of bioremediation. This study began with the purpose of development of Cs-accumulating bacteria by ion beam breeding technology. Previously, we suggested that ${it Deinococcus radiodurans}$ is the most suitable species for the development of high Cs-accumulating bacteria by ion beam breeding technology. Candidates of Cs-accumulating mutant were screened from the mutagenized cells by ion beam breeding technology on the basis of the luminescent intensity and the cell area as indicators of intracellular Cs level and cell growth, respectively. As a result, we obtained 33 candidates, whose Cs-accumulating abilities were over 2-fold higher than that of the wild-type, from the population of mutagenized cells.

論文

Accumulation of cesium and its association with intracellular polyphosphate in ${it Deinococcus radiodurans}$

上田 涼史郎*; 佐藤 勝也; 林 秀謙*; 鳴海 一成*; 大野 豊

JAEA-Review 2015-022, JAEA Takasaki Annual Report 2014, P. 101, 2016/02

It is suggesting a possibility that the polyphosphate granules in cells could accumulate harmful metal ions such as radiocesium to protect the cells from cytotoxicity. In this study, we investigated the accumulation of cesium and its association with intracellular polyphosphate in ${it Deinococcus radiodurans}$ using the gene disruptant and overexpressing strains. The cesium accumulation level in the disruptant and overexpressing strains was compared with that of the wild-type strain. The cesium accumulations in the ${it ppk}$ disruptant and overexpressing strains were slightly decreased and almost same, respectively. Like the intracellular polyphosphate level, the cesium accumulation level was significantly increased in the ${it ppx}$ disruptant strain. These results suggested that the intracellular polyphosphate level was positively correlated with the cesium accumulation level in ${it D. radiodurans}$.

論文

Draft genome sequence of the radioresistant bacterium ${it Deinococcus grandis}$, isolated from freshwater fish in Japan

佐藤 勝也; 小野寺 威文*; 面曽 宏太*; 武田-矢野 喜代子*; 片山 豪*; 大野 豊; 鳴海 一成*

Genome Announcements (Internet), 4(1), p.e01631-15_1 - e01631-15_2, 2016/01

${it Deinococcus grandis}$ was isolated as a gram-negative, red-pigmented, radioresistant, rod-shaped bacterium from freshwater fish. The draft genome sequence of ${it D. grandis}$ ATCC 43672 was 4,092,497 bp, with an average G+C content of 67.5% and comprised of 4 circular contigs and 3 linear contigs that was deposited at DDBJ/EMBL/Genbank under the accession number BCMS00000000. The draft genome sequence indicates that ${it D. grandis}$ possesses a DNA damage response regulator (encoded by ${it pprI}$ homolog) and radiation-desiccation response regulons (${it recA}$, ${it ddrA}$, ${it ddrO}$, ${it pprA}$ and ${it gyrA}$ homologs etc.), which are involved in the unique radiation-desiccation response system in ${it D. radiodurans}$. As it is for ${it D. radiodurans}$, ${it D. grandis}$ seems to employ the same radioresistant mechanisms. In future, the draft genome sequence of ${it D. grandis}$ will be useful for elucidating the common principles of the radioresistance based on the extremely efficient DNA repair mechanisms in Deinococcus species by the comparative analysis of genomic sequences.

論文

${it Arabidopsis pab1}$, a mutant with reduced anthocyanins in immature seeds from $$banyuls$$, harbors a mutation in the MATE transporter $$FFT$$

北村 智; 大野 豊; 鳴海 一成*

Plant Molecular Biology, 90(1-2), p.7 - 18, 2016/01

 被引用回数:8 パーセンタイル:27.67(Biochemistry & Molecular Biology)

種皮にプロアントシアニジンに代わってアントシアニンを蓄積するシロイヌナズナ変異体banを用いて、イオンビームによる変異誘発を実施し、未熟種子におけるアントシアニン量の変化した変異体選抜を行うことで新規変異体pab1を得た。pab1はアントシアニン量が低下しているもののその合成酵素遺伝子の発現は正常であることがわかった。map-based cloningによってpab1変異体ではMATE型輸送体であるFFT遺伝子に変異が生じていることが明らかとなった。pab1変異体にFFT遺伝子を導入するとアントシアニン量が復帰したこと、FFT遺伝子が種皮のフラボノイド蓄積細胞層で発現していること、FFTタンパク質が液胞の膜に局在することから、FFTはアントシアニンの液胞蓄積のために液胞膜で機能していると考えられた。今回実施した変異体選抜を用いると、従来の変異体選抜では見つからないようなアントシアニン関連変異体が得られる可能性が示唆された。

論文

Draft genome sequence of ${it Methylobacterium}$ sp. ME121, isolated from soil as a mixed single colony with ${it Kaistia}$ sp. 32K

藤浪 俊*; 武田 喜代子*; 小野寺 威文*; 佐藤 勝也; 清水 哲*; 若林 佑*; 鳴海 一成*; 中村 顕*; 伊藤 政博*

Genome Announcements (Internet), 3(5), p.e01005-15_1 - e01005-15_2, 2015/09

${it Methylobacterium}$ sp. ME121 was isolated from soil as a mixed single colony with ${it Kaistia}$ sp. 32K during our screening of L-glucose-utilizing microorganisms, and its growth was enhanced by coculture. It was expected that genomic analysis of this bacterium would provide novel information on coculture-dependent growth enhancement. The genomic information of symbiotic bacteria could be of use for studying the molecular mechanisms underlying microbial symbiosis. The draft genome sequence of ${it Methylobacterium}$sp. ME121 is 7,096,979 bp in total length and comprises 197 large contigs ($$>$$ 500 bp) that was deposited at DDBJ/EMBL/GenBank under the accession number BBUX00000000. The draft genome sequence shows that ${it Methylobacterium}$ sp. ME121 has some genes that encode putative methanol/ethanol family PQQ-dependent dehydrogenases involved in methylotrophy. Some unknown factor provided by the coculture may contribute to increase the growth of ${it Methylobacterium}$ sp. ME121.

論文

イオンビーム育種技術による植物・微生物のセシウム吸収特性の改変

大野 豊; 長谷 純宏; 佐藤 勝也; 野澤 樹; 鳴海 一成*

放射線と産業, (138), p.17 - 20, 2015/06

イオンビーム育種技術は、近年植物や微生物の改変に活発に利用され、有用品種の作出に成果をあげている。東京電力福島第一原子力発電所の事故対応のためにイオンビーム育種でなにができるのか。日本原子力研究開発機構量子ビーム応用センターでは、高崎量子応用研究所のサイクロトロンを利用し、セシウム高・低吸収植物やセシウム高濃縮菌の開発に取り組み、変異体の候補を得ることに成功した。

論文

Isolation of carbon ions-induced mutants in ${{it Arabidopsis thaliana}}$ exhibiting different anthocyanin accumulation patterns

北村 智; 大野 豊; 鳴海 一成*

JAEA-Review 2014-050, JAEA Takasaki Annual Report 2013, P. 72, 2015/03

イオンビームは新しい変異体を誘発するのに効果的である。既存のイオンビーム変異体に対するイオンビーム再照射は、イオンビームがゲノムの一部のみを改変する特徴を活かした変異誘発法で、植物の形質を一歩ずつ改変していくのに有効である。今回、シロイヌナズナにおいてイオンビーム照射法を適用し、従来法では得られないような色素蓄積変異体の選抜を行った。

論文

Development of cesium-accumulating bacteria by ion beam breeding technology

佐藤 勝也; 上田 涼史郎; 長谷 純宏; 鳴海 一成*; 大野 豊

JAEA-Review 2014-050, JAEA Takasaki Annual Report 2013, P. 117, 2015/03

Bioremediation uses biological organisms to solve an environmental problem. However, microorganisms are sometimes not enough effective to achieve efficient recovery. Therefore, enhancing the functionality of microorganisms is needed to promote the active use of bioremediation. This study began with the purpose of development of Cs-accumulating bacteria by ion beam breeding technology. We investigated which members are better suited for Cs-accumulation in the genus ${it Deinococcus}$ (${it D. radiodurans}$, ${it D. grandis}$, ${it D. proteolyticus}$, ${it D. radiopugnans}$, ${it D. geothermalis}$ and ${it D. murrayi}$) exhibit extraordinary radioresistant. The Cs concentration in the ${it Deinococcus}$ cells exhibited over 4-fold compared with that of the ${it E. coli}$ by atomic absorption spectrometer analysis. This result suggested that ${it D. radiodurans}$ and ${it D. grandis}$ are best two suitable species for the development of high Cs-accumulating bacteria by ion beam breeding technology.

論文

Molecular analysis of polyphosphate biosynthesis-related genes in ${it Deinococcus radiodurans}$

上田 涼史郎; 佐藤 勝也; 林 秀謙*; 鳴海 一成*; 大野 豊

JAEA-Review 2014-050, JAEA Takasaki Annual Report 2013, P. 118, 2015/03

It is suggesting a possibility that the polyphosphate granules in cells could accumulate harmful metal ions such as radiocesium to protect the cells from cytotoxicity. ${it Deinococcus radiodurans}$ has been considered as a microorganism for bioremediation under highly radioactive contaminated environments. We generated the polyphosphate biosynthesis-related genes (${it ppk}$ and ${it ppx}$) disruptant strains and characterized their disruption effect to clarify the role of polyphosphate for the accumulation of cesium in ${it D. radiodurans}$. The intracellular levels of polyphosphates in the ${it ppk}$ disruptant and overexpressing strains were almost same and slightly increased, respectively. On other hand, the intracellular level of polyphosphates significantly increased in the ${it ppx}$ disruptant strains, suggesting that the ${it ppx}$gene plays an important role in the accumulation of polyphosphate in ${it D. radiodurans}$.

論文

Genome analysis of the high temperature tolerant mutant of ${it Bradyrhizobium japonicum}$ USDA110 generated by ion-beam irradiation

武田 喜代子*; 佐藤 勝也; 鳴海 一成*; 大野 豊; 大津 直子*; 横山 正*

JAEA-Review 2014-050, JAEA Takasaki Annual Report 2013, P. 120, 2015/03

Major constraint of biofertilizer utilization in agricultural practice is a viability loss of beneficial microorganisms in biofertilizers by high temperature stress during both storage and transportation. Therefore, we have tried to improve high temperature tolerance of ${it Bradyrhizobium japonicum}$ USDA110 using carbon-ion beams accelerated by an AVF cyclotron at TIARA. Consequently, we have obtained a high temperature tolerant mutant that can maintain a high survival rate at 42 $$^{circ}$$C for at least 7 days in yeast-mannitol broth medium, and named it as M14. The genome sequence of M14 was determined by a whole-genome shotgun strategy. The acquirement of high temperature tolerance in M14 could be attributed to 18 base mutations and/or a large-scale inversion (1.27 Mb). In the future, we will conduct further analysis on these mutations in order to decipher the molecular mechanisms of high temperature tolerance of M14.

論文

Thermotolerant mutants of entomopathogenic fungi obtained by ion beam- and $$gamma$$ ray-induced mutagenesis

西東 力*; Fitriana, Y.*; 佐藤 勝也; 大野 豊; 鳴海 一成*

JAEA-Review 2014-050, JAEA Takasaki Annual Report 2013, P. 121, 2015/03

Entomopathogenic fungi, such as ${it Metarhizium anisopliae}$, are economically important agents for integrated pest management programmes of insect pests. However, negative effects of temperature, such as heat stress that exceeds 35 $$^{circ}$$C, can have serious deleterious effects on conidial germination and persistence, vegetative growth, sporulation, and/or the infection process in these entomopathogenic fungi. In this study, we attempted to enhance thermotolerance in a model entomopathogenic fungus, ${it M. anisopliae}$, by mutagenesis using ion beams or $$gamma$$ rays and evaluated the relative virulence of the resulting mutants compared with the wild-type isolates. In conclusion, ion beams and $$gamma$$ rays are useful tools for improving biological characteristics, such as thermotolerance and fungicide-tolerance, in entomopathogenic fungi, but the mutants obtained in this study must be carefully evaluated for unpredictable negative side effects.

論文

Role of DNA repair and effect of herbal extract on LOH induced by ion beam radiations in ${it Saccharomyces cerevisiae}$

布柴 達男*; 山内 理子*; 岩田 梨佳*; 佐藤 勝也; 大野 豊; 鳴海 一成*

JAEA-Review 2014-050, JAEA Takasaki Annual Report 2013, P. 124, 2015/03

The LOH (loss of heterozygosity) induction by various ion beam radiations was investigated in diploid ${it Saccharomyces cerevisiae}$. The ion beams $$^{12}$$C$$^{5+}$$, $$^{12}$$C$$^{6+}$$, $$^{4}$$He$$^{2+}$$ and $$^{40}$$Ar$$^{13+}$$ at the lowest dose of 37.5 Gy, which had $$sim$$10% lethality, induced LOH with $$sim$$5$-fold higher frequency, with dose-dependent manner until 150 Gy. The LOH induction was predominantly ($sim$$90%) resulted from homologous recombination. Pol$$zeta$$ may be involved in the induction of LOH by ion beam radiations, because in the strain lacking Rev3, only slight induction of LOH was observed, whereas no remarkable effect of deletion for Pol$$zeta$$ was observed on ion beam induced LOH. In addition, the effects of ${it Eucalyptus regnans}$ extracts were examined on iron-beam radiation induced LOH, and ethanol extract of ${it Eucalyptus regnans}$ possessed inhibitory effect on LOH induced by $$^{12}$$C$$^{6+}$$, $$^{4}$$He$$^{2+}$$ and $$^{40}$$Ar$$^{13+}$$ion-beam radiation.

論文

Benomyl-resistant ${it Beauveria bassiana}$ (Hypocreales: Clavicipitaceae) mutants induced by ion beams

Fitriana, Y.*; 篠原 忍*; 佐藤 勝也; 鳴海 一成*; 西東 力*

Applied Entomology and Zoology, 50(1), p.123 - 129, 2015/02

 パーセンタイル:100(Chemistry, Physical)

Fungicides targeted at phytopathogens can be harmful to entomopathogenic fungi. The purpose of this study was to use ion-beam irradiation technology to produce benomyl-resistant mutants of the entomopathogenic fungus ${it Beauveria bassiana}$. After irradiation of conidia at 150 Gy, two mutant isolates, BB22 and BB24, were selected on media containing the fungicide. In an assay of vegetative growth, BB22 and BB24 were over 500 and 800 times more tolerant to benomyl, respectively, compared with the wild isolate. A mutation was found at position 198 of the $$beta$$-tubulin gene in the mutant isolates, with a substitution of glutamate for alanine (E198A). Ion beams have great potential as a tool to improve the traits of entomopathogenic fungi such as increasing tolerance to fungicides. Fungicide-resistant mutants produced in this way could be useful agents for biological control within IPM programmes where fungicides are also used.

論文

Metabolic alterations in leaves of oxalate-rich plant ${it Rumex obtusifolius}$ L. irradiated by $$gamma$$ rays

北野 沙也佳*; 宮城 敦子*; 大野 豊; 長谷 純宏; 鳴海 一成*; 山口 雅利*; 内宮 博文*; 川合 真紀*

Metabolomics, 11(1), p.134 - 142, 2015/02

 被引用回数:4 パーセンタイル:72.47(Endocrinology & Metabolism)

We investigated on the metabolic effectiveness of $$gamma$$ ray irradiation to seeds of ${it Rumex obtusifolius}$, which are known to accumulate high level of soluble oxalate, to lower oxalate accumulation in leaves. Reduced rate of germination, discoloration of cotyledons, and deformed true leaves were observed by elevated irradiation. Metabolome analysis of primary metabolites using capillary electrophoresis-mass spectrometry showed a decrease in oxalate contents in the leaves of plants from $$gamma$$ ray-irradiated seeds. Moreover, organic acids such as malate and 2-oxoglutarate also decreased, whereas amino acids such as glutamate and glutamine increased. These results indicated that seed irradiation by $$gamma$$ rays leads to dynamic changes in metabolic pathways as well as plant growth/development.

論文

Draft genome sequence of ${it Bacillus alcalophilus}$ AV1934, a classic alkaliphile isolated from human feces in 1934

Attie, O.*; Jayaprakash, A.*; Shah, H.*; Paulsen, I. T.*; 守野 正人*; 高橋 優嘉*; 鳴海 一成*; Sachidanandam, R.*; 佐藤 勝也; 伊藤 政博*; et al.

Genome Announcements (Internet), 2(6), p.e01175-14_1 - e01175-14_2, 2014/11

The alkaliphilic bacterium ${it Bacillus alcalophilus}$ AV1934, isolated from human feces, recent has indicated novel potassium ion coupling to motility in this extremophile. We report draft sequences that will facilitate an examination of whether that coupling is part of a larger cycle of potassium ion-coupled transporters. This draft genome has 4,095 predicted genes and 4,063 predicted proteins. The whole-genome shotgun project has been deposited in Genbank under the accession no. ALPT02000000. In addition to the ${it mot}$ genes encoding the MotPS channel that uses either potassium or sodium coupling, two loci encoding multisubunit Mrp-type antiporters, were found. The ${it B. alcalophilus}$ AV1934 genome reveals no tripartite ATP-independent (TRAP-T family) uptake systems,a major sodium-coupled complement in the other two alkaliphiles. The ${it B. alcalophilus}$ AV1934 genomic data will enable us to test the hypothesis of a major role of potassium in ion coupling in this extremophile.

論文

Interaction of double-stranded DNA with polymerized PprA protein from ${it Deinococcus radiodurans}$

安達 基泰; 平山 裕士; 清水 瑠美; 佐藤 勝也; 鳴海 一成*; 黒木 良太

Protein Science, 23(10), p.1349 - 1358, 2014/10

 被引用回数:7 パーセンタイル:63.85(Biochemistry & Molecular Biology)

DNAの修復に関与する多機能なPprAは、デイノコッカスラジオデュランスの高度な放射線耐性を促進する重要なタンパク質である。PprAによる放射線耐性機構を解明するために、大腸菌で発現した組換え型PprAと2本鎖DNAとの相互作用解析を実施した。ゲルシフトアッセイにより、PprAとスーパーコイル型のpUC19 DNAとの複合体のゲルシフトが2極性であり、それが1mMのMg, Ca, Srイオンで促進されることが示された。シフトしたバンドの相対的な割合からPprAとスーパーコイル型のpUC19 DNAとの複合体形成のヒル係数および解離定数を計算したところ、1mM Mgイオンの存在下で、それぞれ3.3と0.6$$mu$$Mであった。このことは、PprAがスーパーコイル型のpUC19 DNAに少なくとも281分子結合していることを示しており、ゲル濾過での分離後にUV吸収で見積もられた値と一致した。この結果は、PprAが2本鎖DNAに沿って、直鎖状に重合して結合していることを示唆している。一方で、直鎖状の2本鎖DNAとニックがある環状DNAのバンドシフトに関しては、飽和が見られず、1.3$$mu$$M以上のPprA濃度の時、さらに大きな複合体の形成が確認された。この結果は、DNAに結合したPprAが、ダメージを受けたDNAの末端の会合を濃度依存的に促進していることを示している。

論文

Draft genome sequence of calcium-dependent ${it Paenibacillus}$ sp. strain TCA20, isolated from a hot spring containing a high concentration of calcium ions

藤浪 俊*; 武田 喜代子*; 小野寺 威文*; 佐藤 勝也; 佐野 元彦*; 高橋 優嘉*; 鳴海 一成*; 伊藤 政博*

Genome Announcements (Internet), 2(5), p.e00866-14_1 - e00866-14_2, 2014/09

Calcium-dependent ${it Paenibacillus}$ sp. strain TCA20 was isolated from a water sample of a hot spring containing a high concentration of calcium ions. Here, we report the draft genome sequence of this bacterium, which may be the basis for the research of calcium ion homeostasis. The draft genome sequence of ${it Paenibacillus}$ sp. strain TCA20 totals 5,631,463 bp in length and is composed of 33 large contigs ($$>$$ 500 bp) that was deposited at DDBJ/EMBL/Genbank under the accession no. BBIW00000000. The annotation of the draft genome sequence shows that ${it Paenibacillus}$ sp. strain TCA20 has a calcium-specific calcium/proton antiporter, ChaA, and a P-type calcium-transporting ATPase, YloB, was identified as calcium transporters. These transporters may also be important for the growth of ${it Paenibacillus}$ sp. strain TCA20 under the high concentration of calcium ions.

論文

Ion-beam and $$gamma$$-ray irradiations induce thermotolerant mutants in the entomopathogenic fungus ${it Metarhizium anisopliae s.l.}$

Fitriana, Y.*; 佐藤 勝也; 鳴海 一成*; 西東 力*

Biocontrol Science and Technology, 24(9), p.1052 - 1061, 2014/06

 被引用回数:1 パーセンタイル:89.08(Biotechnology & Applied Microbiology)

Entomopathogenic fungi, such as ${it Metarhizium anisopliae}$ Sorokin, are important agents for the biological control of insect pests. However, these fungi are not compatible with high temperatures. In this study, mutagenesis using ion beams or $$gamma$$ rays was used to generate five potentially thermotolerant mutants from two wild-type isolates of ${it M. anisopliae}$ (two using ion beams and three using $$gamma$$ rays). The mutant isolates had a higher upper thermal limit for vegetative growth compared to the wild types (by 2-3$$^{circ}$$C) and enhanced tolerance to wet-heat stress of 45$$^{circ}$$C for conidial germination. These results indicate that ion beams and $$gamma$$ rays are useful tools for improving biological characteristics, such as thermotolerance, in entomopathogenic fungi, but that mutants must be carefully evaluated for unpredictable negative side effects.

論文

Draft genome sequence of potassium-dependent alkaliphilic ${it Bacillus}$ sp. strain TS-2, isolated from a jumping spider

藤浪 俊*; 武田 喜代子*; 小野寺 威文*; 佐藤 勝也; 佐野 元彦*; 鳴海 一成*; 伊藤 政博*

Genome Announcements (Internet), 2(3), p.e00458-14_1 - e00458-14_2, 2014/05

The potassium-dependent alkaliphilic ${it Bacillus}$ sp. strain TS-2 was isolated from the mashed extract of a jumping spider, and its draft genome sequence was obtained. Comparative genomic analysis with a previously sequenced sodium-dependent alkaliphilic ${it Bacillus}$ species may reveal potassium-dependent alkaline adaptation mechanisms. The draft genome sequence of ${it Bacillus}$ sp. strain TS-2 was 4,360,646 bp in total length, comprised 58 large contigs ($$>$$ 500 bp) that was deposited at DDBJ/EMBL/Genbank under the accession number BAWL00000000. In the draft genome sequence of ${it Bacillus}$ sp. strain TS-2, two sets of ${it mrp}$ genes, which encode multisubunit secondary cation/proton antiporter-3 family proteins, were annotated. The Mrp complex acts as an Na$$^{+}$$/H$$^{+}$$ antiporter in typical alkaliphilic ${it Bacillus}$ species and plays a critical role in the sodium-dependent alkaline adaptation mechanism.

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