Functional characterization of chaperonin containing T-complex polypeptide-1 and its conserved and novel substrates in Arabidopsis

Hee Kyung Ahn; Joong Tak Yoon; Ilyeong Choi; Sumin Kim; Ho Seok Lee; Hyun Sook Pai

doi:10.1093/jxb/erz099

Functional characterization of chaperonin containing T-complex polypeptide-1 and its conserved and novel substrates in Arabidopsis

Hee Kyung Ahn, Joong Tak Yoon, Ilyeong Choi, Sumin Kim, Ho Seok Lee, Hyun Sook Pai

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Abstract

Chaperonin containing T-complex polypeptide-1 (CCT) is an evolutionarily conserved chaperonin multi-subunit complex that mediates protein folding in eukaryotes. It is essential for cell growth and survival in yeast and mammals, with diverse substrate proteins. However, only a few studies on plant CCT have been reported to date, due to the essentiality of CCT subunit genes and the large size of the complex. Here, we have investigated the structure and function of the Arabidopsis CCT complex in detail. The plant CCT consisted of eight subunits that assemble to form a high-molecular-mass protein complex, shown by diverse methods. CCT-deficient cells exhibited depletion of cortical microtubules, accompanied by a reduction in cellular α- and β-tubulin levels due to protein degradation. Cycloheximide-chase assays suggested that CCT is involved in the folding of tubulins in plants. Furthermore, CCT interacted with PPX1, the catalytic subunit of protein phosphatase 4, and may participate in the folding of PPX1 as its substrate. CCT also interacted with Tap46, a regulatory subunit of PP2A family phosphatases, but Tap46 appeared to function in PPX1 stabilization, rather than as a CCT substrate. Collectively, our findings reveal the essential functions of CCT chaperonin in plants and its conserved and novel substrates.

Original language	English
Pages (from-to)	2741-2757
Number of pages	17
Journal	Journal of experimental botany
Volume	70
Issue number	10
DOIs	https://doi.org/10.1093/jxb/erz099
Publication status	Published - 2019 May

Bibliographical note

Funding Information:
The authors wish to thank Dr Jen Sheen (Harvard Medical School, USA) for providing seeds of the estradiol-inducible TOR RNAi lines. This research was supported by the Cooperative Research Program for Agriculture Science & Technology Development [Project numbers PJ013212 (PMBC) and PJ013227 (SSAC)] from the Rural Development Administration, and Mid-Career Researcher Program (NRF-2016R1A2B4013180) from the National Research Foundation (NRF) of the Republic of Korea. H-KA was funded by the Global PhD fellowship (2011-0031010).Conflict of interestThe authors declare no conflict of interest.

Funding Information:
The authors wish to thank Dr Jen Sheen (Harvard Medical School, USA) for providing seeds of the estradiol-inducible TOR RNAi lines. This research was supported by the Cooperative Research Program for Agriculture Science & Technology Development [Project numbers PJ013212 (PMBC) and PJ013227 (SSAC)] from the Rural Development Administration, and Mid-Career Researcher Program (NRF- 2016R1A2B4013180) from the National Research Foundation (NRF) of the Republic of Korea. H-KA was funded by the Global PhD fellowship (2011-0031010). Conflict of interest The authors declare no conflict of interest.

Publisher Copyright:
© 2019 The Author(s) 2019. Published by Oxford University Press on behalf of the Society for Experimental Biology.

All Science Journal Classification (ASJC) codes

Physiology
Plant Science

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10.1093/jxb/erz099

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title = "Functional characterization of chaperonin containing T-complex polypeptide-1 and its conserved and novel substrates in Arabidopsis",

abstract = "Chaperonin containing T-complex polypeptide-1 (CCT) is an evolutionarily conserved chaperonin multi-subunit complex that mediates protein folding in eukaryotes. It is essential for cell growth and survival in yeast and mammals, with diverse substrate proteins. However, only a few studies on plant CCT have been reported to date, due to the essentiality of CCT subunit genes and the large size of the complex. Here, we have investigated the structure and function of the Arabidopsis CCT complex in detail. The plant CCT consisted of eight subunits that assemble to form a high-molecular-mass protein complex, shown by diverse methods. CCT-deficient cells exhibited depletion of cortical microtubules, accompanied by a reduction in cellular α- and β-tubulin levels due to protein degradation. Cycloheximide-chase assays suggested that CCT is involved in the folding of tubulins in plants. Furthermore, CCT interacted with PPX1, the catalytic subunit of protein phosphatase 4, and may participate in the folding of PPX1 as its substrate. CCT also interacted with Tap46, a regulatory subunit of PP2A family phosphatases, but Tap46 appeared to function in PPX1 stabilization, rather than as a CCT substrate. Collectively, our findings reveal the essential functions of CCT chaperonin in plants and its conserved and novel substrates.",

author = "Ahn, {Hee Kyung} and Yoon, {Joong Tak} and Ilyeong Choi and Sumin Kim and Lee, {Ho Seok} and Pai, {Hyun Sook}",

note = "Funding Information: The authors wish to thank Dr Jen Sheen (Harvard Medical School, USA) for providing seeds of the estradiol-inducible TOR RNAi lines. This research was supported by the Cooperative Research Program for Agriculture Science & Technology Development [Project numbers PJ013212 (PMBC) and PJ013227 (SSAC)] from the Rural Development Administration, and Mid-Career Researcher Program (NRF-2016R1A2B4013180) from the National Research Foundation (NRF) of the Republic of Korea. H-KA was funded by the Global PhD fellowship (2011-0031010).Conflict of interestThe authors declare no conflict of interest. Funding Information: The authors wish to thank Dr Jen Sheen (Harvard Medical School, USA) for providing seeds of the estradiol-inducible TOR RNAi lines. This research was supported by the Cooperative Research Program for Agriculture Science & Technology Development [Project numbers PJ013212 (PMBC) and PJ013227 (SSAC)] from the Rural Development Administration, and Mid-Career Researcher Program (NRF- 2016R1A2B4013180) from the National Research Foundation (NRF) of the Republic of Korea. H-KA was funded by the Global PhD fellowship (2011-0031010). Conflict of interest The authors declare no conflict of interest. Publisher Copyright: {\textcopyright} 2019 The Author(s) 2019. Published by Oxford University Press on behalf of the Society for Experimental Biology.",

year = "2019",

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T1 - Functional characterization of chaperonin containing T-complex polypeptide-1 and its conserved and novel substrates in Arabidopsis

AU - Ahn, Hee Kyung

AU - Yoon, Joong Tak

AU - Choi, Ilyeong

AU - Kim, Sumin

AU - Lee, Ho Seok

AU - Pai, Hyun Sook

N1 - Funding Information: The authors wish to thank Dr Jen Sheen (Harvard Medical School, USA) for providing seeds of the estradiol-inducible TOR RNAi lines. This research was supported by the Cooperative Research Program for Agriculture Science & Technology Development [Project numbers PJ013212 (PMBC) and PJ013227 (SSAC)] from the Rural Development Administration, and Mid-Career Researcher Program (NRF-2016R1A2B4013180) from the National Research Foundation (NRF) of the Republic of Korea. H-KA was funded by the Global PhD fellowship (2011-0031010).Conflict of interestThe authors declare no conflict of interest. Funding Information: The authors wish to thank Dr Jen Sheen (Harvard Medical School, USA) for providing seeds of the estradiol-inducible TOR RNAi lines. This research was supported by the Cooperative Research Program for Agriculture Science & Technology Development [Project numbers PJ013212 (PMBC) and PJ013227 (SSAC)] from the Rural Development Administration, and Mid-Career Researcher Program (NRF- 2016R1A2B4013180) from the National Research Foundation (NRF) of the Republic of Korea. H-KA was funded by the Global PhD fellowship (2011-0031010). Conflict of interest The authors declare no conflict of interest. Publisher Copyright: © 2019 The Author(s) 2019. Published by Oxford University Press on behalf of the Society for Experimental Biology.

PY - 2019/5

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N2 - Chaperonin containing T-complex polypeptide-1 (CCT) is an evolutionarily conserved chaperonin multi-subunit complex that mediates protein folding in eukaryotes. It is essential for cell growth and survival in yeast and mammals, with diverse substrate proteins. However, only a few studies on plant CCT have been reported to date, due to the essentiality of CCT subunit genes and the large size of the complex. Here, we have investigated the structure and function of the Arabidopsis CCT complex in detail. The plant CCT consisted of eight subunits that assemble to form a high-molecular-mass protein complex, shown by diverse methods. CCT-deficient cells exhibited depletion of cortical microtubules, accompanied by a reduction in cellular α- and β-tubulin levels due to protein degradation. Cycloheximide-chase assays suggested that CCT is involved in the folding of tubulins in plants. Furthermore, CCT interacted with PPX1, the catalytic subunit of protein phosphatase 4, and may participate in the folding of PPX1 as its substrate. CCT also interacted with Tap46, a regulatory subunit of PP2A family phosphatases, but Tap46 appeared to function in PPX1 stabilization, rather than as a CCT substrate. Collectively, our findings reveal the essential functions of CCT chaperonin in plants and its conserved and novel substrates.

AB - Chaperonin containing T-complex polypeptide-1 (CCT) is an evolutionarily conserved chaperonin multi-subunit complex that mediates protein folding in eukaryotes. It is essential for cell growth and survival in yeast and mammals, with diverse substrate proteins. However, only a few studies on plant CCT have been reported to date, due to the essentiality of CCT subunit genes and the large size of the complex. Here, we have investigated the structure and function of the Arabidopsis CCT complex in detail. The plant CCT consisted of eight subunits that assemble to form a high-molecular-mass protein complex, shown by diverse methods. CCT-deficient cells exhibited depletion of cortical microtubules, accompanied by a reduction in cellular α- and β-tubulin levels due to protein degradation. Cycloheximide-chase assays suggested that CCT is involved in the folding of tubulins in plants. Furthermore, CCT interacted with PPX1, the catalytic subunit of protein phosphatase 4, and may participate in the folding of PPX1 as its substrate. CCT also interacted with Tap46, a regulatory subunit of PP2A family phosphatases, but Tap46 appeared to function in PPX1 stabilization, rather than as a CCT substrate. Collectively, our findings reveal the essential functions of CCT chaperonin in plants and its conserved and novel substrates.

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Functional characterization of chaperonin containing T-complex polypeptide-1 and its conserved and novel substrates in Arabidopsis

Abstract

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