Mps1 (Monopolar Spindle 1) Protein Inhibition Affects Cellular Growth and Pro-Embryogenic Masses Morphology in Embryogenic Cultures of Araucaria angustifolia (Araucariaceae)
April
Mps1 (Monopolar Spindle 1) Protein Inhibition Affects Cellular Growth and Pro- Embryogenic Masses Morphology in Embryogenic Cultures of Araucaria angustifolia (Araucariaceae)
Jackellinne C. Douétts-Peres 1 2
Marco Antônio L. Cruz 0 2
Ricardo S. Reis 2
Angelo S. Heringer 2
Eduardo A. G. de Oliveira 0 2
Paula M. Elbl 2
Eny I. S. Floh 2
Vanildo Silveira 2
Claudete Santa-Catarina 1 2
0 Laboratório de Biotecnologia Vegetal, Núcleo em Ecologia e Desenvolvimento Sócio-ambiental de Macaé, Universidade Federal do Rio de Janeiro , Macaé, Rio de Janeiro, Brazil, 3 Laboratório de Biotecnologia, CBB, UENF, Campos dos Goytacazes, Rio de Janeiro , Brazil , 4 Unidade de Biologia Integrativa , Setor de Proteômica, UENF, Campos dos Goytacazes, Rio de Janeiro , Brazil , 5 Laboratório de Biologia Celular de Plantas, Departamento de Botânica, Instituto de Biociências, Universidade de São Paulo , São Paulo, São Paulo , Brazil
1 Laboratório de Biologia Celular e Tecidual, Centro de Biociências e Biotecnologia (CBB), Universidade Estadual do Norte Fluminense Darcy Ribeiro (UENF) , Campos dos Goytacazes, Rio de Janeiro , Brazil
2 Editor: Michael Polymenis, Texas A&M University , UNITED STATES
Somatic embryogenesis has been shown to be an efficient tool for studying processes based on cell growth and development. The fine regulation of the cell cycle is essential for proper embryo formation during the process of somatic embryogenesis. The aims of the present work were to identify and perform a structural and functional characterization of Mps1 and to analyze the effects of the inhibition of this protein on cellular growth and proembryogenic mass (PEM) morphology in embryogenic cultures of A. angustifolia. A singlecopy Mps1 gene named AaMps1 was retrieved from the A. angustifolia transcriptome database, and through a mass spectrometry approach, AaMps1 was identified and quantified in embryogenic cultures. The Mps1 inhibitor SP600125 (10 μM) inhibited cellular growth and changed PEMs, and these effects were accompanied by a reduction in AaMps1 protein levels in embryogenic cultures. Our work has identified the Mps1 protein in a gymnosperm species for the first time, and we have shown that inhibiting Mps1 affects cellular growth and PEM differentiation during A. angustifolia somatic embryogenesis. These data will be useful for better understanding cell cycle control during somatic embryogenesis in plants.
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OPEN ACCESS
Data Availability Statement: All relevant data are
within the paper and its Supporting Information files.
Introduction
The transition from a somatic cell into a somatic embryo, during somatic embryogenesis, is a
complex event, consisting of the following crucial steps: induction, cell dedifferentiation, and
competence acquisition; multiplication, with intense cell division; maturation, which
determines fate; and the germination of somatic embryos [
1
].
During somatic embryo formation, the correct performance of the cell cycle is crucial, and
adequate levels of certain signaling molecules, such as polyamines, carbohydrates, and nitric
oxide (NO), are required [
2–4
]. The maturation induction of somatic embryogenic cultures with
maturation promoters, such as abscisic acid (ABA), or with osmotic agents, such as polyethylene
glycol (PEG) and maltose, induce cell growth inhibition, preventing division and promoting the
differentiation of cell cultures [
5–8
]. However, other compounds, such as auxins, NO, and
putrescine, promote cell division, thereby increasing growth and inhibiting cell differentiation into
somatic embryos [
4,6,7
]. Embryogenic suspension culture systems have been developed for
Araucaria angustifolia, and they have been shown to be efficient systems for studying the effects of
signaling molecules in gymnosperms [
4,9,10
]. Cell cycle regulation can be used as a tool for the
elucidation of metabolism-related events, and it involves signaling compounds that are important
for various processes in plant development [11], including somatic embryogenesis [
12
].
Cell division in eukaryotes is controlled by a complex mechanism that involves
cyclindependent kinases (CDKs) as key regulators [
13,14
]. One of these kinases is Mps1 (monopolar
spindle 1), which has been described in humans and is characterized as a cell cycle regulator
that is evolutionarily conserved in eukaryotes [15]. Mps1 is a dual-specificity protein kinase
that plays a critical role in monitoring the accuracy of chromosome segregation at the mitotic
checkpoint, and it is an important component of the spindle assembly checkpoint (SAC) [
16
].
Among chemical inhibitors, SP600125 acts on Jun N-terminal kinase (JNK) proteins in
humans [
17
] and has been valuable in validating the cellular functions of Mps1. In plants, a
protein was found that was highly similar to human Mps1 in terms of structural characteristics,
such as its catalytic site, and it was conserved relative to the Mps1 protein found i (...truncated)