The role of the MCM2-7 helicase complex during Arabidopsis seed development
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The MINICHROMOSOME MAINTENANCE 2-7 (MCM2-7) complex, a ring-shaped heterohexamer, unwinds the DNA double helix ahead of the other replication machinery. Although there is evidence that individual components might have other roles, the essential nature of the MCM2-7 complex in DNA replication has made it difficult to uncover these. Here, we present a detailed analysis of Arabidopsis thaliana mcm2-7 mutants and reveal phenotypic differences. The MCM2-7 genes are coordinately expressed during development, although MCM7 is expressed at a higher level in the egg cell. Consistent with a role in the egg cell, heterozygous mcm7 mutants resulted in frequent ovule abortion, a phenotype that does not occur in other mcm mutants. All mutants showed a maternal effect, whereby seeds inheriting a maternal mutant allele occasionally aborted later in seed development with defects in embryo patterning, endosperm nuclear size, and cellularization, a phenotype that is variable between subunit mutants. We provide evidence that this maternal effect is due to the necessity of a maternal store of MCM protein in the central cell that is sufficient for maintaining seed viability and size in the absence of de novo MCM transcription. Reducing MCM levels using endosperm-specific RNAi constructs resulted in the up-regulation of DNA repair transcripts, consistent with the current hypothesis that excess MCM2-7 complexes are loaded during G1 phase, and are required during S phase to overcome replicative stress or DNA damage. Overall, this study demonstrates the importance of the MCM2-7 subunits during seed development and suggests that there are functional differences between the subunits.
KeywordsDNA replication DNA helicase DNA repair DNA damage Seed development Arabidopsis
We thank Jane Campbell and Robyn Lee for technical assistance. We would also like to thank Assoc. Prof. Peter Dearden for assistance with microscopy and Dr. Lynette Brownfield for advice on pollen staining. R.P.H. was funded by a Top Achiever Doctoral Scholarship from the Tertiary Education Commission of New Zealand. R.C.D. and R.C.M. were funded by the Marsden Fund of New Zealand.
- Boisnard-Lorig C, Colon-Carmona A, Bauch W, Hodge S, Doerner P, Bancharel E, Dumas C, Haseloff J, Berger F (2001) Dynamic analyses of the expression of the HISTONE: YFP fusion protein in arabidopsis show that syncytial endosperm is divided in mitotic domains. Plant Cell 13:495–509PubMedCentralPubMedGoogle Scholar
- Curtis MD, Grossniklaus U (2003) A gateway cloning vector set for high-throughput functional analysis of genes in planta. Plant Physiol 133:462–469 Google Scholar
- Hisanaga T, Ferjani A, Horiguchi G, Ishikawa N, Fujikura U, Kubo M, Demura T, Fukuda H, Ishida T, Sugimoto K (2013) The ATM-dependent DNA damage response acts as an upstream trigger for compensation in the fas1 mutation during Arabidopsis leaf development. Plant Physiol 162:831–841PubMedCentralPubMedGoogle Scholar
- Le BH, Cheng C, Bui AQ, Wagmaister JA, Henry KF, Pelletier J, Kwong L, Belmonte M, Kirkbride R, Horvath S, Drews GN, Fischer RL, Okamuro JK, Harada JJ, Goldberg RB (2010) Global analysis of gene activity during Arabidopsis seed development and identification of seed-specific transcription factors. Proc Natl Acad Sci USA 107:8063–8070PubMedCentralPubMedGoogle Scholar
- Yi D, Alvim Kamei CL, Cools T, Vanderauwera S, Takahashi N, Okushima Y, Eekhout T, Yoshiyama KO, Larkin J, Van den Daele H, Conklin P, Britt A, Umeda M, De Veylder L (2014) The Arabidopsis SIAMESE-RELATED cyclin-dependent kinase inhibitors SMR5 and SMR7 regulate the DNA damage checkpoint in response to reactive oxygen species. Plant Cell 26:296–309PubMedGoogle Scholar