Monday, 9th December 2019

DNA replication, chromatin and cell division



Crisanto Gutierrez






Research summary

Fig. 1. Identification of DNA replication origins (ORIs) in whole developing organisms: 4 and 10 day-old Arabidopsis thaliana seedlings (left upper panel). The procedure includes the purification of single-stranded DNA of short nascent strands (SNS), conversion to double-stranded DNA and sequencing (right upper panel). Peak identification in several experimental situations allows the genome-wide mapping of ORIs (red in lower panel).  
Fig. 2. Histone H3 dynamics. The canonical H3.1 (green), incorporated in every S-phase, but not the H3.3 variant (red), incorporated in a cell cycle-independent manner, is almost completely evicted in mitotic cells (red mitosis) undergoing their last division before entering the differentiation zone. Mitotic cells in previous cell divisions (yellow mitosis) contain both H3.1 and H3.3.

The transition to multicellularity required the acquisition of novel structures and mechanisms to coordinate cell division, acquisition of cell fates and the differentiation, and the establishment of complex regulatory networks. Our group is interested in understanding the mechanisms that control these processes and how epigenetic mechanisms affect such coordination.

To that end, we use the model plant Arabidopsis thaliana that offers us the possibility of carrying out molecular, cellular, genetic and genomic approaches. In addition, plant development, contrary to the situation in animals, is post-embryonic and occurs during the entire life of the organism. Our research is aimed at understanding fundamental questions on cell proliferation control, cellular homeostasis and genome replication in multicellular organisms.

We have developed genomic strategies to study the functional properties and molecular determinants of DNA replication origins (ORIs) in all cell types of the whole organism to determine the influence of hormonal conditions, developmental signals and the environment (Fig. 1). We found that ORI activity is compatible with multiple signatures although most of them tend to associate with chromatin states present in proximal promoters, TSS and 5’-end of genes. In addition, ORIs are enriched in the tandem arrays of GGN trinucleotides, which can for G4 structures. Our experimental approach is opening a new avenue to use mutants in the analysis of genome replication. We are combining the study of molecular determinants of ORIs with detailed analysis of the pre-RC dynamics during organ development.

Cell proliferation is crucial for organogenesis, which is determined by a strict control of gene expression patterns. We study chromatin dynamics along the cell cycle with special emphasis in two aspects: one, the regulation of cell proliferation potential, very related to the control of gene expression in G1 and G2, and the exit to differentiation, and another, related to the specific chromatin modifications in response to stress. The balance between the canonical histone H3.1 and the variant H3.3 serves to identify the cell population undergoing their last cell cycle before exit to differentiation because most of H3.1 is massively evicted in the last G2 phase (Fig. 2), which is longer in these cells.






Sequeira-Mendes, J., Vergara, Z., Peiro, R., Morata, J., Aragüez, I., Costas, C., Mendez-Giraldez, R., Casacuberta, J.M., Bastolla, U., Gutierrez, C. (2019). Differences in firing efficiency, chromatin and transcription underlie the developmental plasticity of Arabidopsis DNA replication origins. Genome Res. (in press). 


Dvorackova, M., Raposo, B., Matula, P., Fuchs, J., Schubert, V., Peska, V., Desvoyes, B., Gutierrez, C., Fajkus, J. (2018)  Replication of Ribosomal RNA Genes in Arabidopsis thaliana Occurs both inside and outside of the Nucleolus during S-Phase Progression. J. Cell Sci. 131, jcs202416, doi: 10.1242/jcs.202416). 


Desvoyes, B.*, Vergara, Z.*, Sequeira-Mendes, J., Madeira, S., Gutierrez, C. (2017) A rapid and efficient ChIP protocol to profile chromatin binding proteins and epigenetic modifications in bulk Arabidopsis tissue. Methods Mol. Biol. 1675, 71-81. * shared 1st coauthorship.  

Desvoyes, B.*,+, Sequeira-Mendes, J.*, Vergara, Z., Madeira, S., Gutierrez, C+. (2017) Sequential ChIP protocol for profiling bivalent epigenetic modifications in the same chromatin fiber (ReChIP). Methods Mol. Biol. 1675, 83-97.  * shared 1st coauthorship, + Co-corespondent authors

Vergara, Z., Sequeira-Mendes, J., Morata, J., Hénaff, E., Peiró, R., Costas, C., Casacuberta, J.M., Gutierrez, C. (2017) Retrotransposons are specified as DNA replication origins in the gene-poor regions of Arabidopsis heterochromatin. Nucleic Acids Res. 45, 8358-8368.

Fernandez-Marcos, M., Desvoyes, B., Manzano, C., Liberman, L.M., Benfey, P.N., del Pozo, J.C., Gutierrez, C. (2017) Control of Arabidopsis lateral root boundaries by MYB36. New Phytol. 213, 105-112.

Vergara, Z., Gutierrez, C. (2017) Emerging roles of chromatin in the maintenance of genome organization and function in plants. Genome Biol. 18, 96.


Gutierrez, C., Desvoyes, B., Vergara, Z., Otero, S., Sequeira-Mendes, J. (2016) Links of genome replication, transcriptional silencing and chromatin dynamics. Curr. Opin. Plant. Biol. 34, 92-99.

Desvoyes, B., Gutierrez, C. (2016) A plant solution to the CDK conundrum in the DNA damage response. EMBO J. 35, 2061-2063. 

Gutierrez, C. (2016) 25 years of cell cycle research: what’s ahead?  Trends Plant Sci. 21, 823-833.

Sequeira-Mendes, J., Gutierrez, C. (2016) Genome architecture: from linear organisation of chromatin to 3D assembly in the nucleus. Chromosoma 125, 455-469.

Otero, S*., Desvoyes, B.*, Peiró, R., Gutierrez, C. (2016) Histone H3 dynamics uncovers domains with distinct proliferation potential in the Arabidopsis root. Plant Cell 28, 1361-1371 * 1st coauthors.              Plant Cell 28, 1235 (2016) In Brief- Hofmann, N. Last exit to differentiation: histone variants as signspots.

Mauri, N., Fernandez-Marcos, M., Costas, C., Desvoyes, B., Pichel, A., Caro, E., Gutierrez, C. (2016) GEM, a member of the GRAM domain family of proteins, is part of the ABA signaling pathway. Sci. Rep. 6, 22660.

García-Cruz, K., García-Ponce, B., Garay-Arroyo, A., Sanchez, M.P., Ugartechea-Chirino, Y., Desvoyes, B., Pacheco-Escobedo, M., Tapia-López, R., Ransom-Rodriguez, I., Gutierrez, C., Alvarez-Buylla, E. (2016) The MADS-box XAANTAL1 Increases proliferation at the Arabidopsis root stem-cell niche and participates in transition to differentiation by regulating cell cycle components. Ann. Bot.117, 787-796.

Havlová, K., Dvořáčková, M., Peiro, R, Abia, D., Mozgová, I., Vansáčová, L., Gutierrez, C., Fajkus, J. (2016) Variation of 45S rDNA intergenic spacers in Arabidopsis thaliana. Plant. Mol. Biol. 92, 457-471.


Gutierrez, C., Puchta H. (2015) Chromatin and Development: s Special issue. Editorial. Plant J. 83, 1-3.

Sequeira-Mendes, J., Gutierrez, C. (2015) Links between genome replication control and chromatin landscapes. Plant J. 83, 38-51.

Rodriguez-Mega, E., Pyñeiro-Nelson, A., Gutierrez, C., Garcia-Ponce, B., Sanchez, M.P., Zluhen-Martinez, E., Alvarez-Buylla, E.R., Garay-Arroyo, A. (2015) The role of transcriptional regulation in the evolution of plant phenotype. Dev. Dyn. 244, 1074–1095.


Desvoyes, D., Fernandez-Marcos, M., Sequeira-Mendes, J., Otero, S., Vergara, Z., Gutierrez, C. (2014) Looking at plant cell cycle from the chromatin window. Frontiers Plant Sci. 5, 369.

Sequeira-Mendes, J., Aragüez, I., Peiró, R., Zhang, X., Jacobsen, S.E., Bastolla, U., Gutierrez, C. (2014) The Functional Topography of the Arabidopsis Genome is organized in a reduced Number of linear Motifs of Chromatin states. Plant Cell 26, 2351-2366.

Otero, S., Desvoyes, B., Gutierrez, C. (2014) Histone H3 dynamics in plant cell cycle and development. Cytogenet. Genomic Res. 143, 114-124.

Edgar, B.A., Zielke, N., Gutierrez, C. (2014) Endocycles: a recurrent evolutionary innovation for post-mitotic cell growth. Nature Rev Cell Mol. Biol. 15, 197-210.

Coego, A., Brizuela, E., Castillejo, P., Ruiz, S., Koncz, C., del Pozo, J.C., Piñeiro, M., Jarillo, J.A., Paz-Ares, J., Leon J., and the TRANSPLANTA Consortium. (2014) The TRANSPLANTA Collection of Arabidopsis Lines: A resource for Functional Analysis of Transcription Factors based on their conditional Overexpression. Plant J. 77, 944-953.

Desvoyes, B., de Mendoza, A., Ruiz-Trillo, I., Gutierrez, C. (2014) Novel roles of plant RETINOBLASTOMA-RELATED (RBR) protein in cell proliferation and asymmetric cell division. J. Exp. Bot. 65, 2657-2666.

Hénaff, E., Vives, C., Desvoyes, B., Chaurasia, A., Payet, J., Gutierrez, C., Casacuberta, JM. (2014) Extensive amplification of the E2F transcription factor binding sites by transposons during evolution of Brassica species. Plant J. 77, 852-862.


Lario, L., Ramirez-Parra, E., Gutierrez, C., Spampinato, C., Casati, P. (2013) ANTI-SILENCING FUNCTION1 proteins are involved in ultraviolet-induced DNA damage repair and are cell cycle regulated by E2F transcription factors in Arabidopsis. Plant Physiol. 162, 1164-1177.

Triviño, M., Martín-Trillo, M., Ballesteros I., Delgado D., de Marcos, A., Desvoyes, B., Gutierrez, C., Mena, M., Fenoll, C. (2013) Timely expression of Arabidopsis stoma-fate master regulator MUTE is required for specification of other epidermal cell types. Plant J. 75, 808-822.

Gutierrez, C., Sequeira-Mendes, J., Aragüez, I. (2013) Replication of Plant genomes. In: The Plant Sciences: Molecular Biology. M. Tester, R. A. Jorgensen, Eds. Article ID: 349832. Springer.


Stroud, H.+, Otero, S. +, Desvoyes, B., Ramirez-Parra, E., Jacobsen, S.E., Gutierrez, C. Genome-wide analysis of histone H3.1 and H3.3 variants in Arabidopsis thaliana. Proc. Natl. Acad. Sci. USA 109, 5370-5375 (2012). + Co-first authors

Sanchez, M.P. +, Costas, C. +, Sequeira-Mendes, J. +, Gutierrez, C. DNA replication control in plants. (2012) Cold Spring Harb. Perspect. Biol. 4:a01014+ Co-first authors

Manzano, C., Ramirez-Parra, E., Casimiro, I., Otero, S., Desvoyes, B., De Rybel, B., Beeckman, T., Casero, P., Gutierrez, C., del Pozo, J.C. Auxin and epigenetic regulation of SKP2B, an F-box that represses lateral root formation. Plant Physiol. 160, 749-762 (2012).

Caro, E., Desvoyes, B., Gutierrez, C. (2012) GTL1 keeps cell growth and nuclear ploidy under control. EMBO J. 31, 4483-4485 (2012).


Costas, C.+, Sanchez, M.P.+, Stroud, H.+, Yu, Y., Oliveros, J.C., Feng, S., Benguria, A., López-Vidriero, I., Zhang, X., Solano, R., Jacobsen, S.E., Gutierrez, C. (2011) Genome-wide mapping of Arabidopsis origins of DNA replication and their associated epigenetic marks. Nat. Struct. Mol. Biol. 18, 395-400.   + Co-first authors

Lario, L., Ramirez-Parra, E., Gutierrez, C., Casati, P., Spampinato, C. (2011) Regulation of plant MSH2 and MSH6 genes in the UV-B induced DNA damage response. J. Exp. Bot. 62, 2925-2937.

Costas, C. +, Desvoyes, B. +, Gutierrez, C. (2011) A chromatin perspective of cell cycle progression. Biochim. Biophys. Acta 1809, 379-387. + Co-first authors

Costas, C. +, Sanchez, M.P. +, Sequeira-Mendes, J. +, Gutierrez, C. (2011) Progress in understanding DNA replication control. Plant Sci. 181, 203-209. + Co-first authors

Sanmartin, M., Sauer, M., Muñoz, A., Zouhar, J., Ordoñez, A., van de Ven, W.T.G., Caro, E., Sanchez, M.P., Raikhel, N., Gutierrez, C., Sanchez-Serrano, J.J., Rojo, E. (2011) A molecular switch for initiating cell differentiation in Arabidopsis. Curr. Biol. 21, 999-1008.


Desvoyes, B., Sanchez, M.P., Ramirez-Parra, E., Gutierrez, C. (2010) Impact of nucleosome dynamics and histone modifications on cell proliferation during Arabidopsis development. Heredity 105, 80-91.

Jacob, Y.*, Stroud, H.*, LeBlanc, C., Feng, S., Zhou, L., Caro, E., Hassel, C., Gutierrez, C., Michaels, S.D., Jacobsen, S.E. (2010) Two histone H3 lysine methyltransferases, ATXR5 and ATXR6, regulate DNA replication in heterochromatin. Nature 466, 987-991.
















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