Endoreduplication in Arabidopsis thaliana: control mechanisms and its effect on tolerance towards environmental stresses

Endoreduplication is a widespread phenomenon during which the DNA is re-replicated without mitosis phase. Many roles have been proposed for endoreduplication, including metabolism activity, plant growth or development and environmental adaptation. Endoreduplication responds to many different factors, such as hormones and environmental conditions, and is under the control of many cell cycle-related genes. Recently, the E2Fe/DEL1 gene was shown to be a negative regulator of the cell-cycle-to-endoreduplication transition. Thus the role of the members of the DEL family on endoreduplication was investigated. Overexpression of E2Ff/DEL3 led to an endoreduplication augment in leaves while E2Fd/DEL2 seemed to have no effect on DNA endoreplication. While typical E2Fs interplay and are able to activate atypical E2Fs, atypical E2Fs seem unable to activate or repress other E2Fs. Through their effect on cell cycle genes activity and expression, hormones can have an impact on endoreduplication levels. Phytosterols and brassinosteroids are synthesized from identical start compounds, and have distinct effects on plant physiological development. Both components are important for cell division, while only brassinosteroids are seemingly involved in the control of cell size. Fenpropimorph, a drug inhibiting the synthesis of phytosterols and brassinosteroids, was found to severely inhibit leaf expansion, cell expansion and endoreduplication. Interestingly, CDKB1;1.N161 plants presented decreased fenpropimorph sensitivity in terms of ploidy. Cold nights have shown to have an negative impact on endoreduplication extent, cell enlargement and leaf size. The mutants with low and high ploidy levels did not present dramatic differences in growth reduction compared to wild type plants, suggesting that endoreduplication in not conferring any advantage or disadvantage to plants towards cold temperatures conditions. Endoreduplication has been proposed to confer a protection towards DNA-damaging radiations. The E2Fe/DEL1KO mutant plants, displaying increased ploidy levels, performed better after a UV-B treatment, which was primarily due to their increased levels of PHR1, an UV-induced DNA-damage repair enzyme, and only marginally to their elevated ploidy levels. PHR1 was identified as a direct E2Fe/DEL1 target gene. By combining a control on DNA repair mechanisms through PHR1 repression and endoreduplication through CCS52A2 repression, E2Fe/DEL1 may be able to maintain cells in an undifferentiated state

New insights into the control of endoreduplication endoreduplication could be driven by organ growth in Arabidopsis leaves

Enormous progress has been achieved understanding the molecular mechanisms regulating endoreduplication. By contrast, how this process is coordinated with the cell cycle or cell expansion and contributes to overall growth in multicellular systems remains unclear. A holistic approach was used here to give insight into the functional links between endoreduplication, cell division, cell expansion, and whole growth in the Arabidopsis (Arabidopsis thaliana) leaf. Correlative analyses, quantitative genetics, and structural equation modeling were applied to a large data set issued from the multiscale phenotyping of 200 genotypes, including both genetically modified lines and recombinant inbred lines. All results support the conclusion that endoreduplication in leaf cells could be controlled by leaf growth itself. More generally, leaf growth could act as a "hub" that drives cell division, cell expansion, and endoreduplication in parallel. In many cases, this strategy allows compensations that stabilize leaf area even when one of the underlying cellular processes is limiting

Natural DNA transformation is functional in Lactococcus lactis ssp. cremoris KW2.

Lactococcus lactis is one of the most commonly used lactic acid bacteria in the dairy industry. Activation of competence for natural DNA transformation in this species would greatly improve the selection of novel strains with desired genetic traits. Here, we investigated the activation of natural transformation in L. lactis ssp. cremoris KW2, a strain of plant origin whose genome encodes the master competence regulator ComX and the complete set of proteins usually required for natural transformation. In the absence of knowledge about competence regulation in this species, we constitutively overproduced ComX in a reporter strain of late competence phase activation and showed, by transcriptomic analyses, a ComX-dependent induction of all key competence genes. We further demonstrated that natural DNA transformation is functional in this strain and requires the competence DNA uptake machinery. Since constitutive ComX overproduction is unstable, we alternatively expressed comX under the control of an endogenous xylose-inducible promoter. This regulated system was used to successfully inactivate the adaptor protein MecA and subunits of the Clp proteolytic complex, which were previously shown to be involved in ComX degradation in streptococci. In the presence of a low amount of ComX, the deletion of mecA, clpC, or clpP genes markedly increased the activation of the late competence phase and transformability. Altogether, our results report the functionality of natural DNA transformation in L. lactis and pave the way for the identification of signaling mechanisms that trigger the competence state in this species.IMPORTANCELactococcus lactis is a lactic acid bacterium of major importance, which is used as a starter species for milk fermentation, a host for heterologous protein production, and a delivery platform for therapeutic molecules. Here, we report the functionality of natural transformation in L. lactis ssp. cremoris KW2 by the overproduction of the master competence regulator ComX. The developed procedure enables a flexible approach to modify the chromosome with single point mutation, sequence insertion, or sequence replacement. These results represent an important step for the genetic engineering of L. lactis that will facilitate the design of strains optimized for industrial applications. This will also help to discover natural regulatory mechanisms controlling competence in the genus Lactococcus

The DOF transcription factor OBP1 is involved in cell cycle regulation in Arabidopsis thaliana

In contrast to animal growth, plant growth is largely post-embryonic. Therefore plants have developed new mechanisms to precisely regulate cell proliferation by means of internal and external stimuli whilst the general core cell cycle machinery is conserved between eukaryotes. In this work we demonstrate a role for the Arabidopsis thaliana DNA-binding-with-one-finger (DOF) transcription factor OBP1 in the control of cell division upon developmental signalling. Inducible overexpression of OBP1 resulted in a significant overrepresentation of cell cycle genes among the upregulated transcripts. Direct targets of OBP1, as verified by chromatin immunoprecipitation, include at least the core cell cycle gene CYCD3;3 and the replication-specific transcription factor gene AtDOF2;3. Consistent with our molecular data, short-term activation of OBP1 in cell cultures affected cell cycle re-entry, shortening the duration of the G(1) phase and the overall length of the cell cycle, whilst constitutive overexpression of OBP1 in plants influenced cell size and cell number, leading to a dwarfish phenotype. Expression during embryogenesis, germination and lateral root initiation suggests an important role for OBP1 in cell cycle re-entry, operating as a transcriptional regulator of key cell cycle genes. Our findings provide significant input into our understanding of how cell cycle activity is incorporated into plant growth and development

Systems-based analysis of Arabidopsis leaf growth reveals adaptation to water deficit

Leaves have a central role in plant energy capture and carbon conversion and therefore must continuously adapt their development to prevailing environmental conditions. To reveal the dynamic systems behaviour of leaf development, we profiled Arabidopsis leaf number six in depth at four different growth stages, at both the end-of-day and end-of-night, in plants growing in two controlled experimental conditions: short-day conditions with optimal soil water content and constant reduced soil water conditions. We found that the lower soil water potential led to reduced, but prolonged, growth and an adaptation at the molecular level without a drought stress response. Clustering of the protein and transcript data using a decision tree revealed different patterns in abundance changes across the growth stages and between end-of-day and end-of-night that are linked to specific biological functions. Correlations between protein and transcript levels depend on the time-of-day and also on protein localisation and function. Surprisingly, only very few of >1700 quantified proteins showed diurnal abundance fluctuations, despite strong fluctuations at the transcript level

Atypical E2F activity coordinates PHR1 photolyase gene transcription with endoreduplication onset

The atypical E2F transcription factor E2Fe/DEL1 regulates endoreduplication in plants. Here, DEL1 is shown to regulate the UV-B-induced DNA damage response by controlling expression of the DNA repair gene PHR1. Coupling of endoredublication and DNA repair promotes ploidy-driven cell enlargement that compensates for DNA damage-induced reduction in cell number. This feature enables plants to optimize photosynthetic output by maintaining leaf size under stress conditions