Light signaling controls nuclear architecture reorganization during seedling establishment

The spatial organization of chromatin can be subject to extensive remodeling in plant somatic cells in response to developmental and environmental signals. However, the mechanisms controlling these dynamic changes and their functional impact on nuclear activity are poorly understood. Here, we determined that light perception triggers a switch between two different nuclear architectural schemes during Arabidopsis postembryonic development. Whereas progressive nucleus expansion and heterochromatin rearrangements in cotyledon cells are achieved similarly under light and dark conditions during germination, the later steps that lead to mature nuclear phenotypes are intimately associated with the photomorphogenic transition in an organ-specific manner. The light signaling integrators DE-ETIOLATED 1 and CONSTITUTIVE PHOTOMORPHOGENIC 1 maintain heterochromatin in a decondensed state in etiolated cotyledons. In contrast, under light conditions cryptochrome-mediated photoperception releases nuclear expansion and heterochromatin compaction within conspicuous chromocenters. For all tested loci, chromatin condensation during photomorphogenesis does not detectably rely on DNA methylation-based processes. Notwithstanding, the efficiency of transcriptional gene silencing may be impacted during the transition, as based on the reactivation of transposable element-driven reporter genes. Finally, we report that global engagement of RNA polymerase II in transcription is highly increased under light conditions, suggesting that cotyledon photomorphogenesis involves a transition from globally quiescent to more active transcriptional states. Given these findings, we propose that light-triggered changes in nuclear architecture underlie interplays between heterochromatin reorganization and transcriptional reprogramming associated with the establishment of photosynthesis

Reduced dynamics of functional connectivity and cognitive impairment in multiple sclerosis

Background: In multiple sclerosis (MS), abnormalities of brain network dynamics and their relevance for cognitive impairment have never been investigated. Objectives: The aim of this study was to assess the dynamic resting state (RS) functional connectivity (FC) on 62 relapsing-remitting MS patients and 65 sex-matched healthy controls enrolled at 7 European sites. Methods: MS patients underwent clinical and cognitive evaluation. Between-group network FC differences were evaluated using a dynamic approach (based on sliding-window correlation analysis) and grouping correlation matrices into recurrent FC states. Results: Dynamic FC analysis revealed, in healthy controls and MS patients, three recurrent FC states: two characterized by strong intra- and inter-network connectivity and one characterized by weak inter-network connectivity (State 3). A total of 23 MS patients were cognitively impaired (CI). Compared to cognitively preserved (CP), CI-MS patients had reduced RS-FC between subcortical and default-mode networks in the low-connectivity State 3 and lower dwell time (i.e. time spent in a given state) in the high-connectivity State 2. CI-MS patients also exhibited a lower number and a less frequent switching between meta-states, as well as a smaller distance traveled through connectivity states. Conclusion: Time-varying RS-FC was markedly less dynamic in CI- versus CP-MS patients, suggesting that slow inter-network connectivity contributes to cognitive dysfunction in MS

MtNFP and MtLYK3, or AtCERK1 (co-)production in <i>Nicotiana</i> leaves induces defence-like responses.

<p>A, Kinetics of cell death development in <i>Nicotiana</i>. <i>Agrobacterium</i> transformants carrying either <i>MtNFP-3xFLAG</i> or <i>MtLYK3-3xFLAG</i> construct were co-infiltrated into <i>Nicotiana</i> leaves at five different time points (1–5). Macroscopic observation (left panel) and subsequent Evans blue staining (right panel) are depicted 42 hai (region 1), 39 hai (region 2), 36 hai (region 3), 33 hai (region 4) and 30 hai (region 5). Mock infiltration (region 6) was done concomitantly with the infiltration of region 1. Bar is 1 cm. B, Changes in leaf autofluorescence upon MtNFP and MtLYK3 co-production. Leaf regions co-producing MtNFP-3xFLAG and MtLYK3-3xFLAG fusions were analyzed between 24 and 48 hai (here depicted 36 hai) using a stereoscope. Note the decrease in chlorophyll content, as indicated by the decrease of far-red autofluorescence of chlorophyll (left panel), and enhanced accumulation of blue light-excited autofluorescence (right panel) within the infiltrated region. Bar is 1 cm. C, Accumulation of phenolic compounds. The following fusions were (co-)produced in <i>Nicotiana</i> leaves: MtNFP-3xFLAG (1); MtLYK3-3xFLAG (2); MtNFP-3xFLAG+MtLYK3-3xFLAG (3); MtNFP-3xFLAG+MtLYK3[G334E]-3xFLAG (4); AtCERK1-3xFLAG (5); or AtCERK1[K349]-3xFLAG (6). Macroscopic observations (left panel) and subsequent UV-excited autofluorescence of ethanol/lactophenol-cleared (right panel) leaf regions are depicted 36 hai (except for 5–30 hai). Bars are 1 cm. D, Induction of <i>NbHIN1, NbPR1 basic</i>, <i>NbACRE31</i>, and <i>NbACRE132</i> expression in response to separate production or co-production of: MtNFP-3xFLAG (NFP), MtLYK3-3xFLAG (LYK3), MtLYK3[G334E]<i>-</i>3xFLAG (LYK3[G334E]), and AtCERK1-3xFLAG (CERK1). Leaf samples were collected 24 hai and induction of gene expression was analyzed using qRT-PCR. Histograms represent induction of <i>NbHIN1</i> (white columns), <i>NbPR1 basic</i> (grey columns), <i>NbACRE31</i> (hatched columns), and <i>NbACRE132</i> (black columns) normalized by one reference gene, <i>MtEF1 α</i>. Induction of each gene was normalized to that caused by mock infiltration, and then calculated as % induction relative to the induction observed upon co-production of MtNFP and MtLYK3 fusions. Bars represent standard deviation of the mean. At least two technical replicates from two biological replicates were analyzed.</p

Cell death upon MtNFP and MtLYK3 co-production in <i>Nicotiana</i> leaves does not require <i>Sm</i>NF.

<p><i>Agrobacterium</i> transformants carrying either <i>MtNFP-3xFLAG</i> or <i>MtLYK3-3xFLAG</i> construct were co-infiltrated into <i>Nicotiana</i> leaves at a final concentration: OD₆₀₀ [<i>MtNFP</i>] = 0.25 and OD₆₀₀ [<i>MtLYK3</i>] = 0.4 (1); OD₆₀₀ [<i>MtNFP</i>] = 0.15 and OD₆₀₀ [<i>MtLYK3</i>] = 0.25 (2). Twelve hai parts of the transformed regions were syringe-infiltrated with 10⁻⁷mM <i>Sm</i>NF (circled in red) or DMSO diluted to the same concentration (circled in white). Macroscopic observation (left panel) and Evans blue staining (right panel) are depicted 33 hai. Bar is 1 cm.</p

Lanthanum chloride delays the cell death development upon MtNFP and MtLYK3, or AtCERK1 (co-)production.

<p><i>Agrobacterium</i> transformants carrying the following constructs were (co-)infiltrated at a final concentration: OD₆₀₀ [<i>MtNFP-3xFLAG</i>] = 0.125 and OD₆₀₀ [<i>MtLYK3-3xFLAG</i>] = 0.2 (A, B); OD₆₀₀ [<i>AtCERK1-3xFLAG</i>] = 0.2 (C, D). Twelve hai parts of the infiltrated regions were syringe-infiltrated with 5 mM lanthanum chloride (circled in red) or water (circled in white). Macroscopic observations (left panel) and subsequent Evans blue stainings (right panel) are depicted 42 hai for leaf regions co-producing MtNFP and MtLYK3 fusions (A, B), and 33 hai for leaf regions producing AtCERK1 fusion (C, D). Cell death development was scored 42 hai (A, B) or 33 hai (C, D): only infiltrations that showed the lack of tissue collapse and no compromised membrane permeability in the lanthanum chloride- or water-treated region were scored and are presented (right panel) as a fraction of total infiltrations performed. Bars are 1 cm.</p

Cell death induction activity of MtLYK3-sYFP2 mutated variants in <i>Nicotiana</i> leaves.

*<p>-see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0065055#pone.0065055-KlausHeisen1" target="_blank">[20]</a>, except for the T480A (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0065055#pone.0065055.s002" target="_blank">Fig. S2</a>), **-see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0065055#pone.0065055-KlausHeisen1" target="_blank">[20]</a>, except for the P87S <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0065055#pone.0065055-Smit1" target="_blank">[12]</a>, K464A and T480A (this study; number of plants nodulated/number of plants tested).</p><p>The designated constructs were expressed alone or co-expressed with <i>MtNFP-mCherry</i> construct in <i>Nicotiana</i> leaves. Macroscopic symptoms of cell death were scored 48 hai: only infiltrations that resulted in confluent death of (nearly) the entire infiltrated region were scored and are presented as a fraction of total infiltrations performed. # - despite the lack of pronounced macroscopic symptoms, the co-expression of <i>MtLYK3</i>[K464A]-<i>sYFP2</i> and <i>MtNFP-mCherry</i> constructs resulted in increased staining with Evans blue in the entire infiltrated region.</p

Production of AtCERK1 in <i>Nicotiana</i> leaves induces cell death that requires AtCERK1 kinase activity.

<p><i>AtCERK1-sYFP2</i> (A) and <i>AtCERK1</i>[K349E]-<i>sYFP2</i> (B) constructs were expressed in <i>Nicotiana</i> leaves. Macroscopic observations (left panel) and subsequent Evans blue stainings (right panel) are depicted 36 hai. Macroscopic symptoms of cell death were scored 36 hai: only infiltrations that resulted in confluent death of (nearly) the entire infiltrated region were scored and are presented (right panel) as a fraction of total infiltrations performed. Bars are 1 cm.</p