Chiapas : entre el autoritarismo y la transición democrática, desde Acteal hasta el diálogo EZLN y sociedad civil

Obra cuya intención es dar cuenta de los diversos actores sociales involucrados en el conflicto de Chiapas, a nivel político, económico y sociocultural. El texto recupera un taller de análisis de coyuntura sobre el tema, realizado en agosto de 1998.ITESO, A.C

Comparative Live-Cell Imaging Analyses of SPA-2, BUD-6 and BNI-1 in Neurospora crassa Reveal Novel Features of the Filamentous Fungal Polarisome

A key multiprotein complex involved in regulating the actin cytoskeleton and secretory machinery required for polarized growth in fungi, is the polarisome. Recognized core constituents in budding yeast are the proteins Spa2, Pea2, Aip3/Bud6, and the key effector Bni1. Multicellular fungi display a more complex polarized morphogenesis than yeasts, suggesting that the filamentous fungal polarisome might fulfill additional functions. In this study, we compared the subcellular organization and dynamics of the putative polarisome components BUD-6 and BNI-1 with those of the bona fide polarisome marker SPA-2 at various developmental stages of Neurospora crassa. All three proteins exhibited a yeast-like polarisome configuration during polarized germ tube growth, cell fusion, septal pore plugging and tip repolarization. However, the localization patterns of all three proteins showed spatiotemporally distinct characteristics during the establishment of new polar axes, septum formation and cytokinesis, and maintained hyphal tip growth. Most notably, in vegetative hyphal tips BUD-6 accumulated as a subapical cloud excluded from the Spitzenkörper (Spk), whereas BNI-1 and SPA-2 partially colocalized with the Spk and the tip apex. Novel roles during septal plugging and cytokinesis, connected to the reinitiation of tip growth upon physical injury and conidial maturation, were identified for BUD-6 and BNI-1, respectively. Phenotypic analyses of gene deletion mutants revealed additional functions for BUD-6 and BNI-1 in cell fusion regulation, and the maintenance of Spk integrity. Considered together, our findings reveal novel polarisome-independent functions of BUD-6 and BNI-1 in Neurospora, but also suggest that all three proteins cooperate at plugged septal pores, and their complex arrangement within the apical dome of mature hypha might represent a novel aspect of filamentous fungal polarisome architecture

Clonal chromosomal mosaicism and loss of chromosome Y in elderly men increase vulnerability for SARS-CoV-2

The pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2, COVID-19) had an estimated overall case fatality ratio of 1.38% (pre-vaccination), being 53% higher in males and increasing exponentially with age. Among 9578 individuals diagnosed with COVID-19 in the SCOURGE study, we found 133 cases (1.42%) with detectable clonal mosaicism for chromosome alterations (mCA) and 226 males (5.08%) with acquired loss of chromosome Y (LOY). Individuals with clonal mosaic events (mCA and/or LOY) showed a 54% increase in the risk of COVID-19 lethality. LOY is associated with transcriptomic biomarkers of immune dysfunction, pro-coagulation activity and cardiovascular risk. Interferon-induced genes involved in the initial immune response to SARS-CoV-2 are also down-regulated in LOY. Thus, mCA and LOY underlie at least part of the sex-biased severity and mortality of COVID-19 in aging patients. Given its potential therapeutic and prognostic relevance, evaluation of clonal mosaicism should be implemented as biomarker of COVID-19 severity in elderly people. Among 9578 individuals diagnosed with COVID-19 in the SCOURGE study, individuals with clonal mosaic events (clonal mosaicism for chromosome alterations and/or loss of chromosome Y) showed an increased risk of COVID-19 lethality

Subcellular localization patterns of polarisome components during key developmental stages in <i>N. crassa</i>.

<p>(-) no specific subcellular recruitment observed/no colocalization indicated/no unique role observed;</p>‡<p>Colocalization inferred from identical subcellular localization patterns; CSB = cell symmetry breaking, CAT = conidial anastomosis tube, Spk = Spitzenkörper, VHF = vegetative hyphal fusion (in the mature colony), KO = knock-out (gene deletion) mutant;</p><p>References:</p>1<p>Araujo-Palmorales, Master Thesis, CICESE, 2007;</p>2<p>Araujo-Palmorales et al., 2009;</p>3<p>Justa-Schuch et al., 2010.</p

A Δ<i>bni-1</i> strain generated through vegetative homokaryon selection phenocopied growth defects of Δ<i>bud-6</i>.

<p>(<b>A and B</b>) Wild-type like phenotype of the heterokaryotic Δ<i>bni-1</i> strain FGSC11490, including conidia and septa (arrows). Scale bars, 50 µm and 10 µm respectively. (<b>C</b>) The lack of septa in the homokaryotic Δ<i>bni-1</i> strain was confirmed by FM4–64 staining. Scale bar 50 µm. (<b>D</b>) FM4–64 staining also confirmed the absence of an organized apical tip growth apparatus, including the Spk. Scale bar, 10 µm. These defects closely resembled phenotypic key features of Δ<i>bud-6</i> (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0030372#pone-0030372-g007" target="_blank">Figures 7</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0030372#pone-0030372-g008" target="_blank">8</a>).</p

Loss of BUD-6 resulted in a reduced colony extension rate and hyperbranching.

<p>(<b>A</b>) Hyperbranching and polar extension defects resulted in very slowly and extremely dense developing mycelial colonies of Δ<i>bud-6</i>, in comparison to the wild type after 24 hours of incubation. (<b>B</b>) Hyphal morphology of wild type and Δ<i>bud-6</i> at the colony margin. Scale bars, 0.5 mm and 0.25 mm, respectively. (<b>C</b>) Quantification of branching frequency, which on average was more than doubled in the mutant compared to the wild type. (<b>D</b>) Comparison of average colony extension speed between Δ<i>bud-6</i> and wild type.</p

BUD-6 accumulation during conidiogenesis.

<p>(<b>A</b>) BUD-6-GFP accumulated at septation sites in developing macroconidiophores (arrowhead). Scale bars, 10 µm. (<b>B</b>) In cytologically separated, but physically still attached conidia, BUD-6 fluorescence persisted at the cell poles; either at both or only at one pole in case of the terminal conidium. Scale bars, 10 µm. (<b>C</b>) In addition to strong fluorescence at the cell poles, bright clusters of BUD-6-GFP also accumulated at other locations of the cell cortex. Scale bar, 5 µm.</p

BNI-1 localization during septal plugging, tip repolarization and septum formation.

<p>(<b>A</b>) Two minutes after damaging leading hyphae BNI-1 became recruited to the septal plug (position of the Woronin body is indicated with an arrow, 0 min). Fluorescence focused into a smaller area from which a new hyphal tip repolarized, and shortly after condensed into an subapical spot (arrowhead, 16 min) with flanking BNI-1 crescents on either side (inset, 20 min). In parallel, a new septum was being formed about 25 µm behind the severed end, and an additional site of polarity was established (arrowhead, 20 min). Scale bar, 5 µm. See <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0030372#pone.0030372.s014" target="_blank">Movie S11</a> for full sequence. (<b>B</b>) Continuation of (A) but with an extended field of view including an old septum (asterisk). The part of the hypha shown in (A) is outlined with a dashed box. A selection of individual optical slices shows the formation of several septa. Upon septum completion, BNI-1 gradually disappeared from the septal pore. Barely visible remains are indicated with circles at the 68 min time point. BNI-1 fluorescence was usually not observed at ‘old’ septa (asterisk). Recruitment of BNI-1 to a vegetative hyphal fusion site is indicated by an arrowhead.</p

BUD-6 localization in conidia, conidial germlings and sites of septum formation.

<p>(<b>A</b>) Intense and locally defined clusters of BUD-6 were localized at the cell poles of dormant macroconidia. Scale bars, 5 µm. (<b>B–C</b>) Specific recruitment of BUD-6 during isotropic expansion, cell symmetry breaking and germ tube outgrowth was not observed. Scale bars, 2.5 µm. (<b>D</b>) Localized recruitment of BUD-6-GFP to apical caps of growing germ tubes (arrowhead), as well as to septa (arrow), occurred in germlings ≥35 µm in length. FM4–64 stained vesicle clusters were observed at the same positions. (<b>E</b>) Enlarged view of the germ tube tip highlighted in (D). The merged image shows that BUD-6-GFP fluorescence only partially colocalizes with the FM4–64 stained apical cap which extends over a larger crecent. (<b>F</b>) Staining with FM4–64 revealed that BUD-6 recruitment to the incipient septation site preceded plasma membrane invagination (arrowheads), and that it constantly remained associated with the leading edge of the closing septum. Scale bar, 5 µm. See <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0030372#pone.0030372.s004" target="_blank">Movie S1</a> for time course sequences. Note: the bright FM-4–64 stained spot appearing at time point 0 sec does not colocalize with the cortical BUD-6 accumulation indicated by the arrowhead. (<b>G</b>) Reconstruction of BUD-6-GFP localization at the inner perimeter of the closing septal pore. Scale bar, 2.5 µm.</p

BUD-6 dynamics during vegetative hyphal fusion.

<p>(<b>A</b>) BUD-6 became recruited to the tips of homing fusion hyphae, then concentrated at the attachment point and surrounded the opening fusion pore. Shortly after the pore was fully established BUD-6 fluorescence disappeared from this site. Scale bar, 5 µm. See <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0030372#pone.0030372.s011" target="_blank">Movie S8</a> for time course sequence. (<b>B</b>) Transient BUD-6 fluorescence accumulated at incipient fusion sites in the mature colony (arrowhead) and persistent BUD-6 signal at septal pores (all other fluorescently marked sites). Scale bar, 10 µm. (<b>C</b>) SPA-2-GFP became recruited to vegetative hyphal fusion sites (arrow). As it was never seen at completed fusion connections (arrowheads) it must follow the transient dynamics of BUD-6 in this context. Scale bar, 10 µm.</p