12 research outputs found
Assembly and function of the major histocompatibility complex (MHC) I peptide-loading complex are conserved across higher vertebrates
Antigen presentation to cytotoxic T lymphocytes via major histocompatibility complex class I (MHC I) molecules depends on the heterodimeric transporter associated with antigen processing (TAP). For efficient antigen supply to MHC I molecules in the ER, TAP assembles a macromolecular peptide-loading complex (PLC) by recruiting tapasin. In evolution, TAP appeared together with effector cells of adaptive immunity at the transition from jawless to jawed vertebrates and diversified further within the jawed vertebrates. Here, we compared TAP function and interaction with tapasin of a range of species within two classes of jawed vertebrates. We found that avian and mammalian TAP1 and TAP2 form heterodimeric complexes across taxa. Moreover, the extra N-terminal domain TMD0 of mammalian TAP1 and TAP2 as well as avian TAP2 recruits tapasin. Strikingly, however, only TAP1 and TAP2 from the same taxon can form a functional heterodimeric translocation complex. These data demonstrate that the dimerization interface between TAP1 and TAP2 and the tapasin docking sites for PLC assembly are conserved in evolution, whereas elements of antigen translocation diverged later in evolution and are thus taxon specific
Complete male-to-female sex reversal in XY mice lacking the miR-17~92 cluster
Trabajo presentado en EMBO Workshop The evolution of animal genomes, celebrado en Sevilla (España) del 18 al 21 de septiembre de 2023.Peer reviewe
A Transcriptome Meta-Analysis Proposes Novel Biological Roles for the Antifungal Protein AnAFP in Aspergillus niger.
Understanding the genetic, molecular and evolutionary basis of cysteine-stabilized antifungal proteins (AFPs) from fungi is important for understanding whether their function is mainly defensive or associated with fungal growth and development. In the current study, a transcriptome meta-analysis of the Aspergillus niger γ-core protein AnAFP was performed to explore co-expressed genes and pathways, based on independent expression profiling microarrays covering 155 distinct cultivation conditions. This analysis uncovered that anafp displays a highly coordinated temporal and spatial transcriptional profile which is concomitant with key nutritional and developmental processes. Its expression profile coincides with early starvation response and parallels with genes involved in nutrient mobilization and autophagy. Using fluorescence- and luciferase reporter strains we demonstrated that the anafp promoter is active in highly vacuolated compartments and foraging hyphal cells during carbon starvation with CreA and FlbA, but not BrlA, as most likely regulators of anafp. A co-expression network analysis supported by luciferase-based reporter assays uncovered that anafp expression is embedded in several cellular processes including allorecognition, osmotic and oxidative stress survival, development, secondary metabolism and autophagy, and predicted StuA and VelC as additional regulators. The transcriptomic resources available for A. niger provide unparalleled resources to investigate the function of proteins. Our work illustrates how transcriptomic meta-analyses can lead to hypotheses regarding protein function and predict a role for AnAFP during slow growth, allorecognition, asexual development and nutrient recycling of A. niger and propose that it interacts with the autophagic machinery to enable these processes
Analysis of <i>anafp</i> expression using a fluorescent reporter system.
<p>A) Luciferase expression under control of the <i>anafp</i> promoter was measured in a <i>ΔcreA</i> strain (NP6.4) and the corresponding wild type control strain (PK2.9) during 4 days of cultivation in complete medium using microtiter plates. Protein abundance was measured as luminescent counts per second normalized to culture optical density. Data of a representative experiment (n = 4) are shown. B) eYFP expression under control of the <i>anafp</i> promoter was measured in a <i>ΔcreA</i> strain (NP2.8, grey) and the corresponding wild type control strain (PK1.22, black) after 3 days of incubation on a complete medium agar. Expression levels were normalized to that of the control strain and are depicted as mean of three independent experiments performed as duplicate. *, p<0.03.</p
Species owning AnAFP orthologs within the <i>Ascomycota</i> tree of life.
<p>Depicted is an excerpt of the <i>Ascomyota</i> tree of life according to Schoch <i>et al</i>., 2009 [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0165755#pone.0165755.ref064" target="_blank">64</a>]. Species with AnAFP orthologs are listed according to their class. Corresponding superclasses are indicated by numbers 1–3. 1, <i>Sordariomyceta</i>; 2, <i>Leotiomyceta</i>; 3, <i>Dothideomyceta</i>. Numbers of species/genus are indicated in brackets. Dashed line represents the last common ancestor of the three superclasses.</p
Minimal inhibitory concentrations (MICs) of AnAFP and AFP on selected fungi.
<p>Minimal inhibitory concentrations (MICs) of AnAFP and AFP on selected fungi.</p
Genes being positively correlated with <i>anafp</i> expression as inferred from 155 cultivation conditions.
<p>Genes being positively correlated with <i>anafp</i> expression as inferred from 155 cultivation conditions.</p
Expression levels of the <i>anafp</i> gene of <i>A</i>. <i>niger</i> strain N402 at different conditions.
<p>Transcription levels of the <i>anafp</i> gene were analyzed when exposed to different A) carbon sources in shaking flask cultures [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0165755#pone.0165755.ref023" target="_blank">23</a>] and B) to carbon limitation in batch, retentostat [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0165755#pone.0165755.ref027" target="_blank">27</a>] and chemostat bioreactor cultures [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0165755#pone.0165755.ref028" target="_blank">28</a>]. <i>anafp</i> transcription levels are also depicted for batch cultivations in bioreactors with <i>ΔflbA</i> and <i>ΔbrlA</i> mutants, respectively [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0165755#pone.0165755.ref033" target="_blank">33</a>]. Samples have been taken at four time points: t1–t4, exponential growth phase, 16 h, 60 h, 140 h post carbon depletion. In C) mRNA levels at different zones of a plated culture are depicted [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0165755#pone.0165755.ref033" target="_blank">33</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0165755#pone.0165755.ref034" target="_blank">34</a>]. Whereas zone 1 (z1) marks the center of the colony consisting, i.e. the oldest part of the culture, zone 5 (z5) marks the colony’s periphery. Zone 3 lies in between. Absolute values of transcription levels are depicted as arbitrary units of fluorescence intensity in a logarithmic scale and can be assessed using the legend. Color intensities were normalized to the highest absolute transcription level which is indicated by black color.</p
Membrane permeabilization induced by AnAFP in <i>A</i>. <i>niger</i>.
<p>The capability of AnAFP (100 μg/ml) to permeabilize the membranes of viable cells of <i>A</i>. <i>niger</i>, <i>A</i>. <i>nidulans</i> and <i>F</i>. <i>oxysporum</i> was monitored for 2 h of incubation. The fluorescence of the SYTOX Green dye was used as a measure of fungal cells with compromised membranes. Cells with intact membranes do not show a fluorescence signal. Data are averages of triplicate experiments.</p
Alignment of primary and secondary structures of AnAFP orthologs.
<p>Identified or proposed amino-acid sequences of mature proteins were aligned by CLUSTAL W. Names of corresponding organisms owning the orthologue are abbreviated using the first letter of genus name followed by the first three letters of the species name. Protein/GenBank accession numbers from UniProtKB, AspGD or NCBI, respectively, are indicated in brackets. Amino-acid sequences marked by asterisks were derived from an expressed sequence-tag database analysis. Though some of the listed mature proteins are identical (marked by square brackets), all of them differ at least in one amino acid when considering the pre-pro-regions as well. The overall conserved cysteine residues are highlighted by open boxes. Predicted γ-core motifs are highlighted by shaded boxes. Secondary structures derived from NMR solution structures of AFP [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0165755#pone.0165755.ref070" target="_blank">70</a>] and PAF [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0165755#pone.0165755.ref071" target="_blank">71</a>] and from a secondary structure prediction of AFP, PAF or AnAFP, respectively, are shown at the top. Secondary structure predictions have been calculated using the PredictProtein server [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0165755#pone.0165755.ref066" target="_blank">66</a>] for all mature proteins. Blue arrows and capital E-letters on top depict positions of beta strands whereas lines mean loops or turns, respectively. Gaps are depicted by dots.</p