219 research outputs found
Aquaporin splice variation differentially modulates channel function during marine teleost egg hydration
No full text22 pages, 8 figures, supporting information https://doi.org/10.1371/journal.pone.0294814.-- Data Availability: The full-length cDNAs for SaAqp1ab1-WT, HhAqp1ab1-WT and SsAqp1ab2-WT are available in GenBank under accession numbers AY626938, MW960022 and DQ889223, respectively. The full-length transcripts of SaAqp1ab1_v1, SaAqp1ab1_v2, HhAqp1ab1_v1, SsAqp1ab2_v1 and SsAqp1ab2_v2 were deposited in GenBank under accession numbers OR498208, OR498209, OR498210, OR498211 and OR498212, respectively. All relevant data are within the paper and its Supporting information filesAquaporin-mediated oocyte hydration is a developmentally regulated adaptive mechanism that co-occurs with meiosis resumption in marine teleosts. It provides the early embryos with vital water until osmoregulatory systems develop, and in the majority of marine teleosts causes their eggs to float. Recent studies have shown that the subdomains of two water channels (Aqp1ab1 and Aqp1ab2) encoded in a teleost-specific aquaporin-1 cluster (TSA1C) co-evolved with duplicated Ywhaz-like (14-3-3ζ-like) binding proteins to differentially control their membrane trafficking for maximal egg hydration. Here, we report that in species that encode the full TSA1C, in-frame intronic splice variants of Aqp1ab1 result in truncated proteins that cause dominant-negative inhibition of the canonical channel trafficking to the plasma membrane. The inhibition likely occurs through hetero-oligomerization and retention in the endoplasmic reticulum (ER) and ultimate degradation. Conversely, in species that only encode the Aqp1ab2 channel we found an in-frame intronic splice variant that results in an intact protein with an extended extracellular loop E, and an out-of frame intronic splice variant with exon readthrough that results in a truncated protein. Both isoforms cause dominant-negative enhancement of the degradation pathway. However, the extended and truncated Aqp1ab2-type variants can also partially escape from the ER to reach the oocyte plasma membrane, where they dominantly-negatively inhibit water flux. The ovarian follicular expression ratios of the Aqp1ab2 isoforms in relation to the canonical channel are lowest during oocyte hydration, but subsequently highest when the canonical channel is recycled, thus leaving the eggs endowed with >90% water. These findings suggest that the expression of inhibitory isoforms of Aqp1ab1 and Aqp1ab2 may represent a new regulatory mechanism through which the cell-surface expression and the activity of the canonical channels can be physiologically modulated during oocyte hydration in marine teleostsThis work was supported by the Spanish Ministry of Science and Innovation (MCIN/AEI/10.13039/501100011033), and the European Regional Development Fund (ERDF) “A way of making Europe" (European Union), Grant no. AGL2016-76802-R (to J.C.). A.F. was recipient of a predoctoral contract from Spanish MCIN (BES-2014-068745). R.N.F. was supported by the University of Bergen (Norway)With the institutional support of the ‘Severo Ochoa Centre of Excellence’ accreditation (CEX2019-000928-S)Peer reviewe
Polyphony and the anxiety of influence in the fiction of Henry James
No full textJames's fiction, especially in the Middle Phase, centres
on the figure of the artist and is characterized by, the two
interrelated aspects which previous criticism has largely
overlooked: the Bakhtinian 'polyphonic' -creation of
'author-thinkers'; and the conflict between ephebes and
precursors, for which Harold-Bloom's concept of 'the-anxiety of
influence' is the most illuminating model. Polyphony is the
narrative mode, and influence is the intra-artistic, theme.
These, as the Introduction to the thesis makes clear, are
rehearsed in James's inaugural novel, Roderick Hudson. Rowland
Mallet is an author-thinker, and his failure is caused by
authorial limitations. His monologism -is impaired by his
mistaking empathy for the authorial sympathy. Likewise,
Hudson's failure does not arise from a mercurial temperament,
but from a polyphonic shortcoming: not possessing the power of
fiction to contain the fiction of power in, his mentor. And the
relationships among the three artists - Gloriani, Hudson and
Singleton - perfectly exemplify the Bloomian-theme. It is these
two concepts, polyphony and influence, which are the major
preoccupation in the Middle Phase; as, the works chosen
demonstrate. These are a novella, a novel, and a number of
short stories all of which have been unjustifiably neglected.
Chapter One, on The Aspern Papers, argues that Tina Bordereau,
far from being, the artless victim seen by many critics,
actually challenges and defeats the narrator by the very form
of her narrative. Her 'realist' discourse undermines his
language of 'romance', and shows up its internal unstability.
Chapter Two is an extensive study of the critical reception of
The Tragic Muse. The most common areas of critical attention
have been its contemporary topicality, its relation to previous
novels on similar themes, and the possible genealogy of Gabriel
Nash. Those have all missed the core of the work. - Chapter Three
demonstrates how polyphony and the anxiety of influence make
the novel what it really is. Influence arises from the
juxtaposition of, and the wrestling between, artistic ephebes
and their precursors (Nick and Nash,, Miriam and Madame Carre).
The dialogic quality defined by Bakhtin is crucial to the
proper, and even-handed, characterization of all, the conflicts
in the novel. And since most of James's tales in the eighties
and nineties -are about 'masters - and acolytes, the anxiety of
influence remains central. Chapter Four is a study of 'The
Author of Beltraffiol' and 'The Lesson of the Master'. Again the
characters' manipulations are a crucial focus in a way that
G6rard Genette's terminology helps to illuminate. The fact that
the ephebe is the author-thinker emphasizes the inextricability
of the Bakhtinian and the Bloomian in James. Just as
polyphony offers a different focus for explicating the poetics
of James's fiction; so the ephebal conflict provides the basis
for a fresh perception of James's own artistic struggle
Raw RT-PCR gels images.
No full textAquaporin-mediated oocyte hydration is a developmentally regulated adaptive mechanism that co-occurs with meiosis resumption in marine teleosts. It provides the early embryos with vital water until osmoregulatory systems develop, and in the majority of marine teleosts causes their eggs to float. Recent studies have shown that the subdomains of two water channels (Aqp1ab1 and Aqp1ab2) encoded in a teleost-specific aquaporin-1 cluster (TSA1C) co-evolved with duplicated Ywhaz-like (14-3-3ζ-like) binding proteins to differentially control their membrane trafficking for maximal egg hydration. Here, we report that in species that encode the full TSA1C, in-frame intronic splice variants of Aqp1ab1 result in truncated proteins that cause dominant-negative inhibition of the canonical channel trafficking to the plasma membrane. The inhibition likely occurs through hetero-oligomerization and retention in the endoplasmic reticulum (ER) and ultimate degradation. Conversely, in species that only encode the Aqp1ab2 channel we found an in-frame intronic splice variant that results in an intact protein with an extended extracellular loop E, and an out-of frame intronic splice variant with exon readthrough that results in a truncated protein. Both isoforms cause dominant-negative enhancement of the degradation pathway. However, the extended and truncated Aqp1ab2-type variants can also partially escape from the ER to reach the oocyte plasma membrane, where they dominantly-negatively inhibit water flux. The ovarian follicular expression ratios of the Aqp1ab2 isoforms in relation to the canonical channel are lowest during oocyte hydration, but subsequently highest when the canonical channel is recycled, thus leaving the eggs endowed with >90% water. These findings suggest that the expression of inhibitory isoforms of Aqp1ab1 and Aqp1ab2 may represent a new regulatory mechanism through which the cell-surface expression and the activity of the canonical channels can be physiologically modulated during oocyte hydration in marine teleosts.</div
Tagging HhAqp1ab1 and SsAqp1ab2 constructs do not affect channel function.
No full textPf of X. laevis oocytes injected with water (W, controls), or expressing non-tagged or Flag-tagged Atlantic halibut HhAqp1ab1-WT or sole SsAqp1ab2-WT. Oocytes injected with SsAqp1ab2-WT were co-injected with non-tagged halibut YwhazLb and exposed to FSK. The data are the mean ± SEM (n = 12 oocytes per treatment, indicated with dots above each bar). (TIF)</p
Commonwealth caprice [music] : pour piano /
No full textCaption title.; Date approximated from p. 2, Traralgon Record, Tuesday 23 December 1902: "We have received from the composer, Miss Daisy R. Hughes, daughter of Mr E.F. Hughes, pro prietor of the 'Casterton News,' a copy of a special piece of pianoforte music, entitled the 'Commonwealth Caprice' published by Messrs Allan and Co., Melbourne, from whom it can be procured wholesale at a very moderate price."--http://nla.gov.au/nla.news-article64264107; NLA's N copy: Cover inscribed by composer. ANL; Also available online http://nla.gov.au/nla.mus-vn5350188; NLA's N copy from the collection of Keith Watson. ANL
The dominant-negative activity of teleost Aqp1ab1 and -1ab2 splice forms.
No full text(A-C) Changes in Pf of X. laevis oocytes injected with water or expressing SaAqp1ab1-WT (A), HhAqp1ab1-WT (B) or SsAqp1ab2-WT (C) alone or in combination with different amounts of the isoforms of each paralog. For SsAqp1ab2-WT and splice forms, oocytes were co-injected with halibut YwhazLb and exposed to FSK. (D) Effect of the inhibition of the oocyte Pf by the different Aqp1ab1 splice forms, at a concentrations that produced approximately half-maximal reduction (as observed in A-C), in SaAqp1ab1 or HhAqp1ab1 expressing oocytes in the presence or absence of YwhazLa, and exposed to FSK. The percentage of Pf inhibition elicited by each isoform under the two conditions is indicated in each plot. In all panels, data are the mean ± SEM (n indicated above each bar) and were statistically analyzed by an unpaired Student’s t-test (*, P P P < 0.001; with respect to oocytes injected with the WT form alone).</p
Differential regulation of Aqp1ab2 splice variants during ovarian follicle maturation and hydration <i>in vivo</i>.
No full text(A) Representative immunoblots of protein extracts from follicles at the previtellogenic, vitellogenic), hydrating, and mature stage (2.5 follicle equivalents/lane) using the α-Aqp1ab2-Nt antisera. The blots show extracts from three different pools of follicles collected from three different females. The arrowheads indicate the Aqp1ab2-WT, Aqp1ab2_v1 and Aqp1ab2_v2 proteins according to their apparent molecular masses. In all blots, molecular mass markers (kDa) are on the left. (B) Relative amount of the Aqp1ab2_v1 (left) and Aqp1ab2_v2 (right) isoforms with respect to the Aqp1ab2-WT at the different development stages based on the blots shown in A. Bars (mean ± SEM; n = 3 fish) with different superscript are statistically significant (one-way ANOVA; P < 0.05).</p
Functional characterization and subcellular localization of teleost wild-type Aqp1ab1 and -1ab2 and their splice variants.
No full text(A-C) Pf of X. laevis oocytes injected with water (W, controls) or expressing seabream SaAqp1ab1-WT, SaAqp1ab1_v1 or SaAqp1ab1_v2 (A), Atlantic halibut HhAqp1ab1-WT or HhAqp1ab1_v1 (B), or sole SsAqp1ab2-WT, SsAqp1ab2_v1 or SsAqp1ab2_v2 (C). Oocytes injected with SsAqp1ab2-WT or splice forms were co-injected with halibut YwhazLb and exposed to FSK for 1 h prior to the swelling assay. The data are the mean ± SEM (n indicated above each bar) and were statistically analyzed by an unpaired Student’s t-test (***, P D-F) Double immunostaining of oocytes expressing untagged SaAqp1ab1-WT or HA-tagged SaAqp1ab1_v1 or HA-SaAqp1ab1_v2 (D), Flag-tagged HhAqp1ab1-WT or HA-tagged HhAqp1_v1 (E), or Flag-tagged SsAqp1ab2-WT or HA-tagged SsAqp1ab2_v1 or HA-SsAqp1ab2_v2 (F), and the ER marker protein disulfide isomerase (PDI). The plasma membrane is indicated by an arrowhead, whereas the co-localization of aquaporin channels and PDI in the cytoplasm is indicated by arrows. Scale bars, 10 μm (insets 5 μm).</p
Production of Senegalese sole total Aqp1ab2 and Aqp1ab2_v1 specific antisera and immunoblot analysis of Aqp1ab2 splice variants protein expression in developing ovarian follicles.
No full text(A) Amino acid sequence alignment of SsAqp1ab2-WT, SsAqp1ab2_v1 and SsAqp1ab2_v2 splice forms highlighting the sequences employed for the production of the sole-specific Aqp1ab2 and Aqp1ab2_v1 antibodies (α-Aqp1ab2-Nt and α-Aqp1ab2_v1, in red and blue color, respectively). The N- and C-termini of the WT channel, the six transmembrane helices (TM1-TM6) and the two conserved NPA motifs are also indicated. (B) Immunoblots of X. laevis oocytes injected with water or expressing Aqp1ab2-WT, SsAqp1ab2_v1 or SsAqp1ab2_v2 and probed with the α-Aqp1ab2-Nt or α-Aqp1ab2_v1 antisera separately. (C) Representative photomicrographs of sole ovarian follicles at different developmental stages during oocyte growth and maturation. PG, primary growth stage; CA, cortical alveoli stage. Scale bars, 50 and 250 μm. (D) Immunoblots of Aqp1ab2-WT, Aqp1ab2_v1 and Aqp1ab2_v2 in protein extracts from previtellogenic (Pv), vitellogenic (Vt), hydrating (H) and mature (Mt) follicle-enclosed oocytes, as well as from ovulated oocytes (Ov), using the generated antisera. Alpha tubulin (Tuba) was used as loading control. Duplicated blots (right) were run in parallel and incubated with the primary antibodies preadsorbed by the antigenic peptide to test for specificity. The brackets indicate potential post-translational modifications. (E) Hoechst staining of intact vitellogenic follicles and defolliculated oocytes. Scale bar, 100 μm. (F) Immunoblots of Aqp1ab2 isoforms in total protein extracts from intact or defolliculated vitellogenic follicles (+/- follicle cells, Fc). In B, D and E, the arrowheads indicate aquaporin monomers, whereas the asterisks in D and E indicate cross-reactive polypeptides revealed with the Aqp1ab2_v1 antiserum. In all blots, molecular mass markers (kDa) are on the left.</p
Modelling desiccation cracking in a homogenous soil clay layer: comparison between different hypotheses on constitutive behaviour
No full textDesiccation cracks are usually thought to start from the surface of an evaporating soil layer, and the available simplified models for crack initiation and propagation are based on this hypothesis. On the contrary, experimental results on a Dutch river clay showed that cracks in an evaporating soil layer may start and propagate below the surface, confirming earlier findings by other researchers. A simple one-dimensional model was set up to analyse the consequences of different hypotheses about the material behaviour on the crack onset in a homogenous soil layer undergoing surface drying. The results of the model show that dependence of the material behaviour on the rate of water content change is a necessary requirement for cracks to initiate below the surface. The conclusion suggests that, to properly understand cracking in an evaporating soil layer, an intrinsic time scale for the mechanical response must be accounted for, among all the other factors which were previously highlighted by other researchers. The key factor to predict crack onset below the surface is the dependence of the drying branch of the water retention curve of the compressible soil on the rate of drying, which would be justified by a rate dependent fabric evolution
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