Interaction between the Alfalfa mosaic virus movement protein and plasmodesmata

For a full infection of a host, plant viruses should be able to multiply in the initially infected cell and to spread to neighbouring cells as to eventually invade the entire plant. The viral transport pathway can in principle be divided into two steps, i.e. cell-to-cell movement within tissues, and long-distance transport via the vascular system. The first process, movement from cell-to-cell, is an "active" process, in terms of that a viral gene product, the so-called "movement protein" (MP) is essentially involved by modifying the host's plasmodesmata to allow viral infectivity to pass. So far at least two different mechanisms of cell-to-cell movement have been described, i.e. the "tobamovirus-type" mechanism, involving transport of virus in a non-virion form through (cytologically) slightly modified plasmodesmata, and the "comovirus-type" mechanism, involving the formation of transport tubules in heavily modified plasmodesmata and allowing mature virus particles to pass. In both mechanisms the viral MP plays a role in the modification of the host's plasmodesmata, and in case of the tobamovirus movement, an increased size exclusion limit of the plasmodesmata is detected.The PhD research project described in this thesis was part of a programme aiming to unravel the role of the plant's plasmodesmata in virus transport and in cell-cell communication. An important approach in this programme was to use the MPs of two different viruses, each representing both distinct viral transport mechanisms, as probes to identify which host proteins are involved in viral movement and possibly investigate the molecular make up of the plasmodesma. For this purpose the viruses Alfalfa mosaic virus (AMV) and Cowpea mosaic virus (CPMV) were selected, which moreover share a common host, i.e. Nicotiana benthamiana , allowing us to study viral MP-plasmodesma interactions in a single host. Previous work had demonstrated that CPMV uses a tubule-guided movement mechanism, whereas it was anticipated that AMV, in view of its genetic relationship to tomaboviruses, would use the tobamo-type transport. The rational behind the choice for this combination was that the MPs of these two viruses would interact with plasmodesmal constituents or proteins in a distinct way and thus would represent different probes for identifying plasmodesmal proteins and other MP-interacting host proteins in e.g. the yeast two-hybrid system.This thesis covers the cytological studies within the research programme. The first part of this thesis (Chapters 2-5) deals with the unravelling of the cell-to-cell movement mechanism as utilised by AMV, and in the last part (Chapter 6) cytological studies are presented focussing on a host protein (denoted AD3) interacting with the AMV MP in the yeast two-hybrid system (PhD thesis L. Jongejan, UL, The Netherlands, in preparation).At the onset of this PhD research hardly any information on the cell-to-cell movement mechanism of AMV was available. Using cowpea protoplasts as test system, it was first shown (Chapter 2) that AMV, like CPMV, is able to form tubular structures on infected plant cells, unexpectedly indicating that AMV would also use a tubule-guided movement mechanism like CPMV. Further analysis indeed demonstrated that these MP-constructed protrusions contain mature virus particles.To confirm whether AMV, similar to CPMV, would use these structures for a tubule-guided cell-to-cell mechanism, different mutants, defective in systemic spread, were tested for their (in)capability to induce virus-filled tubules. The CP mutant CP4P was previously reported to be unable to form stable virions and was now shown to induce empty tubules on protoplasts (Chapter 3). Moreover, both MP mutants SP6 and SP7 appeared unable to form tubules on infected protoplasts. Hence, the results with all three mutants indicated that the inability to produce virion-filled tubules on single plant protoplasts coincides with a transport-deficient phenotype. These three mutants were completely deficient in cell-cell movement in parenchyma tissues, a restricted spread of these viruses to neighbouring cells was observed in cortex cells surrounding the vascular system. Whether this indicated a second, tissue-dependent minor movement mechanism of AMV or whether this spread reflected distinct properties of plasmodesmata of cortex cells, was not further investigated.During intensive cytological searches in systemically AMV-infected N. benthamiana plants no elongated tubules could be discerned, despite an earlier report. At the front of infection modified plasmodesmata were found containing both the AMV CP and MP, and having a significantly wider diameter than those in non-infected as well as fully infected tissues (Chapter 4). This finding implied that the modification of the plasmodesmata by AMV is only transient, restricted to the front of infection. Cryo-sectioning of such plasmodesmata revealed the presence of rows of virus particles within the interior of the plasmodesmal pore, which might suggest that only short, not-extending tubules are formed (Chapter 4 and Figure 1 of General Discussion)Furthermore, not only a transient modification of plasmodesmata was observed in AMV-infected leaf tissue, also a temporary increase of their number at the infection front was noted (Chapter 5). By inspecting cell walls in mesophyll tissue respectively in front of, at, and after the infection zone it was calculated that the density of plasmodesmata in the infection zone at least doubled. It remains to be resolved whether this increase is due to specific viral induction of new plasmodesmata to become engaged in viral transport or whether this reflects a host response to restore cell-to-cell communication after a major part of the pre-existing plasmodesmata have become modified.In a simultaneously executed PhD project within the same programme (by L. Jongejan, PhD thesis, University of Leiden, in preparation), the AMV MP was used as bait in the yeast two-hybrid system to identify host ( N. benthamiana ) proteins which are specifically targeted by this viral protein during the infection process. Chapter 6 deals with the cytological analysis of the AD3 protein, an AMV MP-binding plant protein of which the expression could be verified in plant tissue. Specific antibodies were raised against heterologously expressed AD3 and used in immuno-cytological localisation studies. These studies revealed that AD3 is specifically found in membrane fractions of both leaf and root tissues of N. benthamiana , and immuno-gold EM demonstrated its localisation in the plasma membrane, which is not in contradiction with a potential function of this protein to support intercellular movement of viruses. Probably due to the low expression levels of AD3 and the transient presence of AMV MP solely at the infection front, a co-localisation between these two proteins in situ could not be established</p

Risk and resilience: health inequalities, working conditions and sickness benefit arrangements: an analysis of the 2010 European Working Conditions survey

In this article we ask whether the level of sickness benefit provision protects the health of employees, particularly those who are most exposed to hazardous working conditions or who have a little education. The study uses the European Working Condition Survey that includes information on 20,626 individuals from 28 countries. Health was measured by self-reported mental wellbeing and self-rated general health. Country-level sickness benefit provision was constructed using spending data from Eurostat. Group-specific associations were fitted using cross-level interaction terms between sickness benefit provision and physical and psychosocial working conditions respectively, as well as those with little education. The mental wellbeing of employees exposed to psychosocial job strain and physical hazards, or who had little education, was better in countries that offer more generous sickness benefit. These results were found in both men and women and were robust to the inclusion of GDP and country fixed effects. In the analyses of self-reported general health, few group-specific associations were found. This article concludes that generous sickness benefit provision may strengthen employee's resilience against mental health risks at work and risks associated with little education. Consequently, in countries with a generous provision of sickness benefit, social inequalities in mental health are smaller

Gustatory responses in primates to the sweetener aspartame and their phylogenetic implications

Two-bottle preference tests have been applied to 70 (sub-) species of the order of Primates and, for comparison, to the tree shrew (Tupaia belangeri) to determine their responses to aspartame (APM), the first known sweet-tasting dipeptide which has to man a sweetness potency of about 200 times that of sucrose. It was found that only the Cercopithecidae, the Hylobatidae and the Pongidae respond like man to this dipeptide and prefer it to water. The other primates tested to date, show no response to this sweetener. From a phylogenetic point of view, we note that APM shows species specificity similar to thaumatin. Thus, a clear dichotomy exists within the order of Primates with respect to both thaumatin and APM. The results here illustrate once more the gustatory diversity among primates and are a compelling argument for the existence of different sweet taste receptors or recognition sites in primate specie

On the dearth of compact, massive, red sequence galaxies in the local universe

We set out to test the claim that the recently identified population of compact, massive, and quiescent galaxies at z similar to 2.3 must undergo significant size evolution to match the properties of galaxies found in the local universe. Using data from the Sloan Digital Sky Survey (SDSS; Data Release 7), we have conducted a search for local red sequence galaxies with sizes and masses comparable to those found at z similar to 2.3. The SDSS spectroscopic target selection algorithm excludes high surface brightness objects; we show that this makes incompleteness a concern for such massive, compact galaxies, particularly for low redshifts (z less than or similar to 0.05). We have identified 63 M(*) > 10(10.7) M(circle dot) (approximate to 5 x 10(10) M(circle dot)) red sequence galaxies at 0.066 < z(spec) < 0.12 which are smaller than the median size mass relation by a factor of 2 or more. Consistent with expectations from the virial theorem, the median offset from the mass velocity dispersion relation for these galaxies is 0.12 dex. We do not, however, find any galaxies with sizes and masses comparable to those observed at z similar to 2.3, implying a decrease in the comoving number density of these galaxies, at fixed size and mass, by a factor of greater than or similar to 5000. This result cannot be explained by incompleteness: in the 0.066 < z < 0.12 interval, we estimate that the SDSS spectroscopic sample should typically be greater than or similar to 75% complete for galaxies with the sizes and masses seen at high redshift, although for the very smallest galaxies it may be as low as similar to 20%. In order to confirm that the absence of such compact massive galaxies in SDSS is not produced by spectroscopic selection effects, we have also looked for such galaxies in the basic SDSS photometric catalog, using photometric redshifts. While we do find signs of a slight bias against massive, compact galaxies, this analysis suggests that the SDSS spectroscopic sample is missing at most a few objects in the regime we consider. Accepting the high-redshift results, it is clear that massive galaxies must undergo significant structural evolution over z less than or similar to 2 in order to match the population seen in the local universe. Our results suggest that a highly stochastic mechanism (e.g., major mergers) cannot be the primary driver of this strong size evolution

An Over-Massive Black Hole in the Compact Lenticular Galaxy NGC1277

All massive galaxies likely have supermassive black holes at their centers, and the masses of the black holes are known to correlate with properties of the host galaxy bulge component. Several explanations have been proposed for the existence of these locally-established empirical relationships; they include the non-causal, statistical process of galaxy-galaxy merging, direct feedback between the black hole and its host galaxy, or galaxy-galaxy merging and the subsequent violent relaxation and dissipation. The empirical scaling relations are thus important for distinguishing between various theoretical models of galaxy evolution, and they further form the basis for all black hole mass measurements at large distances. In particular, observations have shown that the mass of the black hole is typically 0.1% of the stellar bulge mass of the galaxy. The small galaxy NGC4486B currently has the largest published fraction of its mass in a black hole at 11%. Here we report observations of the stellar kinematics of NGC 1277, which is a compact, disky galaxy with a mass of 1.2 x 10^11 Msun. From the data, we determine that the mass of the central black hole is 1.7 x 10^10 Msun, or 59% its bulge mass. Five other compact galaxies have properties similar to NGC 1277 and therefore may also contain over-sized black holes. It is not yet known if these galaxies represent a tail of a distribution, or if disk-dominated galaxies fail to follow the normal black hole mass scaling relations.Comment: 7 pages. 6 figures. Nature. Animation at http://www.mpia.de/~bosch/blackholes.htm

Characterization and temperature dependence of Arctic Micromonas polaris viruses

Global climate change-induced warming of the Artic seas is predicted to shift the phytoplankton community towards dominance of smaller-sized species due to global warming. Yet, little is known about their viral mortality agents despite the ecological importance of viruses regulating phytoplankton host dynamics and diversity. Here we report the isolation and basic characterization of four prasinoviruses infectious to the common Arctic picophytoplankter Micromonas. We furthermore assessed how temperature influenced viral infectivity and production. Phylogenetic analysis indicated that the putative double-stranded DNA (dsDNA) Micromonas polaris viruses (MpoVs) are prasinoviruses (Phycodnaviridae) of approximately 120 nm in particle size. One MpoV showed intrinsic differences to the other three viruses, i.e., larger genome size (205 ± 2 vs. 191 ± 3 Kb), broader host range, and longer latent period (39 vs. 18 h). Temperature increase shortened the latent periods (up to 50%), increased the burst size (up to 40%), and affected viral infectivity. However, the variability in response to temperature was high for the different viruses and host strains assessed, likely affecting the Arctic picoeukaryote community structure both in the short term (seasonal cycles) and long term (global warming)

Gcase and limp2 abnormalities in the liver of niemann pick type c mice

Funding Information: This work was supported by the NWO-Building Blocks of Life: GlcCer grant to J.M.F.G.A: BBOL-2007247202. Publisher Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland.The lysosomal storage disease Niemann–Pick type C (NPC) is caused by impaired cholesterol efflux from lysosomes, which is accompanied by secondary lysosomal accumulation of sph-ingomyelin and glucosylceramide (GlcCer). Similar to Gaucher disease (GD), patients deficient in glucocerebrosidase (GCase) degrading GlcCer, NPC patients show an elevated glucosylsphingosine and glucosylated cholesterol. In livers of mice lacking the lysosomal cholesterol efflux transporter NPC1, we investigated the expression of established biomarkers of lipid-laden macrophages of GD patients, their GCase status, and content on the cytosol facing glucosylceramidase GBA2 and lysoso-mal integral membrane protein type B (LIMP2), a transporter of newly formed GCase to lysosomes. Livers of 80-week-old Npc1−/− mice showed a partially reduced GCase protein and enzymatic activity. In contrast, GBA2 levels tended to be reciprocally increased with the GCase deficiency. In Npc1−/− liver, increased expression of lysosomal enzymes (cathepsin D, acid ceramidase) was observed as well as increased markers of lipid-stressed macrophages (GPNMB and galectin-3). Im-munohistochemistry showed that the latter markers are expressed by lipid laden Kupffer cells. Earlier reported increase of LIMP2 in Npc1−/− liver was confirmed. Unexpectedly, immunohistochemistry showed that LIMP2 is particularly overexpressed in the hepatocytes of the Npc1−/− liver. LIMP2 in these hepatocytes seems not to only localize to (endo)lysosomes. The recent recognition that LIMP2 harbors a cholesterol channel prompts the speculation that LIMP2 in Npc1−/− hepatocytes might mediate export of cholesterol into the bile and thus protects the hepatocytes.publishersversionpublishe

CANDELS Observations of the Structural Properties and Evolution of Galaxies in a Cluster at z=1.62

We discuss the structural and morphological properties of galaxies in a z=1.62 proto-cluster using near-IR imaging data from Hubble Space Telescope Wide Field Camera 3 data of the Cosmic Assembly Near-IR Deep Extragalactic Legacy Survey (CANDELS). The cluster galaxies exhibit a clear color-morphology relation: galaxies with colors of quiescent stellar populations generally have morphologies consistent with spheroids, and galaxies with colors consistent with ongoing star formation have disk-like and irregular morphologies. The size distribution of the quiescent cluster galaxies shows a deficit of compact (< 1kpc), massive galaxies compared to CANDELS field galaxies at z=1.6. As a result the cluster quiescent galaxies have larger average effective sizes compared to field galaxies at fixed mass at greater than 90% significance. Combined with data from the literature, the size evolution of quiescent cluster galaxies is relatively slow from z~1.6 to the present, growing as (1+z)^(-0.6+/-0.1). If this result is generalizable, then it implies that physical processes associated with the denser cluster region seems to have caused accelerated size growth in quiescent galaxies prior to z=1.6 and slower subsequent growth at z<1.6 compared to galaxies in the lower density field. The quiescent cluster galaxies at z=1.6 have higher ellipticities compared to lower redshift samples at fixed mass, and their surface-brightness profiles suggest that they contain extended stellar disks. We argue the cluster galaxies require dissipationless (i.e., gas-poor or "dry") mergers to reorganize the disk material and to match the relations for ellipticity, stellar mass, size, and color of early-type galaxies in z<1 clusters.Comment: Accepted for publication in ApJ. 14 pages in emulateapj format. Replacement includes improvements from referee report, and updates and additions to reference

The relationship between galaxy and dark matter halo size from z ∼ 3 to the present

We explore empirical constraints on the statistical relationship between the radial size of galaxies and the radius of their host dark matter haloes from z similar to 0.1-3 using the Galaxy And Mass Assembly (GAMA) and Cosmic Assembly Near Infrared Deep Extragalactic Legacy Survey (CANDELS) surveys. We map dark matter halo mass to galaxy stellar mass using relationships from abundance matching, applied to the Bolshoi-Planck dissipationless N-body simulation. We define SRHR equivalent to r(e)/R-h as the ratio of galaxy radius to halo virial radius, and SRHR lambda equivalent to r(e)/(lambda R-h) as the ratio of galaxy radius to halo spin parameter times halo radius. At z similar to 0.1, we find an average value of SRHR similar or equal to 0.018 and SRHR. similar or equal to 0.5 with very little dependence on stellar mass. Stellar radius-halo radius (SRHR) and SRHR lambda have a weak dependence on cosmic time since z similar to 3. SRHR shows a mild decrease over cosmic time for low-mass galaxies, but increases slightly or does not evolve formoremassive galaxies. We find hints that at high redshift (z similar to 2-3), SRHR. is lower for more massive galaxies, while it shows no significant dependence on stellar mass at z less than or similar to 0.5. We find that for both the GAMA and CANDELS samples, at all redshifts from z similar to 0.1-3, the observed conditional size distribution in stellar mass bins is remarkably similar to the conditional distribution of lambda R-h. We discuss the physical interpretation and implications of these results

Discovery of a Quadruple Lens in CANDELS with a Record Lens Redshift z=1.53

Using spectroscopy from the Large Binocular Telescope and imaging from the Hubble Space Telescope we discovered the first strong galaxy lens at z(lens)>1. The lens has a secure photometric redshift of z=1.53+/-0.09 and the source is spectroscopically confirmed at z=3.417. The Einstein radius (0.35"; 3.0 kpc) encloses 7.6 x 10^10 Msol, with an upper limit on the dark matter fraction of 60%. The highly magnified (40x) source galaxy has a very small stellar mass (~10^8 Msol) and shows an extremely strong [OIII]_5007A emission line (EW_0 ~ 1000A) bolstering the evidence that intense starbursts among very low-mass galaxies are common at high redshift.Comment: accepted for publication in ApJ Letter