Biochemical and cytological interactions between callose synthase and microtubules in the tobacco pollen tube

Key message: The article concerns the association between callose synthase and cytoskeleton by biochemical and ultrastructural analyses in the pollen tube. Results confirmed this association and immunogold labeling showed a colocalization. Abstract: Callose is a cell wall polysaccharide involved in fundamental biological processes, from plant development to the response to abiotic and biotic stress. To gain insight into the deposition pattern of callose, it is important to know how the enzyme callose synthase is regulated through the interaction with the vesicle-cytoskeletal system. Actin filaments likely determine the long-range distribution of callose synthase through transport vesicles but the spatial/biochemical relationships between callose synthase and microtubules are poorly understood, although experimental evidence supports the association between callose synthase and tubulin. In this manuscript, we further investigated the association between callose synthase and microtubules through biochemical and ultrastructural analyses in the pollen tube model system, where callose is an essential component of the cell wall. Results by native 2-D electrophoresis, isolation of callose synthase complex and far-western blot confirmed that callose synthase is associated with tubulin and can therefore interface with cortical microtubules. In contrast, actin and sucrose synthase were not permanently associated with callose synthase. Immunogold labeling showed colocalization between the enzyme and microtubules, occasionally mediated by vesicles. Overall, the data indicate that pollen tube callose synthase exerts its activity in cooperation with the microtubular cytoskeleton

Influence of Televised Crime news on Audience Actions

Most television newscasts often relay crime stories with detailed descriptions and vivid images. Literally, the media set the agenda of what the audience should pay attention to and think about by selecting crime stories, the audiences are however not usually told how to think about those selected stories. This could mean that the degree to which exposure to televised crime news stories influences audience perception about crime is not explicit and therefore not clear. This study aimed at documenting audience perception of prevalent crime topics in televised news content in order to interpret the actions/behaviour of the audiences. This paper presents results obtained from a qualitative study undertaken in Milimani geographical area of Nakuru town. The study was guided by McCombs and Shaw’s Agenda Setting and Gerbner’s Media Effects theories to collect and analyse data. The population comprised all categories of people residing, working or learning within Milimani area of Nakuru town. The total sample was 30 participants, an acceptable size for a qualitative study. The results show that different viewers react in varied ways to televised crime news and consequently act differently in varied contexts. This is an indicator that televised crime news influence audience behaviour and consequently, affirming the influence of news bulletins on audience perception of social issues, and in the process, they influence the audience with what to think about because of the emotions they evoke. The question lingers: is televised crime news useful? How much can the journalist reveal? How do audience actions affect the journalists work

EFFECTS OF ACUTE HEAT STRESS DURING ANTHER AND POLLEN DEVELOPMENT IN LYCOPERSICON ESCULENTUM CV MICRO-TOM

The exposure of plants to extreme temperatures affects their geographic distribution and their yield quality and quantity (1). One of the most sensitive process to high temperature is plant reproduction, particularly the male apparatus (2). Thanks to its small size, rapid growth and easy transformation, Lycopersicon esculentum cv Micro- Tom has been proposed as one of the preferred variety of tomato plants to carry out molecular research (3). Although it is widely used as model organism, its use is quite recent and little is known about its response to heat stress. Plants at three flower bud stages: meiotic stage (2 mm long), microspore stage (4 mm long) and mature stage (6 mm long) were stressed with high temperature (42 °C for 3 hours) to study the effects on mature anthers. Investigations by light microscopy were carried out with three different stains: Alcian Blue, Toluidine Blue O and Periodic Acid Schiff. The only remarkable difference in morphological features was the absence of starch in cells of anthers treated at microspore stage. Pressman (4) reported that in tomato high temperature inhibited starch deposition in developing pollen, either through a decrease in the availability of assimilates or through the impairment of the activities of enzymes involved in starch biosynthesis or in sucrose hydrolysis, namely invertase and sucrose synthase (SuS). The immunolocalization evidenced the absence of the SuS enzyme in mature anthers treated at microspore stage, confirming that probably the HS interfered with the presence of SuS and starch. Scanning electron microscope observations revealed several alterations in shape and size of pollen grains. Pollen grains from anthers treated at the meiotic stage were reduced to collapsed exine coat; the majority of pollen grains from anthers treated at the microspore stage was crushed, whereas most of pollen grains treated at the mature stage was round in shape. Pollen viability and germinability tests showed that only pollen grains of mature treated anthers had a viability percentage similar to the control; the germination percentage of pollen grains of anthers treated at all developmental stages was extremely low. Molecular analysis on two heat shock factors (HSFA2 and HSFB3a) highlighted an attempt of HS response by treated anthers, especially for HSFA2, which showed a significant increase of expression after HS exposure. The severe physiological alterations caused by HS irreversibly affected the male apparatus of Micro-Tom plants, thus impairing its reproductive performance

EFFECTS OF ACUTE HEAT STRESS ON THE FEMALE REPRODUCTIVE APPARATUS OF LYCOPERSICON ESCULENTUM CV MICRO-TOM

Temperatures above the normal optimum are sensed as heat stress (HS) by all living organisms, including plants. HS affects fertility, yield and production quality of crops with important consequences on food security and the well-being of human population. These negative effects can depend on the fact that all the phases of sexual reproduction in angiosperms, namely gametophyte development (from meiosis to pollination), progamic phase (from pollination to zygote formation) and embryo development (from zygote to seed), are more vulnerable to temperature stress than vegetative processes (1). Presently there are limited data about the effects of HS on the female part (pistil) and its interaction with pollen grains. The present research gives more insight into the effects of acute HS on the pistil's morphology and the progamic phase of tomato (Lycopersicon esculentum cv Micro-Tom). A morphological analysis of Micro-Tom pistils treated at 42°C for 3 h at different developmental stages has been performed by light microscopy using Toluidine Blue O, Periodic Acid Schiff, Alcian Blue and Calcofluor White stains. Moreover, the pattern distribution of α-tubulin has been investigated in the heat-treated pistils. In addition, the carbohydrate epitope of arabinogalactan proteins (AGPs), involved in pollen tube guidance to the ovary, has been localized by JIM8 antibody. The consequences of acute HS have been also evaluated on stigmatic receptivity, pollen performance and in vivo pollen tube growth using epifluorescence microscopy. Results herein reported show that the general organization of the pistil tissues is substantially unaltered under HS. A significant observation is that HS seems to alter the microtubule distribution in the ovules: in control ovules, microtubules resulted well preserved in the inner integument tissue, whereas in treated ovules they were diffused in the cytoplasm. The epitope immunorecognized by JIM8 antibody seemed to disappear under HS in the transmitting tissue of the styles, thus denoting a possible alteration in the pollen tube growth pathway; on the contrary its localization was similar to the control in heat-treated ovules. Both stigma receptivity and pollen performance resulted sensitive to HS. The present work confirmed the sensitivity of sexual plant reproduction to HS: like in other species, such as peach (2) and sweet cherry (3), the receptivity of Micro-Tom stigma was reduced under HS and thus, the possibility of fertilization. Molecules involved in pollen tube guidance to the ovary (AGPs) and structural proteins (microtubules) showed an altered pattern with possible functional consequences on pistil function

Group membership and adolescents' alcohol intake: The mediating role of drinking motives

Objective: Group norms and drinking motives are crucial predictors of adolescents’ alcohol intake. The current study examined the role of drinking motives in the association between descriptive group norms and alcohol intake. Method: A sample of 525 Italian adolescents (56% men) was surveyed. Participants completed measures of group norms, drinking motives, and personal drinking. Results: Hierarchical multiple regression analysis revealed that stronger group norms were positively related to increased alcohol intake. Drinking for enhancement and social motives also were related to increased alcohol intake. Mediation analysis showed that group norms were related to alcohol intake through social and enhancement drinking motives. Conclusions: Drinking motives provide fruitful insights into the relationship between descriptive group norms and excessive alcohol use among adolescents. Implications for future research are discussed

Temperature-Dependent Compatible and Incompatible Pollen-Style Interactions in Citrus clementina Hort. ex Tan. Show Different Transglutaminase Features and Polyamine Pattern

In clementine, failure of fertilization can result in parthenocarpic fruit development, which has several advantages, such as seedless fruit, longer shelf-life, and greater consumer appeal. Recently, S-RNases have been identified in Citrus grandis, thus revealing that the self-incompatibility (SI) reaction relies on the S-RNase gametophytic mechanism. The fundamental role of environmental factors, mostly temperature, in determining the numbers of pollen tubes reaching the ovary is also well established in Citrus. In the present work, temperature-dependent pollen\u2013pistil interactions in C. clementina were analyzed, focusing on several morphological aspects, as well as on polyamine (PA) content and the activity and distribution of transglutaminase (TGase), both reported to be involved in the SI response in pear and in pummelo. Results clearly indicate that temperature contributed to a different activation of the SI response, which occurs at optimal temperature of 25\ub0C but was by-passed at 15\ub0C. TGase activity was stimulated during the SI response, and it localized differently in the compatible and incompatible interaction: in compatible pollinated styles, TGase localized inside the style canal, while it was detected all around it in incompatible crosses. TGase localization and activity were congruent with the levels of soluble and insoluble conjugated PAs and with morphological evidences, which highlighted cell wall modification occurring as a result of SI

Genetic basis of fr-h2 in barley (Hordeum vulgare L.): significant improvements in the understanding of the “CBF number game”

Freezing temperatures can severely compromise barley yield potential. Freezing tolerance depends on a period of cold acclimation mediated by CBFs (C-repeat binding factors), a class of transcriptional factors that modulate the expression of multiple effector genes. In order to investigate the relationship between frost tolerance effect due to the major QTL Fr-H2 and copy number variation (CNV) for a specific cluster of CBF genes, a qPCR approach was used on a contrasting set of barley cultivars. The panel, composed of 41 genotypes with different growth habits, was accurately phenotyped by field survival and Fv/Fm measurements. Data showed that there was a positive correlation between frost tolerance and CNV for the HvCBF2-HvCBF4 genomic region in the proximal cluster and a lower yet significant negative correlation between CNV for HvCBF3 in the distal cluster and frost tolerance. These results strongly support the hypothesis that Fr-H2 effect is due to a sort of“HvCBF-number game” rather than to a CNV at a single HvCBF gene. In addition to this, a protocol to discriminate the HvCBF2 forms showed an increased copy number of HvCBF2A form in highly resistant facultative and winter genotypes. Resequencing by third generation sequencing technology (PacBio) of Fr-H2 in the frost tolerant cv. ‘Nure’ is in progress. A comparison of the region with the structure of the locus in both the susceptible reference genotype ‘Morex’ and the wheat genomes allowed a detailed characterization of the genic and intergenic space of the proximal subcluster harbouring HvCBF4 and HvCBF2. The obtained results are being validated by detailed expression analysis aimed to define how each gene involved in CNV contribute in terms of CBF transcripts accumulation. These results represent a significant step forward the understanding of the genetic bases of frost tolerance in barley