Journal of Biological Research-Thessaloniki最新文献

Background: Cortical microtubules regulate cell expansion by determining cellulose microfibril orientation in the root apex of Arabidopsis thaliana. While the regulation of cell wall properties by cortical microtubules is well studied, the data on the influence of cell wall to cortical microtubule organization and stability remain scarce. Studies on cellulose biosynthesis mutants revealed that cortical microtubules depend on Cellulose Synthase A (CESA) function and/or cell expansion. Furthermore, it has been reported that cortical microtubules in cellulose-deficient mutants are hypersensitive to oryzalin. In this work, the persistence of cortical microtubules against anti-microtubule treatment was thoroughly studied in the roots of several cesa mutants, namely thanatos, mre1, any1, prc1-1 and rsw1, and the Cellulose Synthase Interacting 1 protein (csi1) mutant pom2-4. In addition, various treatments with drugs affecting cell expansion were performed on wild-type roots. Whole mount tubulin immunolabeling was applied in the above roots and observations were performed by confocal microscopy.

Results: Cortical microtubules in all mutants showed statistically significant increased persistence against anti-microtubule drugs, compared to those of the wild-type. Furthermore, to examine if the enhanced stability of cortical microtubules was due to reduced cellulose biosynthesis or to suppression of cell expansion, treatments of wild-type roots with 2,6-dichlorobenzonitrile (DCB) and Congo red were performed. After these treatments, cortical microtubules appeared more resistant to oryzalin, than in the control.

Conclusions: According to these findings, it may be concluded that inhibition of cell expansion, irrespective of the cause, results in increased microtubule stability in A. thaliana root. In addition, cell expansion does not only rely on cortical microtubule orientation but also plays a regulatory role in microtubule dynamics, as well. Various hypotheses may explain the increased cortical microtubule stability under decreased cell expansion such as the role of cell wall sensors and the presence of less dynamic cortical microtubules.

Background: MADS-box transcription factors function as homo- or heterodimers and regulate many aspects of plant development; moreover, MADS-box genes have undergone extensive duplication and divergence. For example, the morphological diversity of floral organs is closely related to the functional divergence of the MADS-box gene family. B-class genes (such as Arabidopsis thaliana APETALA3 [AP3] and PISTILLATA [PI]) belong to a subgroup of MADS-box genes. Here, we collected 97 MADS-box B protein sequences from 21 seed plant species and examined their motifs to better understand the functional evolution of B proteins.

Results: We used the MEME tool to identify conserved sequence motifs in these B proteins; unique motif arrangements and sequences were identified in these B proteins. The keratin-like domains of Malus domestica and Populus trichocarpa B proteins differed from those in other angiosperms, suggesting that a novel regulatory network might have evolved in these species. The MADS domains of Nelumbo nucifera, Glycine max, and Amborella trichopoda B-proteins contained motif 9; in contrast, those of other plants contained motif 1. Protein modelling analyses revealed that MADS domains with motif 9 may lack amino acid sites required for DNA-binding. These results suggested that the three species might share an alternative mechanism controlling floral development.

Conclusions: Amborella trichopoda has B proteins with either motif 1 or motif 9 MADS domains, suggesting that these two types of MADS domains evolved from the ancestral domain into two groups, those with motif 9 (N. nucifera and G. max), and those with motif 1. Moreover, our results suggest that the homodimer/heterodimer intermediate transition structure first appeared in A. trichopoda. Therefore, our systematic analysis of the motifs in B proteins sheds light on the evolution of these important transcription factors.

Background: The Indo-Pacific sea urchin Diadema setosum has invaded the Mediterranean Sea and has spread along many locations in the southeastern part of the basin, where established populations exist on the shallow subtidal rocky shore. Diadema setosum is a ubiquitous species, of particular ecological importance due to the high levels of grazing pressure it imposes on benthic communities. Its biology, however, is not adequately studied, especially along its introduced range of distribution. The present study examines the population status of D. setosum outside its native range, in the Dodecanese island complex, south Aegean Sea. Thirty-four stations located across 16 islands were surveyed by scientific SCUBA-diving (up to a depth of 10 m) in December 2019 and June-July 2020. Samplings included: (i) visual census along transects to estimate relative abundance and population density, and (ii) random collection of specimens from densely populated stations to assess biometry and reproductive condition (histological examination of gonads) of D. setosum.

Results: Diadema setosum was found in 21 out of the 34 surveyed stations. The species had sparse populations of well-hidden individuals in rocky crevices, but with dense localized patches in Agathonisi, Leros, Kalymnos, Pserimos, Symi, Alimia and Chalki islands. In those seven islands, mean population density was 2.5 ± 1.48 individuals m^-2. Diadema setosum had denser populations in shallower depths but larger dimensions in deeper; these results suggest segregated density and size patterns along a depth gradient. The size structure, according to the size frequency distribution of the test diameter, was unimodal with a fitted mode at 4.0-4.5 and 6.5-7.0 cm in shallow and deep populations, respectively. The examined morphometric relationships followed negative allometry, as previously suggested for the species within its native range of distribution, and test diameter appeared to be a good predictor of biomass. Diadema setosum specimens had immature gonads in winter and mature in summer, suggesting a synchronous reproductive pattern. These results conform to previous data from temperate populations of the species.

Conclusions: Differences in local environmental conditions, e.g. hydrodynamics and habitat type, together with biotic interactions, e.g. recruitment and competition, probably shape D. setosum population in the south Aegean distributional range. The establishment of D. setosum has severe implications on benthic communities and local sea urchin populations demanding management measures to prevent the forecasted further expansion of this invasive species.

Background: Lipoxygenases are one of the critical signaling mediators which can be targeted for human prostate cancer (PC) therapy. In this study, 4-methyl-2-(4-methylpiperazinyl)pyrimido[4,5-b]benzothiazine (4-MMPB) and its two analogs, 4-propyl-2-(4-methylpiperazinyl)pyrimido[4,5-b]benzothiazine (4-PMPB) and 4-ethyl-2-(4-methylpiperazinyl)pyrimido[4,5-b]benzothiazine (4-EMPB), were proposed to have anti-tumor properties in prostate cancer.

Methods: After synthesizing the compounds, cytotoxic effects of 4-MMPB and its two analogs against PC-3 cancerous and HDF normal cells were investigated by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay and then mechanism of cell death was assessed by flow cytometry. Finally, the anti-tumor effects of the mentioned compounds were investigated in an immunocompromised C57BL/6 mouse model.

Results: 4-PMPB and 4-EMPB had similar anti-cancer effects on PC-3 cells as compared with 4-MMPB, while they were not effective on normal cells. Moreover, apoptosis and ferroptosis were the main mechanisms of induced cell death in these cancerous cells. Furthermore, in vivo results indicated that both analogs had similar anti-cancer effects as 4-MMPB, leading to delayed tumor growth without any noticeable side effects in weight loss and histological investigations.

Conclusion: Thus, our results suggest that specific targeting of lipoxygenases via 4-MMPB analogs can be considered as a treatment of choice for PC therapy, although it requires further investigations.

Background: Somatic embryogenesis is a phenomenon carried out in an environment that generates abiotic stress. Thus, regenerants may differ from the source of explants at the morphological, genetic, and epigenetic levels. The DNA changes may be the outcome of induction media ingredients (i.e., copper and silver ions) and their concentrations and time of in vitro cultures.

Results: This study optimised the level of copper and silver ion concentration in culture media parallel with the induction medium longevity step towards obtaining barley regenerants via somatic embryogenesis with a minimum or maximum level of tissue culture-induced differences between the donor plant and its regenerants. The optimisation process is based on tissue culture-induced variation evaluated via the metAFLP approach for regenerants derived under varying in vitro tissue culture conditions and exploited by the Taguchi method. In the optimisation and verification experiments, various copper and silver ion concentrations and the different number of days differentiated the tested trials concerning the tissue culture-induced variation level, DNA demethylation, and de novo methylation, including symmetric (CG, CHG) and asymmetric (CHH) DNA sequence contexts. Verification of optimised conditions towards obtaining regenerants with minimum and maximum variability compared to donor plants proved useful. The main changes that discriminate optimised conditions belonged to DNA demethylation events with particular stress on CHG context.

Conclusions: The combination of tissue culture-induced variation evaluated for eight experimental trials and implementation of the Taguchi method allowed the optimisation of the in vitro tissue culture conditions towards the minimum and maximum differences between a source of tissue explants (donor plant) and its regenerants from somatic embryos. The tissue culture-induced variation characteristic is mostly affected by demethylation with preferences towards CHG sequence context.

Background: Microcystins are emerging marine biotoxins, produced by potentially toxic cyanobacteria. Their presence has been reported in aquatic animals in Greek freshwater, while data are few in marine environments. Since the climate change induces eutrophication and harmful algal blooms in coastal marine ecosystems affecting the public health, further research on microcystins' presence in marine waters is required. The aim of this study was to examine the potential presence of microcystins in mussels Mytilus galloprovincialis in the largest farming areas in Thermaikos gulf, in Northern Greece, and to investigate their temporal and spatial distribution, adding to the knowledge of microcystins presence in Greek Mediterranean mussels.

Results: A 4-year microcystins' assessment was conducted from 2013 to 2016, in farmed Mediterranean mussels M. galloprovincialis, in five sampling areas in Thermaikos gulf, in northern Greece, where the 90% of the Greek mussels' farming activities is located. The isolation of potentially toxic cyanobacteria was confirmed by molecular methods. An initial screening was performed with a qualitative and quantitative direct monoclonal (DM) ELISA and results above 1 ng g^-1 were confirmed for the occurrence of the most common microcystins-RR, -LR and -YR, by Ultra High Performance Liquid Chromatography (UHPLC) coupled with a high- resolution mass spectrometer (HRMS) (Orbitrap analyzer). Microcystin-RR and microcystin-LR were detected, while the intensity of microcystin-YR was below the method detection limit. Most samples that exhibited concentrations above 1 ng g^-1 were detected during the warm seasons of the year and especially in spring. Results indicated an overestimation of the ELISA method, since concentrations ranged between 0.70 ± 0.15 ng g^-1 and 53.90 ± 3.18 ng g^-1, while the confirmation denoted that the levels of microcystins were 6 to 22 times lower.

Conclusions: Microcystin-RR and microcystin-LR were detected for the first time in mussel M. galloprovincialis, harvested from farms in Thermaikos gulf, in Central Macedonia, Greece. Their presence was linked to potentially toxic cyanobacteria. Bioaccumulation was observed in digestive gland, while the concentrations in muscles were found extremely low. Samples with levels above 1 ng g^-1 were observed mostly during spring, confirming the seasonal distribution of microcystins. The comparison of the results by the ELISA and the LC-Orbitrap MS method indicated an overestimation of concentration by the ELISA method.

Background: Circular RNAs (circRNA) have been shown to be involved in the pathogenesis of colorectal cancer (CRC). CircCTNNA1 was found to be one of the upregulated circRNAs in CRC. However, there are few studies on circCTNNA1, so it is necessary to carry out further studies.

Methods: The expression of circCTNNA1, microRNA (miR)-363-3p, and chemokine C-X-C motif ligand 5 (CXCL5) was detected by quantitative real-time PCR (qRT-PCR). The protein levels of CXCL5 and metastasis markers were measured using western blot (WB) analysis. Cell proliferation, apoptosis, cell cycle, migration, and invasion were determined by cell counting kit 8 (CCK8) assay, colony formation assay, flow cytometry, and transwell assay. The relationship between miR-363-3p and circCTNNA1 or CXCL5 was evaluated via dual-luciferase reporter assay and RNA immunoprecipitation assay. Animal study was performed to explore the function of circCTNNA1 on CRC tumorigenesis.

Results: CircCTNNA1 and CXCL5 were highly expressed in CRC. Knockdown of circCTNNA1 could inhibit the proliferation, cell cycle, metastasis, and promote the apoptosis of CRC cells. MiR-363-3p could be sponged by circCTNNA1, and the inhibition effect of circCTNNA1 silencing on CRC progression could be reversed by miR-363-3p inhibitor. Moreover, miR-363-3p could interact with CXCL5, and CXCL5 overexpression also could reverse the suppressive effect of miR-363-3p on CRC progression. Downregulation of circCTNNA1 also could hinder the tumor growth of CRC in vivo.

Conclusion: CircCTNNA1 enhanced CRC progression via regulating the miR-363-3p/CXCL5 axis.

Background: Parkinson's disease (PD) is the second most common neurodegenerative disease after Alzheimer's disease. The oxidative stress is an important component of the pathogenesis of PD. Artemisinin (ART) has antioxidant and neuroprotective effects. The purpose of this study is to explore the neuroprotective effect of ART on 1-methyl-4-phenyliodine iodide (MPP ⁺)-treated SH-SY5Y cells and underlying mechanism.

Methods: We used MPP⁺-treated SH-SY5Y cells to study the neuroprotective effect of ART. Cell viability was measured by MTT assay after incubating the cells with MPP⁺ and/or ART for 24 h. DCFH-DA was used to detect the level of intracellular reactive oxygen species (ROS), and WST-8 was used to detect the level of superoxide dismutase (SOD). The level of intracellular reduced glutathione (GSH) was detected with 5,5΄-dithiobis-(2-nitrobenzoic acid), and the level of malondialdehyde (MDA) was assessed based on the reaction of MDA and thiobarbituric acid. A mitochondrial membrane potential detection kit (JC-1) was used to detect changes in the mitochondrial membrane potential (MMP), and an Annexin V-FITC cell apoptosis kit was used to detect cell apoptosis. The expression levels of caspase-3, cleaved caspase-3 and the autophagy-related proteins LC3, beclin-1, and p62 were detected by Western blotting. In addition, to verify the change in autophagy, we used immunofluorescence to detect the expression of LC3 and p62.

Results: No significant cytotoxicity was observed at ART concentrations up to 40 μM. ART could significantly increase the viability of SH-SY5Y cells treated with MPP⁺ and reduce oxidative stress damage and apoptosis. In addition, the Western blotting and immunofluorescence results showed that MPP⁺ treatment could increase the protein expression of beclin1 and LC3II/LC3I and decrease the protein expression of p62, indicating that MPP⁺ treatment could induce autophagy. Simultaneous treatment with ART and MPP⁺ could decrease the protein expression of beclin1 and LC3II/LC3I and increase the protein expression of p62, indicating that ART could decrease the level of autophagy induced by MPP⁺.

Conclusion: Our results indicate that ART has a protective effect on MPP⁺-treated SH-SY5Y cells by the antioxidant, antiapoptotic activities and inhibition of autophagy. Our findings may provide new hope for the prevention and treatment of PD.

Background: Because of the highly heterogeneous nature of breast cancer, each subtype differs in response to several treatment regimens. This has limited the therapeutic options for metastatic breast cancer disease requiring exploration of diverse therapeutic models to target tumor specific biomarkers.

Methods: Differentially expressed breast cancer genes identified through extensive data mapping were studied for their interaction with other target proteins involved in breast cancer progression. The molecular mechanisms by which these signature genes are involved in breast cancer metastasis were also studied through pathway analysis. The potential drug targets for these genes were also identified.

Results: From 50 DEGs, 20 genes were identified based on fold change and p-value and the data curation of these genes helped in shortlisting 8 potential gene signatures that can be used as potential candidates for breast cancer. Their network and pathway analysis clarified the role of these genes in breast cancer and their interaction with other signaling pathways involved in the progression of disease metastasis. The miRNA targets identified through miRDB predictor provided potential miRNA targets for these genes that can be involved in breast cancer progression. Several FDA approved drug targets were identified for the signature genes easing the therapeutic options for breast cancer treatment.

Conclusion: The study provides a more clarified role of signature genes, their interaction with other genes as well as signaling pathways. The miRNA prediction and the potential drugs identified will aid in assessing the role of these targets in breast cancer.

Background: One third of the worldwide amphibian species are threatened, therefore, efficient monitoring efforts are needed. Amphibians which adopt a hidden lifestyle, such as the common spadefoot toad, are often missed with standard surveying efforts. Spadefoot toads can be identified in regurgitated pellets of the barn owl, which provides an effective way to estimate toad activity. In our study we analyzed frequency of spadefoot toad remains from 2004 to 2016 in a steppe landscape in eastern Austria.

Methods: We used an automated model selection procedure together with a GLM analysis using a zero inflated error Poisson distribution, to analyze the presence of Pelobates fuscus in barn owl pellets. All analyses were done in the statistical software R, and the scripts to reproduce our results are available within this publication. Our approach may provide a template for other researchers to use for their own pellet data.

Conclusions: Our analysis suggested that activity of the common spadefoot toad is mainly influenced by rainfalls, while time of the year and temperature had small but significant effects. Interestingly, our data confirmed the possibility of a second breeding period in summer, triggered by heavy rainfalls. There were no indications for a population decrease in the observed years and locations. Our study shows that barn owl pellets can be used effectivley to assess pelobatid activity in an area. This might constitute a useful monitoring tool for conservation management for amphibians.