imagine (Institute of molecular genetics and genetic engineering)
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The long non-coding RNA GAS5 contributes to the suppression of inflammatory responses by inhibiting NF-κB activity
No full textIntroductionNuclear factor kappa B (NF-κB) is a key regulator of immune and inflammatory responses. Glucocorticoid drugs (GC) act through the glucocorticoid receptor (GR) as immunosuppressant also in pediatric patients inhibiting NF-κB activity. The long non-coding RNA GAS5 interacts with the GR, influencing GC activity. No data on the role of GAS5 on GR-dependent inhibition of NF-κB activity have been published.MethodsThis study investigated the impact of GAS5 on NF-κB activity in HeLa cells overexpressing GAS5, both under basal conditions and during GC treatment. The study used EMSA, RNA-immunoprecipitation (RIP), Western blotting, and bioinformatic analyses to assess NF-κB DNA binding, GAS5-p65 interaction, and NF-κB signaling pathway modulation.ResultsGAS5 overexpression increased NF-κB DNA binding activity in untreated cells. RNA-IP confirmed a direct interaction between GAS5 and the NF-κB subunit p65, suggesting a potential regulatory mechanism. GAS5 overexpression led to downregulation of NF-κB target genes, TNF-α, and NR3C1. GC treatment reduced NF-κB DNA binding activity in GAS5-overexpressing cells, indicating a potential synergistic effect. Furthermore, GAS5 overexpression increased IκB levels and reduced p-p65/pan-p65 levels during GC treatment.DiscussionGAS5 appears to modulate NF-κB activity in a complex manner, influencing both basal and GC-induced signaling. The interaction between GAS5, GCs, and NF-κB is multi-faceted, and further research is needed to fully elucidate the underlying mechanisms. These findings suggest that GAS5 could be a potential target for personalized therapy, particularly in pediatric patients with inflammatory conditions
Beyond 10,000 Ancient Human Genomes; Ancestral Origins at the Balkans
No full textWith more than 10,000 ancient human genomes published in 2023, thanks to new
technological developments on DNA sequencing, we are now able to investigate multiple
ancestry layers associated to past migrations that have shaped the genomes of modern
populations. These studies have been able to unravel past social structures, as well as
selective processes, that left genomic marks. In the Balkans, the recent analysis of some
hundreds of ancient genomes from the last three thousand years have uncovered the
genetic signals of globalisation during the Roman Empire and also the signals of the Slavic
migrations after the 6th century BCE. Getting into historical periods, these population
movements have strong cultural and even political implications, showing the complex
nature of ancestry, genetics and identity. Genetics can offer objective data on human past
and yet, their interpretation in terms of identity is complex. A multidisciplinary approach,
involving different disciplines such as archaeology, anthropology, history and even
linguistics is recommended.Book of abstracts: 5th Belgrade Bioinformatics Conference, Serbia, Belgrade,17-20 june 2024
Antimicrobial Activities of Essential Oils of Different Pinus Species from Bosnia and Herzegovina
No full textBackground/Objectives: The emergence of antimicrobial resistance has urged researchers to explore new antimicrobial agents, such as essential oils (EOs). The aim of this study was to examine chemical composition and antimicrobial activity of the EOs from the needles and green cones of four Pinus species (Pinus mugo Turra., P. nigra J.F., P. syilvestris L., and P. halepensis Miller) from Bosnia and Herzegovina. Methods: Chemical profiles of EOs were assessed by gas chromatography, while microdilution method was used to test their antimicrobial activity. A synergistic action of EOs and gentamicin was investigated by the checkerboard assay. Results: The chemical composition of the tested EOs showed a high percentage of α-pinene, (E)-caryophyllene, limonene, germacrene D, myrcene, and δ-3-carene. EO from green cones of P. sylvestris showed high efficiency against S. aureus and E. faecalis. The MIC of P. nigra cones’ EO was 100 μg/mL against E. coli. The EO of P. halepensis green cones demonstrated the strongest activity against E. faecalis. EOs of P. halepensis needles and green cones exhibited the highest activity against C. albicans. Further, synergistic interaction was detected in combination of the selected EOs/gentamicin toward S. aureus and K. pneumoniae. Conclusions: Among the tested EOs, oils of P. sylvestris cones and P. halepensis cones and needles showed the greatest antimicrobial activity. The same EOs and EO from P. nigra cones displayed synergistic potential in combination with gentamicin, supporting their utilization as antimicrobial agents alone or in combination with antibiotics, which is in line with their ethnopharmacological usage and circular bioeconomy principles
Multi-drug resistant Staphylococcus epidermidis from chronic wounds impair healing in human wound model
No full textVenous leg ulcers (VLUs) represent one of the most prevalent types of chronic wounds characterised by perturbed microbiome and biofilm-forming bacteria. As one of the most abundant skin-commensal, Staphylococcus epidermidis is known as beneficial for the host, however, some strains can form biofilms and hinder wound healing. In this study, S. epidermidis distribution in VLUs and associated resistome were analysed in ulcer tissue from patients. Virulence of S. epidermidis isolates from VLUs were evaluated by whole genome sequencing, antimicrobial susceptibility testing, in vitro biofilm and binding assays, and assessment of biofilm-forming capability and pro-inflammatory potential using human ex vivo wound model. We demonstrated that S. epidermidis isolates from VLUs inhibit re-epithelialization through biofilm-dependent induction of IL-1β, IL-8, and IL-6 which was in accordance with impaired healing outcomes observed in patients. High extracellular matrix binding ability of VLU isolates was associated with antimicrobial resistance and expression levels of the embp and sdrG, responsible for bacterial binding to fibrinogen and fibrin, respectively. Finally, we showed that S. epidermidis from VLUs demonstrate pathogenic features with ability to impair healing which underscores the emergence of treatment-resistant virulent lineages in patients with chronic ulcers
TESTING THE EFFECTS OF MOTHERS CURSE HYPOTHESIS ON TWO DROSOPHILA SPECIES
No full textThe Mother's Curse hypothesis proposes an accumulation of a male-specific genetic load within
the mitochondrial DNA (mtDNA). This is due to selection acting on mtDNA genes only in
females because of the uniparental mechanism of mitochondrial inheritance through mothers.
Consequently, harmful mutations that affect only males go unnoticed by selection, allowing them
to persist and even increase in frequency within the population through genetic drift or even
selection if a specific mutation has antagonistic effects between sexes.
While the theoretical framework of the Mother’s Curse hypothesis is straightforward, measuring
the effects of the Mother’s Curse in natural populations has proven challenging, yielding many
contradictory results. Most experiments that were aligned with the theory used specially
engineered introgression lines that were designed to maximise the effect of male-specific load,
minimizing the influence of compensatory nuclear mutations that could mitigate these effects.
Using two model species (Drosophila subobscura and D. obscura) we conducted life history
experiments with sympatric mito-nuclear lines. Measured traits were: desiccation resistance,
developmental time, egg-to-adult viability and percentage of males. For each component that
showed sex-specific differences, we tested the magnitude of the mitochondrial effects between
the sexes to test for signatures of the Mother’s Curse. Contrary to theoretical predictions, our
results in both model species revealed that mitochondrial variation had a bigger impact on female
fitness. These findings suggest a complex interplay between the two genomes and underscore the
need for further investigation into the relevance and magnitude of its effects in natural and
experimental populations.VII Congress of the Serbian Genetic Society Zlatibor; October 2 to 5, 2024
PHB AND PHB-PHO COMPOSITES WITH IMMOBILIZED Streptomyces DG19 SPORES FOR ENHANCED POLYMER BIODEGRADATION
No full textDue to its established biological carbon cycle, poly(hydroxybutyrate) (PHB) is
a promising substitute for petrochemical plastic. Although PHB is susceptible to
microbial degradation in the environment, its biodegradation rate can significantly vary
with environmental conditions and the distribution of microbial populations. To
overcome these shortcomings augmented PHB biodegradation was analyzed in this
work. As Streptomyces species are known for thriving in soil landfill environments they
were chosen as promising catalysts for enhanced landfilling biodegradation. Therefore,
newly isolated Streptomyces sp. DG19 was selected based on its ability to degrade
biopolymers, including PHB and its genome was analyzed for plastic presence of genes
that encode common plastic hydrolyses. Biodegradation of PHB material supplemented
with demonstrated PHB degrader, Streptomyces sp. DG19 was assessed under the
landfilling and home composting conditions. The spore-forming capacity of
Streptomyces sp. DG19 was crucial for cell immobilization as live cells are sensitive to
hydration, osmotic pressure, temperature and pH. Furthermore, the biodegradation of
PHB-PHO blends is impacted by the increased crystallinity and hydrophobicity with
water contact angle shifting from 93° for PHB to 99° and 105° for 80:20 and 70:30
PHB/PHO. In conclusion, a scalable approach to producing biocomposite materials
offering a promising solution for sustainable waste management is presented.Compiled poster abstract booklet for the 19th Int’l Symposium on Biopolymers 20th-23rd October 2024 School of Biological Sciences, Universiti Sains Malaysi
Plant beneficial bacteria Paraburkholderia phytofirmans PsJN protects Arabidopsis thaliana against fungal pathogen Botrytis cinerea
No full textAs plants face a wide range of pathogens, their innate immunity provides the first line of defense.
This response can be triggered by beneficial microorganisms resulting in plant priming and
increased resilience during stress. Paraburkholderia phytofirmans PsJN is an endophytic and rhizospheric
beneficial bacterial strain helping plants during abiotic and biotic stress. Botrytis cinerea,
a phytopathogenic fungus with a wide host range infecting more than 1400 plant species, is a
destructive necrotrophic pathogen responsible for significant crop losses, before and after harvest.
Beneficial microorganisms, including PsJN, have demonstrated the ability to enhance the
plant’s defense response against this pathogen. In our experiments on Arabidopsis thaliana, PsJN
significantly reduced Botrytis-induced necrosis four days after inoculation, indicating its protective
effect against fungal infection. To investigate interactions among the plant, PsJN, and the pathogen
B. cinerea, RNA sequencing was performed on Arabidopsis roots and leaves that were or not
inoculated with PsJN, mock-treated, or infected with B. cinerea spores at 24 hours of treatment.
Transcriptome analysis revealed significant alterations in gene expression patterns in both roots
and leaves in the presence of PsJN during fungal infection. In PsJN inoculated roots of Botrytis infected
plants, 1452 of DEG were upregulated and 1668 downregulated, while in leaves there were
953 upregulated and 1010 downregulated genes. The differentially expressed genes in roots were
associated with various biological processes such as response to fungus, cytoplasmic translation,
cell wall organization, and secretory vesicle pathways, whereas in leaves, they were related to processes
like photosynthesis, carbohydrate metabolism, and homeostasis.Book of abstract: 5th International Conference on Plant Biology (24th SPPS Meeting
Biomarker Potential of TNS3-203 and LRRFIP1-211 Alternative Transcripts in Oral Squamous Cell Carcinoma
No full textA recent pan-cancer transcriptome analysis showed differential activity of alternative promoters of
genes TNS3 and LRRFIP1 in head and neck squamous cell carcinoma compared to non-cancerous
tissue. Upregulated promoters give rise to TNS3-203 and LRRFIP1-211 transcripts. Our aim was to
investigate the biomarker potential of TNS3-203 and LRRFIP1-211 alternative transcripts in oral
cancer, the most common type of head and neck cancer.
An in silico approach was used to characterize the promoters and transcripts of interest. Promoter
regions were analyzed for the presence of CpG islands and potential binding of transcription factors
using multiple bioinformatic tools. Analysis included prediction of coding potential, transcript
localization and potential interactions with miRNA molecules. Expression of transcripts and miRNA
molecules was retrieved from GSE246050 Gene Expression Omnibus dataset and corresponding
Sequence Read Archive entry. Relative expression of TNS3-203 and LRRFIP1-211 transcripts was
evaluated by qRT-PCR in a group of 46 oral cancer patients in samples of cancer and adjacent noncancerous
tissue.
There are no predicted CpG islands in sequences of the upregulated promoter of TNS3 and LRRFIP1
genes, which could possibly explain their upregulation in cancer. Different transcription factors are
identified to regulate up- and downregulated LRRFIP1 promoter as HNF-1 and SRF transcriptional
factor. HNF-1 is involved in regulating the activity of various genes and members of the HNF family
are also associated with various types of cancer. SRF promotes cell proliferation while induces cell
death resistance, invasion and metastasis.
Results of this study showed that TNS3-203 was significantly overexpressed in oral cancer compared
with matched non-cancerous tissue and the level of this transcript may serve as a reliable indicator of
oral cancer presence. There were no differences in LRRFIP1-211 level between analyzed samples.
None of the investigated transcripts has prognostic potential in oral cancer. LRRFIP-211 is more likely
to be long non-coding RNA as it is longer and probably more stable due to the retained intron.
The results obtained indicate the diagnostic potential of TNS3-203, but not LRRFIP1-211 transcript
and its role in oral carcinogenesis. Future studies should investigate the expression of TNS3-203 in
liquid biopsies in oral cancer patients to confirm if TNS3-203 can be used as a universal diagnostic
biomarker.Book of abstract: “HDIR-7: Advances in Cancer Research & Treatment” The 7th Meeting of the Croatian Association for Cancer Research with International Participation November 7 & 8, 2024 Hotel International, Zagreb, Croati
Enhancing Antibacterial Efficacy: Combining Novel Bacterial Topoisomerase Inhibitors with Efflux Pump Inhibitors and Other Agents Against Gram-Negative Bacteria
No full textBackground: The novel bacterial topoisomerase inhibitors (NBTIs) developed in our laboratory show potent on-target enzyme inhibition but suffer from low activity against Gram-negative bacteria. Methods: With the aim of improving the antibacterial activity of our compounds against Gram-negative bacteria, we tested them in combination with different efflux pump inhibitors (EPIs), a strategy that showed promise in several other classes of antimicrobials. We also investigated the combined effect of NBTIs with ATP-competitive inhibitors of bacterial type II topoisomerases (ACIs), as well as the antibiofilm properties of our compounds and the combination with EPIs against early and mature Acietobacter baumannii biofilm. Results: Our results demonstrate that combinations of NBTIs with EPI Phenylalanine-arginyl-β-naphthylamide significantly reduce the corresponding NBTIs’ minimal inhibitory concentration values and show potentiation of A. baumannii biofilm inhibition as compared to NBTIs alone. Although combinations of NBITs and ACIs did not show synergistic effects, the FIC index value calculations revealed additive effects for all the combinations of a selected NBTI in combination with three ACIs in all the assayed Gram-negative bacteria from the ESKAPE group. Conclusions: These results show for the first time that combinations of NBTIs with either EPIs or a different class of the topoisomerase inhibitors may be a beneficial strategy to combat difficult-to-treat bacterial infections
Recombinant Expression of Se1JFR Esterase from Streptomyces sp. WAC04770 in Different Escherichia coli and Bacillus subtilis Strains
No full textPlastics are among the most prevalent materials in the world today, known for their lightweight,
moldability, resilience, and durability. However, their non-biodegradable nature poses a significant
threat to environmental sustainability (Amobonye et al., 2021; Topakas et al., 2022). Biodegradable
plastics, which can be degraded by enzymatic activity, such as Se1JFR esterase, offer a promising
alternative to conventional polymers (Wei et al., 2020). In this study, Escherichia coli and Bacillus
subtilis were employed as expression systems for Se1JFR esterase, with bioinformatic tools used to
analyze the enzyme’s properties and 3D structure, highlighting the significance of disulfide bonds for
system compatibility. The expression efficiency and esterase activity were evaluated under varying
incubation conditions using an esterase assay, while spectrophotometric absorbance was measured to
assess enzyme activity. E. coli was identified as the more effective expression platform.
Among the tested bacterial strains, E. coli BL21 (DE3) demonstrated the highest levels of protein
expression and enzymatic activity, with a rate of 0,024 μmol/mL*min, and a protein yield of 0,025-
0,05 mg/mL. The purified enzyme's capacity to degrade poly(caprolactone) was further demonstrated
through an agar plate assay, where clear zones indicated successful plastic degradation. Despite these
promising results, the protein yield and activity were below the standard values reported in the
literature. Future research should focus on optimizing incubation conditions, testing different vectors
and signal peptides for improved expression in the B. subtilis system.Book of abstract: Young BMEco, April 29th and May 4th 2025 , Czech Republic, Pragu