Purpose: To characterize the effect of variants identified in the 5’-untranslated region of the EYS gene in patients with autosomal recessive retinitis pigmentosa (ARRP).

Methods: Variant screening included gene panel, Sanger, exome, and genome sequencing. Functional validation included an electrophoretic mobility shift assay (EMSA) and various luciferase assays. Clinical examination included visual acuity testing, electroretinography (ERG) testing, and retinal imaging.

Results: Patients with RP from six EYS biallelic Arab-Muslim families harbored a 5’-noncoding EYS variant, c.-453G>T, and four harbored a structural variant affecting the 5’-noncoding exons. The EMSA analysis revealed an effect on the binding of transcription factors for c.-453G>T and a neighboring variant, c.-454G>T, which was reported previously in a patient with RP. Dual luciferase assays using the overexpression of various transcription factors showed distinct effects on expression. c.-453G>T was associated with higher luciferase expression with CRX overexpression, and c.-454G>C was associated with higher luciferase expression with OTX2 overexpression. In addition, these two variants were found to influence translation by affecting upstream initiation codons. Interestingly, the visual functions (including age of onset, visual acuity and ERG responses) of EYS RP patients who harbor c.-453G>T is better than that of those with biallelic null EYS mutations.

Conclusions: Our analysis revealed both single-nucleotide and structural variants in the EYS promoter as the cause of ARRP. These variants may affect EYS expression via a dual mechanism by altering transcription factor binding affinity at the EYS promoter and by affecting upstream open reading frames.

The biosynthesis of Iturin A, a compound with potent antimicrobial properties, has attracted considerable interest. However, challenges persist in achieving high yields from wild-type strains and optimizing Bacillus amyloliquefaciens for industrial-scale production. This study investigates the genetic foundations of Iturin A biosynthesis, emphasizing the roles of endogenous plasmids and Rap phosphatases. By optimizing and updating CRISPR/Cas9-based gene-editing tools, the modifications were successfully applied to the genetic manipulation of Bacillus amyloliquefaciens. The deletion of the endogenous plasmid plas1 from B. amyloliquefaciens HM618 significantly enhances Iturin A production, likely due to the removal of the Rap phosphatase gene located on plas1. Additionally, the targeted inactivation of the rapC, rapF, and rapH genes within the chromosomal DNA further increases Iturin A yield and specific productivity, although this is accompanied by reduced cell growth. By strategically combining plasmid deletion with targeted gene inactivation, a balance between cellular growth and Iturin A production is established. The engineered strain subsequently produced 849.9 mg/L of Iturin A within 48 hours, demonstrating a significant improvement in production efficiency. These findings underscore the crucial role of microbial genetics in optimizing Iturin A biosynthesis and offer a novel approach for enhancing the industrial production of this valuable antimicrobial compound in B. amyloliquefaciens.

Mycorrhizal symbioses, crucial for plant nutrition and ecosystem functioning, exhibit varying degrees of specificity between plant hosts and fungal symbionts. This study explores the genomic underpinnings of mycorrhizal fungal specificity, synthesizing recent advances in understanding the molecular mechanisms governing these intricate associations. Comparative genomic analyses of over 100 mycorrhizal fungal species have revealed that the loss of plant cell wall-degrading enzymes is a common feature, with ectomycorrhizal fungi retaining only 10-15% of these enzymes compared to their saprotrophic ancestors. This adaptation reflects the transition from saprotrophic to symbiotic lifestyles. Studies on effector proteins have identified several mycorrhiza-induced small secreted proteins (MiSSPs) that play key roles in symbiosis establishment. For instance, the MiSSP7 gene in Laccaria bicolor is essential for the colonization of Populus roots, highlighting the importance of effectors in host specificity. Genomic investigations have also uncovered expansions in certain gene families, such as those encoding for nutrient transporters, with some ectomycorrhizal fungi possessing up to three times more ammonium transporter genes than their non-mycorrhizal relatives. Furthermore, the analysis of plant genomes has revealed that up to 5% of plant genes are differentially expressed during mycorrhizal colonization, with common symbiosis genes being conserved across 80-90% of land plants. This study underscores the complex genomic landscape underlying mycorrhizal specificity, emphasizing the co-evolution of plant and fungal genomes. Future research directions include the functional characterization of candidate genes and exploration of epigenetic mechanisms in symbiosis regulation.

The frequency of clear-cell renal-cell carcinoma (ccRCC) metastasis has reached 25-30%, which makes the search for prognostic biomarkers urgent. Long non-coding RNAs (lncRNAs) regulate biological processes and may serve as prognostic markers. The aim of our study was to compare the expression levels of lncRNAs GAS5, HAND2-AS1, LINC00152 and LINC00339 in tumors and normal kidney tissue and analyze the association of their changes with clinicopathological parameters of ccRCC.

We used archival RNA samples (RIN>7) from 70 primary tumors with a confirmed diagnosis of ccRCC and 70 adjacent normal kidney tissues. Expression analysis was performed by RT-qPCR. Statistical analysis was performed using a multivariate ANOVA test (p ≤ 0.05).

A decrease in the expression of GAS5 by 5.25, HAND2-AS1 by 5.92, LINC00152 by 2.04 and LINC00339 by 2.34 times was revealed in the tumor tissues. The expression of LINC00152 did not change at T1, T2 and T3 (the TNM classification) and decreased by 33.43 times when the tumor grew beyond Gerota’s fascia (T4). LINC00152 expression was lower in T4 tumors compared to tumors of a smaller size: T1/T4—10.86 times; T2/T4—22.76 times; and T3/T4—27.11 times. LINC00152 expression decreased by 4.71 times in cases with multiple lesions of regional lymph nodes (normal/N2), including compared with tumors with no metastases in the lymph nodes (N0/N2) or one affected lymph node (N1/N2), by 4.36 and 7.26 times, respectively. HAND2-AS1 had a lower level of expression in tumors with multiple lesions of regional lymph nodes: normal/N2—15.02 times; N0/N2—5.84 times; N1/N2—4.04 times.

The significant decrease in the expression of HAND2-AS1 and LINC00152 when more than one regional lymph node is affected (N2 according to the TNM classification) in ccRCC allows us to propose them as prognostic markers of lymphogenous metastasis in the absence of the ability to evaluate regional lymph nodes (Nx).

Transcriptomics provides a link between the genome, proteome, and cellular phenotype; therefore, it is regarded as a unique approach to revealing the molecular basis of phenotypes. Despite the availability of huge datasets for a wide range of taxa in public databases, RNA-Seq data analysis and interpretation remain challenging. This is due to factors including limited processing capability and the need for specialized coding expertise and expensive software. Here, we used publicly available RNA-Seq datasets and emphasized an in-depth description of each cutting-edge system/tool, especially Galaxy platform and TRAPID 2. We also conducted a downstream analysis ofthe authenticity of the taxonomic identity, the global transcriptome map/network, and the functional enrichment of overrepresented genes in silkworm (Bombyx mori L.) and mulberry (Morus spp.). We provided a flexible de novo assembly, annotations, and sufficient visualization to interpret the RNA-Seq analysis data. The current data-mining approach sheds light on the significant influence of silkworm black dilute (bd) mutants on melanogenesis and tyrosine degradation, which is mediated by dopachrome tautomerase (DTC). However, M. serrata revealed significant polyploid trade-offs, such as investments in biochemical expenditure, nucleic acid metabolism, the intrinsic activity of transposable elements, and a functional association with chloroplasts. Overall, the current data-mining method reduces costs and requires no prior coding expertise to comprehend the genetic foundation of phenotype variance. The present approach can also help to decode the genetic architecture of novel phenotypes, including trait-specific accessions, ploidy-associated traits, and the hybridity effect of not only Seri-genetic resources but also other model and non-model organisms.

Cancer research largely focuses on the epithelial-to-mesenchymal transition (EMT) as a critical mechanism required for the acquisition of the invasive potential of cancer cells, which can culminate in the formation of metastases. This process involves the transformation of epithelial cells into mesenchymal cells by acquiring suppressed levels of E-cadherin, an anti-EMT marker with a role in the establishment of intercellular junctions. Also, the expression of pro-EMT markers, such as the regulatory marker SNAIL, as well as the effectors N-cadherin and Vimentin, rises as cancer advances. Therefore, there is an emerging need for bioactive substances able to target and modulate EMT markers and reverse this process. The unsaturated fatty acid 10H2DA has not been investigated so far regarding its potential to target specific EMT markers in colorectal cancer (CRC), which was the aim of this study.

Colorectal cancer cell line SW-480 isolated from stage II CRC was treated with 10H2DA in two selected concentrations of 10 and 100 μM. After 24 h, the gene expression of E-cadherin, SNAIL, N-cadherin, and Vimentin markers was assessed by the qPCR method. Additionally, changes in the protein concentration of these markers between control (untreated) and treated cells were evaluated by immunofluorescent assay.

Queen bee acid successfully upregulated the expression of the anti-EMT marker E-cadherin. Our results also point towards a downregulated expression of the pro-EMT regulatory marker SNAIL, as well as the effector markers N-cadherin and Vimentin. This successively led to a significant elevation in E-cadherin on the protein level, and also to an inhibition of pro-EMT proteins, namely, SNAIL, N-cadherin, and Vimentin.

The potential of the unsaturated acid 10H2DA to modulate the expression of specific and significant EMT markers in CRC is obvious and prominent, and it should not be neglected in future studies regarding anticancer therapeutic approaches.

According to the literature, the frequency of mutations in genes associated with the development of acute myeloid leukemia (AML) varies significantly.

The objective of this study was to obtain up-to-date data on the prevalence of mutations in the "hot spots" of the FLT3, NPM1, IDH1, IDH2, and DNMT3A genes in AML.

Methods: An analysis of NGS data from 1567 patients with AML presented in the cancer genomics database on C-Bioportal and 124 Russian patients with AML was performed.

Results: According to the database analysis, at the time of the diagnosis of the disease, 46.6% of patients had mutations in DNMT3A p.R882, NPM1 p.W288Cfs*12, FLT3-ITD, FLT3-TKD1, IDH1 p.R132, and IDH2 p.R140. Only in a third of cases (30.1%) did the DNMT3A mutation of R.R882 occur in patients in an isolated variant. In 47.4% of cases, it was combined with NPM1 p.W288Cfs*12, in 34.1% with mutations in the hot spots of the FLT3 gene, and in 23.0% with recurrent mutations in IDH1 and IDH2. At the same time, the combination revealed by our data remains highly significant (p<0.001) even after adjusting for the multiplicity of comparisons (q<0.001). In Russian AML patients, the mutation rates in the "hot spots" of genes generally corresponded to the data from the C-Bioportal cancer genomics database and amounted to the following: DNMT3A p.R882—7.3%; NPM1 p.W288Cfs*12—15.3%; FLT3-ITD—14.5%; FLT3-TKD1—4.0%; IDH1 p.R132—5.6%; IDH2 p.R140—10.5%.

Conclusions: The data obtained indicate that in 40-50% of AML cases, clinically significant recurrent mutations in one or more of the studied genes are detected at the onset of the disease. Mutations for which targeted drugs (FLT3, IDH1, and IDH2 inhibitors) have been developed occur in 35% of patients. In one-fifth of cases (18.1%) of AML, NPM1 p.W288Cfs*12 is detected, which can be used as an independent target for the molecular assessment of minimal residual disease.

Purpose:
To report CREB3 as a novel genetic basis of inherited retinal diseases (IRDs) in families mostly of North African Jewish origin.
Methods:
Thirteen patients with a clinical diagnosis of retinitis pigmentosa or macular degeneration were analyzed using next-generation sequencing (NGS). Homozygosity mapping was performed on the Franklin platform for seven patients from four unrelated families. Expression analysis was performed on patient-derived skin fibroblasts using the reverse transcription–polymerase chain reaction (RT-PCR) and Western blot analysis, as well as by interrogating previously published retinal single-cell RNA-seq data. Immunohistochemistry staining (IHC) was performed on wild-type mouse retinal sections using an anti-CREB3 antibody.
Results:
A founder homozygous nonsense variant in CREB3 (c.881G>A, p.Trp294*) was identified in thirteen patients from four unrelated families. All patients manifested retinal degeneration with varying levels of severity. Homozygosity mapping revealed that out of the seven patients, six patients of North African Jewish descent had an identical haplotype. In patient-derived fibroblasts, the mutant mRNA transcript generated a truncated CREB3 protein. Expression analysis and IHC revealed CREB3 RNA and protein expression in various retinal cell types, indicating its vital role in photoreceptor function.
Conclusion:
We report here for the first time the involvement of CREB3 in IRDs. CREB3 was previously shown to be upregulated following UV radiation, which might contribute to extensive clinical variability observed in this relatively large cohort of homozygous patients for the same truncating variant.

Introduction

A major problem in biomedical research is poor reproducibility, with many scientific results not being successfully replicated. Many biological factors can explain this, from sampling effects to the influence of environmental factors such as microbiota. But other factors that are much easier to address, such as a lack of access to methodological details, raw data, and research materials, are also playing a major role in the reproducibility crisis. In particular, inadequate reporting of the genetics of laboratory animals in research papers is common.

Methods

ARRIVE has been a pioneer in demonstrating to researchers using in vivo animals the importance of structured documentation for animal research. In the same vein, we brought together a large number of leading international experts, consortia, and learned societies in the field of animal genetics to define the LAG-R recommendations for Laboratory Animal Genetic Reporting.

Results

Our recommendations have been published in Nature Communications (https://doi.org/10.1038/s41467-024-49439-y). This LAG-R framework defines a set of guidelines to support more complete documentation of the genetic make-up of animals that are used in research.

Conclusions

The purpose of LAG-R is to help authors better describe the information they should already have in scientific papers, but also to provide reviewers with a checklist of essential genetic information that needs to be included in a publication. LAG-R provides an easy-to-understand list of recommendations with the goal of improving reproducibility, reliability, and overall scientific rigor.