Genetic Recombination Research Articles

PARP inhibitors alone or in combination for prostate cancer.

DNA repair genomic aberrations in the Homologous Recombination pathway are identifiable in up to 25% of patients with advanced prostate cancer, making them more likely to benefit from treatment with poly (ADP-ribose) polymerase inhibitors (PARPi) alone or in combination with other therapies, particularly when BRCA driver genomic aberrations are documented. Although several clinical trials have demonstrated the efficacy of this approach, the validation of reliable biomarkers predictive of response still needs further improvement to refine patient selection. In this setting, the characterization of resistance mechanisms and the validation of novel biomarkers are critical to maximize clinical benefit and to develop novel treatment combinations to improve outcomes. In this review, we summarize the development of PARPi in prostate cancer as single agent as well as the efficacy of their combination with other drugs, and the future directions for their implementation in the management of advanced prostate cancer.

Are there discrete recombination pathways in the case of semi-localized thermoluminescence transitions: The significance of thermal treatments towards their discrimination

This study explores an advanced luminescence protocol, aiming to reveal recombination mechanisms behind stimulated luminescence in MgB4O7: Dy, Na (MBO), and BeO: Cr, Mg, Sn (BeOR). The two-stage protocol integrates elements from prior methodologies. In the initial phase, materials undergo precise annealing at varying temperatures, followed by Thermoluminescence (TL) measurements. The second stage involves repeating annealing procedures with a single high dose. Materials exhibit localized peaks within dosimetric regions. The analysis reveals MBO's two distinct excited states, valuable for understanding recombination mechanisms and potential transitions. While BeOR undergoes a complete transition, MBO shows indications of a similar transformation. The confirmed presence of two distinct excited states within MBO establishes a fundamental premise. Validation includes peak shape methods (PSM), initial rise (IR), and prompt isothermal decay (PID). This approach enhances the understanding of stimulated luminescence phenomena, providing insights into trapping parameters and transitions between states in the studied materials.

Fluorescent human RPA to track assembly dynamics on DNA

DNA metabolic processes including replication, repair, recombination, and telomere maintenance occur on single-stranded DNA (ssDNA). In each of these complex processes, dozens of proteins function together on the ssDNA template. However, when double-stranded DNA is unwound, the transiently open ssDNA is protected and coated by the high affinity heterotrimeric ssDNA binding Replication Protein A (RPA). Almost all downstream DNA processes must first remodel/remove RPA or function alongside to access the ssDNA occluded under RPA. Formation of RPA-ssDNA complexes trigger the DNA damage checkpoint response and is a key step in activating most DNA repair and recombination pathways. Thus, in addition to protecting the exposed ssDNA, RPA functions as a gatekeeper to define functional specificity in DNA maintenance and genomic integrity. RPA achieves functional dexterity through a multi-domain architecture utilizing several DNA binding and protein-interaction domains connected by flexible linkers. This flexible and modular architecture enables RPA to adopt a myriad of configurations tailored for specific DNA metabolic roles. To experimentally capture the dynamics of the domains of RPA upon binding to ssDNA and interacting proteins we here describe the generation of active site-specific fluorescent versions of human RPA (RPA) using 4-azido-L-phenylalanine (4AZP) incorporation and click chemistry. This approach can also be applied to site-specific modifications of other multi-domain proteins. Fluorescence-enhancement through non-canonical amino acids (FEncAA) and Förster Resonance Energy Transfer (FRET) assays for measuring dynamics of RPA on DNA are also described. The fluorescent human RPA described here will enable high-resolution structure-function analysis of RPA-ssDNA interactions.

Multifunctional Surface Treatment against Imperfections and Halide Segregation in Wide-Band Gap Perovskite Solar Cells.

Mixed-halide wide-band gap perovskites (WBPs) still suffer from losses due to imperfections within the absorber and the segregation of halide ions under external stimuli. Herein, we design a multifunctional passivator (MFP) by mixing bromide salt, formamidinium bromide (FABr) with a p-type self-assembled monolayer (SAM) to target the nonradiative recombination pathways. Photoluminescence measurement shows considerable suppression of nonradiative recombination rates after treatment with FABr. However, WBPs still remained susceptible to halide segregation for which the addition of 25% p-type SAM was effective to decelerate segregation. It is observed that FABr can act as a passivating agent of the donor impurities, shifting the Fermi-level (Ef) toward the mid-band gap, while p-type SAM could cause an overweight of Ef toward the valence band. Favorable band bending at the interface could prevent the funneling of carriers toward I-rich clusters. Instead, charge carriers funnel toward an integrated SAM, preventing the accumulation of polaron-induced strain on the lattice. Consequently, n-i-p structured devices with an optimal MFP treatment show an average open-circuit voltage (VOC) increase of about 20 mV and fill factor (FF) increase by 4% compared with the control samples. The unencapsulated devices retained 95% of their initial performance when stored at room temperature under 40% relative humidity for 2800 h.

Balanced spatiotemporal arrangements of histone H3 and H4 posttranslational modifications are necessary for meiotic prophase I chromosome organization.

Dynamic nuclear architecture and chromatin organizations are the key features of the mid-prophase I in mammalian meiosis. The chromatin undergoes major changes, including meiosis-specific spatiotemporal arrangements and remodeling, the establishment of chromatin loop-axis structure, pairing, and crossing over between homologous chromosomes, any deficiencies in these events may induce genome instability, subsequently leading to failure to produce gametes and infertility. Despite the significance of chromatin structure, little is known about the location of chromatin marks and the necessity of their balance during meiosis prophase I. Here, we show a thorough cytological study of the surface-spread meiotic chromosomes of mouse spermatocytes for H3K9,14,18,23,27,36, H4K12,16 acetylation, and H3K4,9,27,36 methylation. Active acetylation and methylation marks on H3 and H4, such as H3K9ac, H3K14ac, H3K18ac, H3K36ac, H3K56ac, H4K12ac, H4K16ac, and H3K36me3 exhibited pan-nuclear localization away from heterochromatin. In comparison, repressive marks like H3K9me3 and H3K27me3 are localized to heterochromatin. Further, taking advantage of the delivery of small-molecule chemical inhibitors methotrexate(heterochromatin enhancer), heterochromatin inhibitor, anacardic acid (histone acetyltransferase inhibitor), trichostatin A (histone deacetylase inhibitor), IOX1 (JmjC demethylases inhibitor), and AZ505 (methyltransferase inhibitor) in seminiferous tubules through the rete testis route, revealed that alteration in histone modifications enhanced the centromere mislocalization, chromosome breakage, altered meiotic recombination and reduced sperm count. Specifically, IOX1 and AZ505 treatment shows severe meiotic phenotypes, including altering chromosome axis length and chromatin loop size via transcriptional regulation of meiosis-specific genes. Our findings highlight the importance of balanced chromatin modifications in meiotic prophase I chromosome organization and instability.

Enrichment and Diversification of the Wheat Genome via Alien Introgression

Wheat, including durum and common wheat, respectively, is an allopolyploid with two or three homoeologous subgenomes originating from diploid wild ancestral species. The wheat genome’s polyploid origin consisting of just three diploid ancestors has constrained its genetic variation, which has bottlenecked improvement. However, wheat has a large number of relatives, including cultivated crop species (e.g., barley and rye), wild grass species, and ancestral species. Moreover, each ancestor and relative has many other related subspecies that have evolved to inhabit specific geographic areas. Cumulatively, they represent an invaluable source of genetic diversity and variation available to enrich and diversify the wheat genome. The ancestral species share one or more homologous genomes with wheat, which can be utilized in breeding efforts through typical meiotic homologous recombination. Additionally, genome introgressions of distant relatives can be moved into wheat using chromosome engineering-based approaches that feature induced meiotic homoeologous recombination. Recent advances in genomics have dramatically improved the efficacy and throughput of chromosome engineering for alien introgressions, which has served to boost the genetic potential of the wheat genome in breeding efforts. Here, we report research strategies and progress made using alien introgressions toward the enrichment and diversification of the wheat genome in the genomics era.

Coordinated evolution of the SNORD115 and SNORD116 tandem repeats at the imprinted Prader–Willi/Angelman locus

Abstract The Prader–Willi/Angelman syndrome (PWS/AS) locus is regulated by the epigenetic mechanism of parental genomic imprinting. This region holds two eutherian-specific, large tandem repeats of box C/D small nucleolar RNA (Snord) genes called SNORD115 and SNORD116, whose loss of paternal expression is key in the development of the PWS. Snords represent an ancient class of noncoding RNAs that typically direct the 2′-O-methylation of specific nucleotides of ribosomal RNAs. However, Snord115 and Snord116 belong to the large class of orphan Snords whose functions remain unclear. The constraints that generated and maintained their unusual genetic organization for mammalian genomes have been poorly addressed to date. Here, a comparative analysis of the evolutionary history of both tandem repeats reveals that several genetic events affected them concomitantly, including copy gains and losses between species, emergence of gene subfamilies in catarrhines or partial tandem duplication in rats. Several indications suggest that parental genomic imprinting orchestrated this coordination of events, adding a new effect on mammalian genome structure and evolution to its roles in gene dosage, meiotic recombination and replication timing. Finally, our work provides a functional rationale for the existence of closely located tandem repeats of small RNA genes in mammalian genomes.

Structural variation and DNA methylation shape the centromere-proximal meiotic crossover landscape in Arabidopsis.

Centromeres load kinetochore complexes onto chromosomes, which mediate spindle attachment and allow segregation during cell division. Although centromeres perform a conserved cellular function, their underlying DNA sequences are highly divergent within and between species. Despite variability in DNA sequence, centromeres are also universally suppressed for meiotic crossover recombination, across eukaryotes. However, the genetic and epigenetic factors responsible for suppression of centromeric crossovers remain to be completely defined. To explore the centromere-proximal meiotic recombination landscape, we map 14,397 crossovers against fully assembled Arabidopsis thaliana (A. thaliana) genomes. A. thaliana centromeres comprise megabase satellite repeat arrays that load nucleosomes containing the CENH3 histone variant. Each chromosome contains a structurally polymorphic region of ~3-4 megabases, which lack crossovers and include the satellite arrays. This polymorphic region is flanked by ~1-2 megabase low-recombination zones. These recombination-suppressed regions are enriched for Gypsy/Ty3 retrotransposons, and additionally contain expressed genes with high genetic diversity that initiate meiotic recombination, yet do not crossover. We map crossovers at high-resolution in proximity to CEN3, which resolves punctate centromere-proximal hotspots that overlap gene islands embedded in heterochromatin. Centromeres are densely DNA methylated and the recombination landscape is remodelled in DNA methylation mutants. We observe that the centromeric low-recombining zones decrease and increase crossovers in CG (met1) and non-CG (cmt3) mutants, respectively, whereas the core non-recombining zones remain suppressed. Our work relates the genetic and epigenetic organization of A. thaliana centromeres and flanking pericentromeric heterochromatin to the zones of crossover suppression that surround the CENH3-occupied satellite repeat arrays.

Metabolic stress-induced long ncRNA transcription governs the formation of meiotic DNA breaks in the fission yeast fbp1 gene.

Meiotic recombination is a pivotal process that ensures faithful chromosome segregation and contributes to the generation of genetic diversity in offspring, which is initiated by the formation of double-strand breaks (DSBs). The distribution of meiotic DSBs is not uniform and is clustered at hotspots, which can be affected by environmental conditions. Here, we show that non-coding RNA (ncRNA) transcription creates meiotic DSBs through local chromatin remodeling in the fission yeast fbp1 gene. The fbp1 gene is activated upon glucose starvation stress, in which a cascade of ncRNA-transcription in the fbp1 upstream region converts the chromatin configuration into an open structure, leading to the subsequent binding of transcription factors. We examined the distribution of meiotic DSBs around the fbp1 upstream region in the presence and absence of glucose and observed several new DSBs after chromatin conversion under glucose starvation conditions. Moreover, these DSBs disappeared when cis-elements required for ncRNA transcription were mutated. These results indicate that ncRNA transcription creates meiotic DSBs in response to stress conditions in the fbp1 upstream region. This study addressed part of a long-standing unresolved mechanism underlying meiotic recombination plasticity in response to environmental fluctuation.

Type 2 cytokine signaling in macrophages protects from cellular senescence and organismal aging

Accumulation of senescent cells in organs and tissues is a hallmark of aging and known to contribute to age-related diseases. Although aging-associated immune dysfunction, or immunosenescence, is known to contribute to this process, the underlying mechanism remains elusive. Here, we report that type 2 cytokine signaling deficiency accelerated aging and, conversely, that the interleukin-4 (IL-4)-STAT6 pathway protected macrophages from senescence. Mechanistically, activated STAT6 promoted the expression of genes involved in DNA repair both via homologous recombination and Fanconi anemia pathways. Conversely, STAT6 deficiency induced release of nuclear DNA into the cytoplasm to promote tissue inflammation and organismal aging. Importantly, we demonstrate that IL-4 treatment prevented macrophage senescence and improved the health span of aged mice to an extent comparable to senolytic treatment, with further additive effects when combined. Together, our findings support that type 2 cytokine signaling protects macrophages from immunosenescence and thus hold therapeutic potential for improving healthy aging.

LOX-1 regulation of H-type vascular endothelial cell regeneration in hyperglycemia.

Diabetic osteoporosis (DOP) is the most common secondary form of osteoporosis. Diabetes mellitus affects bone metabolism; however, the underlying pathophysiological mechanisms remain unclear. Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) expression is upregulated in conditions characterized by vascular injury, such as atherosclerosis, hypertension, and diabetes. Additionally, Notch, HIF-1α, and VEGF are involved in angiogenesis and bone formation. Therefore, we aimed to investigate the expression of Notch, HIF-1α, and VEGF in the LOX-1 silencing state. Rat bone H-type vascular endothelial cells (THVECs) were isolated and cultured in vitro. Cell identification was performed using immunofluorescent co-expression of CD31 and Emcn. Lentiviral silencing vector (LV-LOX-1) targeting LOX-1 was constructed using genetic recombination technology and transfected into the cells. The experimental groups included the following: NC group, HG group, LV-LOX-1 group, LV-CON group, HG + LV-LOX-1 group, HG + LV-CON group, HG + LV-LOX-1 + FLI-06 group, HG + LV-CON + FLI-06 group, HG + LV-LOX-1 + LW6 group, and HG + LV-CON + LW6 group. The levels of LOX-1, Notch, Hif-1α, and VEGF were detected using PCR and WB techniques to investigate whether the expression of LOX-1 under high glucose conditions has a regulatory effect on downstream molecules at the gene and protein levels, as well as the specific molecular mechanisms involved. High glucose (HG) conditions led to a significant increase in LOX-1 expression, leading to inhibition of angiogenesis, whereas silencing LOX-1 can reverse this phenomenon. Further analysis reveals that changes in LOX-1 will promote changes in Notch/HIF-1α and VEGF. Moreover, Notch mediates the activation of HIF-1α and VEGF. The activation of LOX-1 and the inhibition of Notch/HIF-1α/VEGF in THVECs are the main causes of DOP. These findings contribute to our understanding of the pathogenesis of DOP and offer a novel approach for preventing and treating osteoporosis.

In vitro replicative potential of an HIV-1/MO intergroup recombinant virus compared to HIV-1/M and HIV-1/O parental viruses

Genetic recombination is one of the major evolution processes of HIV-1. Despite their great genetic divergence, HIV-1 groups M and O can generate HIV-1/MO intergroup recombinants. The current description of 20 HIV-1/MO unique recombinant forms suggests a possible benefit of the recombination. The aim of this work was to study in vitro the replicative potential of HIV-1/MO recombinant forms. This analysis was based on a simple recombination pattern, [Ogag/pol-Menv], harboring a breakpoint in Vpr. A chimeric infectious molecular clone, pOM-TB-2016 was synthesized from HIV-1/M subtype B and HIV-1/O subgroup T and recombinant viruses were obtained by transfection/co-culture. To compare the replicative potential of these viruses, two markers were monitored in culture supernatants: Reverse Transcriptase (RT) activity and P24 antigen concentration. The results showed a superiority of the group M parental virus compared to group O for both markers. In contrast, for the recombinant virus, RT activity data did not overlap with the concentration of P24 antigen, suggesting a hybrid behavior of the recombinant, in terms of enzyme activity and P24 production. These results highlighted many hypotheses about the impact of recombination on replicative potential and demonstrated again the significant plasticity of HIV genomes and their infinite possibility of evolution.

RNF20 Regulates Oocyte Meiotic Spindle Assembly by Recruiting TPM3 to Centromeres and Spindle Poles.

Previously a ring finger protein 20 (RNF20) is found to be essential for meiotic recombination and mediates H2B ubiquitination during spermatogenesis. However, its role in meiotic division is still unknown. Here, it is shown that RNF20 is localized at both centromeres and spindle poles, and it is required for oocyte acentrosomal spindle organization and female fertility. RNF20-depleted oocytes exhibit severely abnormal spindle and chromosome misalignment caused by defective bipolar organization. Notably, it is found that the function of RNF20 in spindle assembly is not dependent on its E3 ligase activity. Instead, RNF20 regulates spindle assembly by recruiting tropomyosin3 (TPM3) to both centromeres and spindle poles with its coiled-coil motif. The RNF20-TPM3 interaction is essential for acentrosomal meiotic spindle assembly. Together, the studies uncover a novel function for RNF20 in mediating TPM3 recruitment to both centromeres and spindle poles during oocyte spindle assembly.

SOG1 and BRCA1 Interdependently Regulate RAD54 Expression for Repairing Salinity-Induced DNA Double-Strand Breaks in Arabidopsis.

As sessile organisms, land plants experience various forms of environmental stresses throughout their life span. Therefore, plants have developed extensive and complicated defense mechanisms, including a robust DNA damage response (DDR) and DNA repair systems for maintaining genome integrity. In Arabidopsis, the NAC [NO APICAL MERISTEM (NAM), ARABIDOPSIS TRANSCRIPTION ACTIVATION FACTOR (ATAF), CUP-SHAPED COTYLEDON (CUC)] domain family transcription factor SUPPRESSOR OF GAMMA RESPONSE 1 (SOG1) plays an important role in regulating DDR. Here, we show that SOG1 plays a key role in regulating the repair of salinity-induced DNA double-strand breaks (DSBs) via the homologous recombination (HR) pathway in Arabidopsis. The sog1-1 mutant seedlings display a considerably slower rate of repair of salinity-induced DSBs. Accumulation of SOG1 protein increases in wild-type Arabidopsis under salinity stress, and it enhances the expression of HR pathway-related genes, including RAD51, RAD54 and BReast CAncer gene 1 (BRCA1), respectively, as found in SOG1 overexpression lines. SOG1 binds specifically to the AtRAD54 promoter at the 5'-(N)4GTCAA(N)3C-3' consensus sequence and positively regulates its expression under salinity stress. The phenotypic responses of sog1-1/atrad54 double mutants suggest that SOG1 functions upstream of RAD54, and both these genes are essential in regulating DDR under salinity stress. Furthermore, SOG1 interacts directly with BRCA1, an important component of the HR-mediated DSB repair pathway in plants, where BRCA1 appears to facilitate the binding of SOG1 to the RAD54 promoter. At the genetic level, SOG1 and BRCA1 function interdependently in modulating RAD54 expression under salinity-induced DNA damage. Together, our results suggest that SOG1 regulates the repair of salinity-induced DSBs via the HR-mediated pathway through genetic interactions with RAD54 and BRCA1 in Arabidopsis.

Photonic Crystal Surface Mode Real-Time Imaging of RAD51 DNA Repair Protein Interaction with the ssDNA Substrate.

Photonic crystals (PCs) are promising tools for label-free sensing in drug discovery screening, diagnostics, and analysis of ligand-receptor interactions. Imaging of PC surface modes has emerged as a novel approach to the detection of multiple binding events at the sensor surface. PC surface modification and decoration with recognition units yield an interface providing the highly sensitive detection of cancer biomarkers, antibodies, and oligonucleotides. The RAD51 protein plays a central role in DNA repair via the homologous recombination pathway. This recombinase is essential for the genome stability and its overexpression is often correlated with aggressive cancer. RAD51 is therefore a potential target in the therapeutic strategy for cancer. Here, we report the designing of a PC-based array sensor for real-time monitoring of oligonucleotide-RAD51 recruitment by means of surface mode imaging and validation of the concept of this approach. Our data demonstrate that the designed biosensor ensures the highly sensitive multiplexed analysis of association-dissociation events and detection of the biomarker of DNA damage using a microfluidic PC array. The obtained results highlight the potential of the developed technique for testing the functionality of candidate drugs, discovering new molecular targets and drug entities. This paves the way to further adaption and bioanalytical use of the biosensor for high-content screening to identify new DNA repair inhibitor drugs targeting the RAD51 nucleoprotein filament or to discover new molecular targets.

Genetic recombination and genotype diversity of norovirus GI in children with acute gastroenteritis in Thailand, 2015-2021

BackgroundHuman norovirus is a predominant etiological agent responsible for acute gastroenteritis across all age groups. Recently, norovirus recombinant strains have been reported as the cause of norovirus outbreaks in several settings and the strains that cause outbreaks mostly belong to the norovirus GII. However, yet, the norovirus GI recombinant strains have never been reported previously in Thailand. The aims of this study were to investigate the genetic recombination and genotype diversity of norovirus GI strains in children hospitalized with acute gastroenteritis in Chiang Mai, Thailand during a period of seven years from 2015 to 2021. MethodsA total of 2829 stool specimens were screened for norovirus GI by real-time PCR, and the polymerase and capsid genes were sequenced and analyzed. ResultsOf 2829 specimens tested, 12 (0.4%) were positive for norovirus GI. Of these, 7 out of 12 (58.3%) strains were identified as norovirus GI recombinant strains. Among 7 norovirus GI recombinant strains, 3, 3, and 1 were identified as GI.3[P13], GI.5[P4], and GI.6[P11], respectively. The remaining five strains were identified as non-recombinant strains of the GI.4[P4], GI.5[P5], and GI.6[P6] genotypes. ConclusionsThe findings highlight the genetic diversity and multiple intergenotype recombinant strains of norovirus GI circulating in children with acute gastroenteritis in Chiang Mai, Thailand from 2015 to 2021. The detection of multiple intergenotype norovirus GI recombinant strains further underscore the complexity of norovirus GI strains circulating in this region.

Regulatory T cells in skin mediate immune privilege of the hair follicle stem cell niche.

Immune tolerance is maintained in lymphoid organs (LOs). Despite the presence of complex immune cell networks in non-LOs, it is unknown whether self-tolerance is maintained in these tissues. We developed a technique to restrict genetic recombination to regulatory T cells (Tregs) only in skin. Selective depletion of skin Tregs resulted in T cell-mediated inflammation of hair follicles (HFs). Suppression did not rely on CTLA-4, but instead on high-affinity interleukin-2 (IL-2) receptor expression by skin Tregs, functioning exclusively in a cell-extrinsic manner. In a novel model of HF stem cell (HFSC)-driven autoimmunity, we reveal that skin Tregs immunologically protect the HFSC niche. Finally, we used spatial transcriptomics to identify aberrant IL-2 signaling at stromal-HF interfaces in a rare form of human alopecia characterized by HFSC destruction and alopecia areata. Collectively, these results reveal the fundamental biology of Tregs in skin uncoupled from the systemic pool and elucidate a mechanism of self-tolerance.

Leveraging machine learning for taxonomic classification of emerging astroviruses.

Astroviruses are a family of genetically diverse viruses associated with disease in humans and birds with significant health effects and economic burdens. Astrovirus taxonomic classification includes two genera, Avastrovirus and Mamastrovirus. However, with next-generation sequencing, broader interspecies transmission has been observed necessitating a reexamination of the current host-based taxonomic classification approach. In this study, a novel taxonomic classification method is presented for emergent and as yet unclassified astroviruses, based on whole genome sequence k-mer composition in addition to host information. An optional component responsible for identifying recombinant sequences was added to the method's pipeline, to counteract the impact of genetic recombination on viral classification. The proposed three-pronged classification method consists of a supervised machine learning method, an unsupervised machine learning method, and the consideration of host species. Using this three-pronged approach, we propose genus labels for 191 as yet unclassified astrovirus genomes. Genus labels are also suggested for an additional eight as yet unclassified astrovirus genomes for which incompatibility was observed with the host species, suggesting cross-species infection. Lastly, our machine learning-based approach augmented by a principal component analysis (PCA) analysis provides evidence supporting the hypothesis of the existence of human astrovirus (HAstV) subgenus of the genus Mamastrovirus, and a goose astrovirus (GoAstV) subgenus of the genus Avastrovirus. Overall, this multipronged machine learning approach provides a fast, reliable, and scalable prediction method of taxonomic labels, able to keep pace with emerging viruses and the exponential increase in the output of modern genome sequencing technologies.

Population genetic diversity of tomato spotted wilt orthotospovirus isolates from tobacco in Yunnan Province, China

We explored the genetic diversity and molecular evolutionary characteristics of tomato spotted wilt orthotospovirus (TSWV) on tobacco in Yunnan. In this study, 523 diseased tobacco leaf samples with suspected TSWV infection from nine different geographical sources in Yunnan were tested, and nucleocapsid protein (NP) of some isolates was amplified and cloned by reverse transcription-polymerase chain reaction (RT‒PCR). Bioinformatics tools were used to analyze the population genetic structure and molecular evolutionary relationships of the tobacco TSWV population. TSWV was detected in 284 of 523 samples, with an average positivity rate of 54.3%. NP gene sequences of the 159 tobacco TSWV isolates obtained were all 777 bp, with 265 nucleic acid polymorphic sites, with 197 variant sites, including 66 parsimony informative and 131 single variant sites. Results for virus consistency and variation analysis based on the NP gene showed that the nucleotide sequence consistency of Yunnan TSWV isolates was 96∼100%, the haplotype diversity 0.963, and the nucleic acid diversity 0.006. Phylogenetic analysis revealed that Yunnan tobacco TSWV isolates belong to the same group, with lower genetic differentiation and more frequent gene exchange among tobacco isolates of different geographical origins; their geographical characteristics were not obvious, and significant recombination signals were detected in the gene sequences of the DL-20 and DL-42 isolates. Many of the NP gene mutations of tobacco TSWV isolates from different geographic origins were deleterious in the nine different planting regions of Yunnan tobacco TSWV isolates, with only the population in the QJ planting region experiencing past population expansion events. The findings suggest that genetic recombination, mutation, and negative selection pressure are major drivers of the genetic diversity of tobacco TSWV in Yunnan. This study report of genetic variation in populations of TSWV isolates from various tobacco regions in Yunnan and provides information on the population structure of Yunnan tobacco TSWV isolates, which can help to prevent and control TSWV on Yunnan tobacco and provide a reference for research of disease-resistant varieties of tobacco.

Genome and GWAS analysis identified genes significantly related to phenotypic state of Rhododendron bark.

As an important horticultural plant, Rhododendron is often used in urban greening and landscape design. However, factors such as the high rate of genetic recombination, frequent outcrossing in the wild, weak linkage disequilibrium, and the susceptibility of gene expression to environmental factors limit further exploration of functional genes related to important horticultural traits, and make the breeding of new varieties require a longer time. Therefore, we choose bark as the target trait which is not easily affected by environmental factors, but also has ornamental properties. Genome-wide association study (GWAS) of Rhododendron delavayi (30 samples), R. irroratum (30 samples) and their F1 generation R. agastum (200 samples) was conducted on the roughness of bark phenotypes. Finally, we obtained 2416.31Gbp of clean data and identified 5 328 800 high-quality SNPs. According to the P-value and the degree of linkage disequilibrium of SNPs, we further identified 4 out of 11 candidate genes that affect bark roughness. The results of gene differential expression analysis further indicated that the expression levels of Rhdel02G0243600 and Rhdel08G0220700 in different bark phenotypes were significantly different. Our study identified functional genes that influence important horticultural traits of Rhododendron, and illustrated the powerful utility and great potential of GWAS in understanding and exploiting wild germplasm genetic resources of Rhododendron.