Search Results (48)

Whole-genome duplication, or tetraploidization, occurs in cells, tissues, or entire organisms. In human cancers, tetraploidization promotes aneuploidy and genomic instability, accelerating tumor progression, metastasis, and drug resistance. These adaptations demand metabolic rewiring, including mitochondrial plasticity. Here, we investigate the relationship between mitochondrial quantity/activity, including the mitochondrial transmembrane potential, the intracellular calcium, and the oxidative stress in diploid versus tetraploid cancer cells (colon, sarcoma, liver) and fungal and yeast models (C. albicans diploid/tetraploid strains; S. cerevisiae haploid/diploid/tetraploid strains). We demonstrate that tetraploid cells, whether from human carcinomas or yeast, exhibit consistently enlarged cell size, elevated mitochondrial content, and heightened metabolic activity compared to diploids. Our findings underscore mitochondrial adaptation as a hallmark of tetraploidization, offering novel therapeutic targets for chromosomally unstable tumors. Full article

Haploid breeding technology offers a promising means of significantly shortening the breeding cycle by rapidly generating homozygous inbred lines. Previous studies have shown that DMP8 is involved in haploid induction across various plant species. In this study, we performed whole-genome identification and bioinformatics analyses to investigate the evolutionary relationships, gene structures, conserved domains, and expression patterns of DMP gene family members in tomato (Solanum lycopersicum), pepper (Capsicum annuum) and eggplant (S. melongena). A total of seven, six, and eight DMP genes were identified in the genomes of tomato, pepper, and eggplant, respectively. All encoded proteins contained the DUF679 domain, and the DMP family members were clustered into three distinct groups. Collinearity analysis revealed species-specific expansions of DMP genes in the Solanaceae family. Phylogenetic analysis indicated that CaDMP8 and SmDMP8 are homologous to SlDMP8, with conserved gene and protein structures, suggesting that CaDMP8 and SmDMP8 are potential targets for developing haploid induction lines. Expression pattern analysis demonstrated that SlDMP4 and SlDMP8 are highly expressed in tomato flower tissues, suggesting their potential functional synergy. This study provides the first comprehensive insight into the evolutionary characteristics and functional diversification of the DMP gene family in Solanaceous vegetables. The findings offer a theoretical foundation for the targeted editing of DMP8 homologs to create haploid induction lines, which is critical for accelerating the genetic improvement of Solanaceous crops. Full article

Genome-wide paternal uniparental isodisomy mosaicism (GWpUPIDM) is an extremely rare condition characterized by varying proportions of an androgenetic cell line across different tissues. It is primarily associated with severe congenital hyperinsulinism (CHI), Beckwith–Wiedemann syndrome (BWS) stigmata, a high risk (69–79%) of developing neoplasia and, in some cases, additional manifestations of multilocus paternal imprinting disorders (MPIDs). We herein report the first Mexican/Latin American female patient GWpUPIDM presenting with non-syndromic CHI requiring subtotal pancreatectomy and persistent but unexplained asymptomatic diffuse hepatopathy. When she was 8.5 years old, whole-exome sequencing (WES) in blood revealed an unexpectedly high (~92%) proportion of regions of homozygosity. DNA profiling confirmed a single haploid set of paternal chromosomes in both biparental and androgenetic cell lines, with varying proportions of the androgenetic lineage in leukocytes (84%), resected pancreas (74%), buccal cells (47%), and hair follicles (0.7%). Additional WES trio analysis using gDNA from the patient’s buccal cells and blood samples from both parents revealed an allelic frequency of ~75% for the paternally inherited variant NM_000158.4(GBE1):c.555+1G>T [ClinVar:632422; dbSNP:rs759707498]. At age 8.5, the patient exhibited no clinical features of BWS, MPIDs, or neoplasia. However, she presented persistent hepatic abnormalities that warrant further investigation to rule out an unmasked glycogen storage disease type IV (OMIM#232500). Our findings emphasize the critical need for early diagnosis of GWpUPIDM using SNP-based microarray or WES with further confirmation through DNA profiling in patients presenting with CHI, placental mesenchymal dysplasia, BWS stigmata, or other MPID-related conditions, including neoplasia, to facilitate timely cancer surveillance and management. Full article

Peanut (Arachis hypogaea L.) is a globally important oilseed cash crop, yet its limited genetic diversity and unique reproductive biology present persistent challenges for conventional crossbreeding. Traditional breeding approaches are often time-consuming and inadequate, mitigating the pace of cultivar development. Essential for double fertilization and programmed cell death (PCD), DUF679 membrane proteins (DMPs) represent a membrane protein family unique to plants. In the present study, a comprehensive analysis of the DMP gene family in peanuts was conducted, which included the identification of 21 family members. Based on phylogenetic analysis, these genes were segregated into five distinct clades (I–V), with AhDMP8A, AhDMP8B, AhDMP9A, and AhDMP9B in clade IV exhibiting high homology with known haploid induction genes. These four candidates also displayed significantly elevated expression in floral tissues compared to other organs, supporting their candidacy for haploid induction in peanuts. Subcellular localization prediction, confirmed through co-localization assays, demonstrated that AhDMPs primarily localize to the plasma membrane, consistent with their proposed roles in the reproductive signaling process. Furthermore, chromosomal mapping and synteny analyses revealed that the expansion of the AhDMP gene family is largely driven by whole-genome duplication (WGD) and segmental duplication events, reflecting the evolutionary dynamics of the tetraploid peanut genome. Collectively, these findings establish a foundational understanding of the AhDMP gene family and highlight promising targets for future applications in haploid induction-based breeding strategies in peanuts. Full article

Drought severely limits crop yield every year, making it critical to clarify the genetic basis of drought tolerance for breeding of improved varieties. As drought tolerance is a complex quantitative trait, we analyzed three phenotypic groups: (1) agronomic traits under well-watered (WW) and water-deficit (WD) conditions, (2) stress tolerance indices of these traits, and (3) phenotypic plasticity, using a multi-parent doubled haploid (DH) population assessed in multi-environment trials. Genome-wide association studies (GWAS) identified 130, 171, and 71 quantitative trait loci (QTL) for the three groups of phenotypes, respectively. Only one QTL was shared among all trait groups, 25 between stress indices and agronomic traits, while the majority of QTL were specific to their group. Functional annotation of candidate genes revealed distinct pathways of the three phenotypic groups. Candidate genes under WD conditions were enriched for stress response and epigenetic regulation, while under WW conditions for protein synthesis and transport, RNA metabolism, and developmental regulation. Stress tolerance indices were enriched for transport of amino/organic acids, epigenetic regulation, and stress response, whereas plasticity showed enrichment for environmental adaptability. Transcriptome analysis of 26 potential candidate genes showed tissue-specific drought responses in leaves, ears, and tassels. Collectively, these results indicated both shared and independent genetic mechanisms underlying drought tolerance, providing novel insights into the complex phenotypes related to drought tolerance and guiding further strategies for molecular breeding in maize. Full article

Auxins play a critical role in establishing the embryo axis and embryonic pattern. Our study aimed to determine the developmental stage of 21-day old oat (Avena sativa L.) haploid embryos, obtained by distant crossing with maize, and examined oat zygotic embryos at different developmental stages for their levels of endogenous indole-3-acetic acid (IAA), its metabolites, and IAA localization. The content of auxin metabolites was determined by HPLC-MS/MS, while IAA visualization in embryos was performed by immunohistochemistry and observed under confocal microscopy. We found that 21-day-old haploid embryos contained half the IAA concentration of age-matched zygotic embryos. Simultaneously, the total conjugated auxins (IAA-Asp, IAA-Glu, meIAA) were higher than in zygotic embryos, regardless of their age. Immunolocalization revealed IAA accumulation in embryos aligned with regions of tissue differentiation (e.g., shoot apical meristem, radicle primordium, and coleptile). We conclude that limited morphogenetic progression, evidenced by microscopic sections accompanied by changes in IAA content and distribution in haploid embryos, indicates a developmental stage earlier than the coleoptilar stage of zygotic embryos which occurs 9 days after pollination. Our findings may be useful in embryo rescue techniques, suggesting modulation of auxin concentration in in vitro culture. Full article

Cannabis sativa L. is a dioecious species known to produce over 1600 chemical constituents, including more than 180 cannabinoids classified into 11 structural groups. These bioactive compounds are predominantly synthesised in the glandular trichomes of female inflorescences. However, sex determination in C. sativa is influenced by both genetic and environmental factors, often leading to the development of male flowers on female plants. This unintended fertilisation reduces cannabinoid yield and increases genetic heterogeneity and challenges in medical cannabis production. Haploid and doubled haploid (DH) technologies offer a promising solution by rapidly generating homozygous lines from gametophytic (e.g., unpollinated ovaries and ovules) or sporophytic tissues (e.g., anthers and microspores) via in vitro culture or chromosome reduction during hybridisation. In land plants, the life cycle alternates between a diploid sporophyte and a haploid gametophyte generation, both capable of mitotic division to form multicellular bodies. A single genome regulates this phase transition and encodes the molecular, genetic, and epigenetic mechanisms that precisely control the developmental processes unique to each generation. While the application of haploid technology in C. sativa remains limited, through recent progress in haploid induction (HI) and CRISPR-based genome editing, the direct modification of haploid gametes or embryos enables the creation of null homozygous lines following chromosome doubling, improving genetic uniformity. Understanding the molecular mechanisms of spontaneous chromosome doubling may further facilitate the development of elite cannabis genotypes. Ultimately, enhancing the efficiency of DH production and optimising genome editing approaches could significantly increase the speed of genetic improvement and cultivar development in Cannabis sativa. Full article

The anther culture-based breeding of rice is a plant tissue culture technique that utilizes rice pollen to rapidly obtain haploid plants. In comparison with traditional breeding methods, this technique shortens the breeding cycle and enables the quick generation of homozygous plants, which is of great significance for the development of new rice varieties and the expansion of germplasm resources. With the advancement of technologies, the use of the anther culture technique in rice breeding has matured and has been applied to the development and utilization of new varieties with high yield, multiple resistances, and superior quality, in combination with other breeding methods. This technique has gained widespread attention globally, with many countries adopting it to create new germplasm resources. This study reviews advances in the rice anther culture technique, the factors influencing anther culture efficiency, and the progress in breeding rice varieties using this technique, as well as analyzes the current challenges and future prospects of anther culture breeding. Full article

Genome editing-mediated haploid inducer systems (HISs) present a promising strategy for enhancing breeding efficiency in legume crops, which are vital for sustainable agriculture due to their nutritional benefits and ability to fix nitrogen. Traditional legume breeding is often slow and complicated by the complexity of legumes’ genomes and the challenges associated with tissue culture. Recent advancements have broadened the applicability of HISs in legume crops, facilitating a reduction in the duration of the breeding cycle. By integrating genome editing technology with haploid breeding systems, researchers can achieve precise genetic modifications and rapidly produce homozygous lines, thereby significantly accelerating the development of desired traits. This review explores the current status and future prospects of genome editing-mediated HISs in legumes, emphasizing the mechanisms of haploid induction; recent breakthroughs; and existing technical challenges. Furthermore, we highlight the necessity for additional research to optimize these systems across various legume species, which has the potential to greatly enhance breeding efficiency and contribute to the sustainability of legume production. Full article

Background: Busulfan is an FDA-approved alkylating drug used in the chemotherapy of advanced acute myeloid leukemia. The precise mechanisms by which Busulfan kills spermatogonia stem cells (SSCs) are not yet completely understood. Methods: Using a murine model, we evaluated Busulfan-induced apoptosis and DNA damage signaling between testis and ovary tissues. We executed RT-qPCR, analyzed single-nuclei RNA sequencing data and performed in situ hybridization for the localization of the gene expression in the tissues. Results: The results indicate that, in contrast to female germ cells, haploid male germ cells undergo significant apoptosis following Busulfan chemotherapy. Moreover, a gene enrichment analysis revealed that reactive oxygen species may activate the inflammatory response in part through the TNF-α/NF-κB signaling pathway. Interestingly, in the testis, the mRNA levels of TNF-α and TAF7 (TATA box-binding protein-associated factor 7) are downregulated, and testosterone levels suppressed. Mechanistically, the promoter of TNF-α has a conserved motif for binding TAF7, which is necessary for its transcriptional activation and may require further in-depth study. We next analyzed the tumorigenic function of TAF7 and revealed that it is highly overexpressed in several types of human cancers, particularly testicular germ cell tumors, and associated with poor patient survival. Therefore, we executed in situ hybridization and single-nuclei RNA sequencing, finding that less TAF7 mRNA is present in SSCs after chemotherapy. Conclusions: Thus, our data indicate a possible function of TAF7 in the regulation of SSCs and spermatogenesis following downregulation by Busulfan. These findings may account for the therapeutic effects of Busulfan and underlie its potential impact on cancer chemotherapy prognosis. Full article

Increasing survival rates of children following cancer treatment have resulted in a significant population of adult survivors with the common side effect of infertility. Additionally, the availability of genetic testing has identified Klinefelter syndrome (classic 47,XXY) as the cause of future male infertility for a significant number of prepubertal patients. This study explores new spermatogonia stem cell (SSC)-based fertility therapies to meet the needs of these patients. Testicular cells were isolated from cryopreserved human testes tissue stored from XY and XXY prepubertal patients and propagated in a two-dimensional culture. Cells were then incorporated into a 3D human testicular organoid (HTO) system. During a 3-week culture period, HTOs maintained their structure, viability, and metabolic activity. Cell-specific PCR and flow cytometry markers identified undifferentiated spermatogonia, Sertoli, Leydig, and peritubular cells within the HTOs. Testosterone was produced by the HTOs both with and without hCG stimulation. Upregulation of postmeiotic germ cell markers was detected after 23 days in culture. Fluorescence in situ hybridization (FISH) of chromosomes X, Y, and 18 identified haploid cells in the in vitro differentiated HTOs. Thus, 3D HTOs were successfully generated from isolated immature human testicular cells from both euploid (XY) and Klinefelter (XXY) patients, supporting androgen production and germ cell differentiation in vitro. Full article

Members of DOMAIN OF UNKNOWN FUNCTION 679 membrane proteins (DMPs) have the DUF679 domain, which plays an important role in the process of plant fertilization. In this study, bioinformatics methods were used to identify and analyze the DMP gene family in pepper. The location of the expression of the DMP gene family was explored according to the transient expression of Nicotiana benthamiana, and its expression patterns in different tissues and abiotic stress treatments were analyzed by qRT-PCR. A total of 17 CaDMP genes were identified from the three capsicum varieties, and sub-cellular localization prediction showed that CaDMPs were located on the cell membrane. Phylogenetic analysis showed that CaDMP5 in subgroup Ⅳ was highly homologous with haploid induction genes in Arabidopsis and maize, and its expression level in reproductive organs was significantly higher than that in other tissues, suggesting that CaDMP5 could be a candidate gene for haploid induction in pepper. The expression of CaDMPs increased to varying degrees after different stress treatments, indicating that the DMP gene plays an important role in plant growth and development. The CaDMP gene family was systematically analyzed in this study, which provided preliminary insights for the further research of Capsicum haploid breeding. Full article

Vanilla orchids are members of the Vanilloideae orchid subfamily, and they hold significant economic value as a spice crop in tropical regions. Despite the presence of 180 known species within this subfamily, commercial production focuses on only three species (Vanilla planifolia, V. odorata, and V. pompona) and one hybrid (V. × tahitensis), prized for their aromatic qualities and bioactive compounds. Limited modern breeding initiatives have been undertaken with vanilla orchids, although recent advancements in genomic research are shedding light on this crop’s potential. The protracted breeding cycle of vanilla, coupled with increasing demand for germplasm, underscores the importance of research and breeding efforts in vanilla. This paper outlines a protocol for haploid production in V. planifolia using unfertilized ovaries in tissue culture conditions. Additionally, we present a methodology to confirm the haploid nature of putative haploid lines through stomatal size comparison, chromosome counting, and flow cytometry analysis, proving the successful development of haploid vanilla plants. These findings contribute to the advancement of breeding programs and genetic improvement strategies for the vanilla industry. Full article

The brown macroalga Saccharina japonica (Laminariales, Phaeophyta) is the most productive cultured seaweed in the world. In order to improve the biosafety of transgenic kelp, it is necessary to develop endogenous constitutive promoters, replacing those of virus origin. In this study, the housekeeping actin gene from S. japonica (SjACT) was found to contain three exons and two introns, representing a unique actin gene structure pattern in brown algae. Additionally, the 5′ upstream region was obtained using genome walking, and fused to the reporter gene lacZ or EGFP to construct promoter-detective vectors. Using an established genetic transformation system, kelps in different life-cycle stages were transformed. The detection results showed that, in the diploid sporophyte stage, the transient expression from the lacZ gene could be observed in the frond, stipe, or holdfast of kelps, indicating a manner of being non-tissue-specific. And, in the haploid gametophyte stage of S. japonica, the fluorescence of the expressed EGFP were detected in vivo in gametophyte cells of both genders. These results indicate that the promoter of the SjACT gene (pSjACT) functions in a constitutive manner and is expected to be a key endogenous element in the genetic manipulation of kelps. Full article

The spatiotemporal expression patterns of genes are crucial for maintaining normal physiological functions in animals. Conditional gene knockout using the cyclization recombination enzyme (Cre)/locus of crossover of P1 (Cre/LoxP) strategy has been extensively employed for functional assays at specific tissue or developmental stages. This approach aids in uncovering the associations between phenotypes and gene regulation while minimizing interference among distinct tissues. Various Cre-engineered mouse models have been utilized in the male reproductive system, including Dppa3-MERCre for primordial germ cells, Ddx4-Cre and Stra8-Cre for spermatogonia, Prm1-Cre and Acrv1-iCre for haploid spermatids, Cyp17a1-iCre for the Leydig cell, Sox9-Cre for the Sertoli cell, and Lcn5/8/9-Cre for differentiated segments of the epididymis. Notably, the specificity and functioning stage of Cre recombinases vary, and the efficiency of recombination driven by Cre depends on endogenous promoters with different sequences as well as the constructed Cre vectors, even when controlled by an identical promoter. Cre mouse models generated via traditional recombination or CRISPR/Cas9 also exhibit distinct knockout properties. This review focuses on Cre-engineered mouse models applied to the male reproductive system, including Cre-targeting strategies, mouse model screening, and practical challenges encountered, particularly with novel mouse strains over the past decade. It aims to provide valuable references for studies conducted on the male reproductive system. Full article