Genetic variation mining of the Chinese mitten crab (Eriocheir sinensis) based on transcriptome data from public databases.

At present, public databases house an extensive repository of transcriptome data, with the volume continuing to grow at an accelerated pace. Utilizing these data effectively is a shared interest within the scientific community. In this study, we introduced a novel strategy that harnesses SNPs and InDels identified from transcriptome data, combined with sample metadata from databases, to effectively screen for molecular markers correlated with traits. We utilized 228 transcriptome datasets of Eriocheir sinensis from the NCBI database and employed the Genome Analysis Toolkit software to identify 96 388 SNPs and 20 645 InDels. Employing the genome-wide association study analysis, in conjunction with the gender information from databases, we identified 3456 sex-biased SNPs and 639 sex-biased InDels. The KOG and KEGG annotations of the sex-biased SNPs and InDels revealed that these genes were primarily involved in the metabolic processes of E. sinensis. Combined with SnpEff annotation and PCR experimental validation, a highly sex-biased SNP located in the Kelch domain containing 4 (Klhdc4) gene, CHR67-6415071, was found to alter the splicing sites of Klhdc4, generating two splice variants, Klhdc4_a and Klhdc4_b. Additionally, Klhdc4 exhibited robust expression across the ovaries, testes, and accessory glands. The sex-biased SNPs and InDels identified in this study are conducive to the development of unisexual cultivation methods for E. sinensis, and the alternative splicing event caused by the sex-biased SNP in Klhdc4 may serve as a potential mechanism for sex regulation in E. sinensis. The analysis strategy employed in this study represents a new direction for the rational exploitation and utilization of transcriptome data in public databases.

Post-implantation analysis of genomic variations in the progeny from developing fetus to birth.

The analysis of genomic variations in offspring after implantation has been infrequently studied. In this study, we aim to investigate the extent of de novo mutations in humans from developing fetus to birth. Using high-depth whole-genome sequencing, 443 parent-offspring trios were studied to compare the results of de novo mutations (DNMs) between different groups. The focus was on fetuses and newborns, with DNA samples obtained from the families' blood and the aspirated embryonic tissues subjected to deep sequencing. It was observed that the average number of total DNMs in the newborns group was 56.26 (54.17-58.35), which appeared to be lower than that the multifetal reduction group, which was 76.05 (69.70-82.40) (F = 2.42, P = 0.12). However, after adjusting for parental age and maternal pre-pregnancy body mass index (BMI), significant differences were found between the two groups. The analysis was further divided into single nucleotide variants (SNVs) and insertion/deletion of a small number of bases (indels), and it was discovered that the average number of de novo SNVs associated with the multifetal reduction group and the newborn group was 49.89 (45.59-54.20) and 51.09 (49.22-52.96), respectively. No significant differences were noted between the groups (F = 1.01, P = 0.32). However, a significant difference was observed for de novo indels, with a higher average number found in the multifetal reduction group compared to the newborn group (F = 194.17, P < 0.001). The average number of de novo indels among the multifetal reduction group and the newborn group was 26.26 (23.27-29.05) and 5.17 (4.82-5.52), respectively. To conclude, it has been observed that the quantity of de novo indels in the newborns experiences a significant decrease when compared to that in the aspirated embryonic tissues (7-9 weeks). This phenomenon is evident across all genomic regions, highlighting the adverse effects of de novo indels on the fetus and emphasizing the significance of embryonic implantation and intrauterine growth in human genetic selection mechanisms.

Ornaments for efficient allele-specific expression estimation with bias correction.

Allele-specific expression plays a crucial role in unraveling various biological mechanisms, including genomic imprinting and gene expression controlled by cis-regulatory variants. However, existing methods for quantification from RNA-sequencing (RNA-seq) reads do not adequately and efficiently remove various allele-specific read mapping biases, such as reference bias arising from reads containing the alternative allele that do not map to the reference transcriptome or ambiguous mapping bias caused by reads containing the reference allele that map differently from reads containing the alternative allele. We present Ornaments, a computational tool for rapid and accurate estimation of allele-specific transcript expression at unphased heterozygous loci from RNA-seq reads while correcting for allele-specific read mapping biases. Ornaments removes reference bias by mapping reads to a personalized transcriptome and ambiguous mapping bias by probabilistically assigning reads to multiple transcripts and variant loci they map to. Ornaments is a lightweight extension of kallisto, a popular tool for fast RNA-seq quantification, that improves the efficiency and accuracy of WASP, a popular tool for bias correction in allele-specific read mapping. In experiments with simulated and human lymphoblastoid cell-line RNA-seq reads with the genomes of the 1000 Genomes Project, we demonstrate that Ornaments improves the accuracy of WASP and kallisto, is nearly as efficient as kallisto, and is an order of magnitude faster than WASP per sample, with the additional cost of constructing a personalized index for multiple samples. Additionally, we show that Ornaments finds imprinted transcripts with higher sensitivity than WASP, which detects imprinted signals only at gene level.

Molecular mechanisms of DNA lesion and repair during antibody somatic hypermutation.

Antibody diversification is essential for an effective immune response, with somatic hypermutation (SHM) serving as a key molecular process in this adaptation. Activation-induced cytidine deaminase (AID) initiates SHM by inducing DNA lesions, which are ultimately resolved into point mutations, as well as small insertions and deletions (indels). These mutational outcomes contribute to antibody affinity maturation. The mechanisms responsible for generating point mutations and indels involve the base excision repair (BER) and mismatch repair (MMR) pathways, which are well coordinated to maintain genomic integrity while allowing for beneficial mutations to occur. In this regard, translesion synthesis (TLS) polymerases contribute to the diversity of mutational outcomes in antibody genes by enabling the bypass of DNA lesions. This review summarizes our current understanding of the distinct molecular mechanisms that generate point mutations and indels during SHM. Understanding these mechanisms is critical for elucidating the development of broadly neutralizing antibodies (bnAbs) and autoantibodies, and has implications for vaccine design and therapeutics.

Single-nucleus multi-omics of Parkinson's disease reveals a glutamatergic neuronal subtype susceptible to gene dysregulation via alteration of transcriptional networks.

The genetic architecture of Parkinson's disease (PD) is complex and multiple brain cell subtypes are involved in the neuropathological progression of the disease. Here we aimed to advance our understanding of PD genetic complexity at a cell subtype precision level. Using parallel single-nucleus (sn)RNA-seq and snATAC-seq analyses we simultaneously profiled the transcriptomic and chromatin accessibility landscapes in temporal cortex tissues from 12 PD compared to 12 control subjects at a granular single cell resolution. An integrative bioinformatic pipeline was developed and applied for the analyses of these snMulti-omics datasets. The results identified a subpopulation of cortical glutamatergic excitatory neurons with remarkably altered gene expression in PD, including differentially-expressed genes within PD risk loci identified in genome-wide association studies (GWAS). This was the only neuronal subtype showing significant and robust overexpression of SNCA. Further characterization of this neuronal-subpopulation showed upregulation of specific pathways related to axon guidance, neurite outgrowth and post-synaptic structure, and downregulated pathways involved in presynaptic organization and calcium response. Additionally, we characterized the roles of three molecular mechanisms in governing PD-associated cell subtype-specific dysregulation of gene expression: (1) changes in cis-regulatory element accessibility to transcriptional machinery; (2) changes in the abundance of master transcriptional regulators, including YY1, SP3, and KLF16; (3) candidate regulatory variants in high linkage disequilibrium with PD-GWAS genomic variants impacting transcription factor binding affinities. To our knowledge, this study is the first and the most comprehensive interrogation of the multi-omics landscape of PD at a cell-subtype resolution. Our findings provide new insights into a precise glutamatergic neuronal cell subtype, causal genes, and non-coding regulatory variants underlying the neuropathological progression of PD, paving the way for the development of cell- and gene-targeted therapeutics to halt disease progression as well as genetic biomarkers for early preclinical diagnosis.

Immunotherapy Applications for Thymine Dimers and WT1 Antigen in Renal Cancers: A Comparative Statistical Analysis.

Renal cell carcinoma (RCC) remains incurable in advanced stages. Biomarkers have proven to be quite useful in cancer therapeutics. Herein, we provide a comparative/integrative statistical analysis of seminal immunohistochemistry (IHC) findings for Wilms' Tumor 1 antigen (WT1) and thymine dimers (TDs), emerging as atypical, yet promising, potential biomarkers for RCCs. We assessed WT1/TD reactivity in adult RCC tumor cells, tumor microenvironment (TME), and tumor-adjacent healthy renal tissue (HRT). WT1 positivity was scarce and strictly nuclear in tumor cells, whereas TD-reactive tumor tissues were prevalent. We report statistically significant positive correlations between the density of reactive RCC cellularity and the intensity of nuclear staining for both biomarkers (WT1 - rho = 0.341, p-value = 0.036; TDs - rho = 0.379, p-value = 0.002). RCC stromal TME TD-positivity was much more frequent than WT1 reactivity, apparently proportional to that of the proper RCC cellularity and facilitated by extensive RCC inflammatory infiltration. TDs exhibited nuclear reactivity for most TME cell lines, while RCC TME WT1 expression was rare and inconsistent. In HRTs, TDs were entirely restricted to renal tubular cells, the likely cellular progenitor of most conventional RCC subtypes. In lieu of proper validation, these early findings have significant implications regarding the origins/biology of RCCs and may inform RCC therapeutics, both accounting for the high frequency of immunotherapy-permissive frameshift indels in RCCs, but also hinting at novel predictive clinical tools for WT1-targeted immunotherapy. Overall, the current study represents a meek yet hopefully significant step towards understanding the molecular biology and potential therapeutic targets of RCCs.

MONET: a database for prediction of neoantigens derived from microsatellite loci.

Background: Microsatellite instability (MSI) secondary to mismatch repair (MMR) deficiency is characterized by insertions and deletions (indels) in short DNA sequences across the genome. These indels can generate neoantigens, which are ideal targets for precision immune interception. However, current neoantigen databases lack information on neoantigens arising from coding microsatellites. To address this gap, we introduce The MicrOsatellite Neoantigen Discovery Tool (MONET). Method: MONET identifies potential mutated tumor-specific neoantigens (neoAgs) by predicting frameshift mutations in coding microsatellite sequences of the human genome. Then MONET annotates these neoAgs with key features such as binding affinity, stability, expression, frequency, and potential pathogenicity using established algorithms, tools, and public databases. A user-friendly web interface (https://monet.mdanderson.org/) facilitates access to these predictions. Results: MONET predicts over 4 million and 15 million Class I and Class II potential frameshift neoAgs, respectively. Compared to existing databases, MONET demonstrates superior coverage (>85% vs. <25%) using a set of experimentally validated neoAgs. Conclusion: MONET is a freely available, user-friendly web tool that leverages publicly available resources to identify neoAgs derived from microsatellite loci. This systems biology approach empowers researchers in the field of precision immune interception.

Forensic features and phylogenetic structure survey of four populations from southwest China via the autosomal insertion/deletion markers.

Insertion/Deletion (InDel) polymorphisms, characterized by their smaller amplicons, reduced mutation rates, and compatibility with the prevalent capillary electrophoresis (CE) platforms in forensic laboratories, significantly contribute to the advancement and application of genetic analysis. Guizhou province in China serves as an important region for investigating the genetic structure, ethnic group origins, and human evolution. However, DNA data and the sampling of present-day populations are lacking, especially about the InDel markers. Here, we reported data on 47 autosomal InDels from 592 individuals from four populations in Guizhou (Han, Dong, Yi, and Chuanqing). Genotyping was performed with the AGCU InDel 50 kit to evaluate their utility for forensic purposes and to explore the population genetic structure. Our findings showed no significant deviations from Hardy-Weinberg and linkage equilibriums. The combined power of discrimination (CPD) and the combined power of exclusion (CPE) for each population demonstrated that the kit could be applied to forensic individual identification and was an effective supplement for parentage testing. Genetic structure analyses, including principal component analysis, multidimensional scaling, genetic distance calculation, STRUCTURE, and phylogenetic analysis, highlighted that the genetic proximity of the studied populations correlates with linguistic, geographical, and cultural factors. The observed genetic variances within four research populations were less pronounced than those discerned between populations across different regions. Notably, the Guizhou Han, Dong, and Chuanqing populations showed closer genetic affiliations with linguistically similar groups than the Guizhou Yi. These results underscore the potential of InDel markers in forensic science and provide insights into the genetic landscape and human evolution in multi-ethnic regions like Guizhou. Key points: InDel markers show promise for forensic individual identification and parentage testing via the AGCU InDel 50 kit.Genetic analysis of Guizhou populations reveals correlations with linguistic, geographical, and cultural factors.Guizhou Han, Dong, and Chuanqing populations showed closer genetic affiliations with linguistically similar groups than the Guizhou Yi.

Genome-Wide Profile of Mutations Induced by Carbon Ion Beam Irradiation of Dehulled Rice Seeds.

As a physical mutagen, carbon ion beam (CIB) irradiation can induce high-frequency mutation, which is user-friendly and environment-friendly in plant breeding. In this study, we resequenced eight mutant lines which were screened out from the progeny of the CIB-irradiated dehulled rice seeds. Among these mutants, CIB induced 135,535 variations, which include single base substitutions (SBSs), and small insertion and deletion (InDels). SBSs are the most abundant mutation, and account for 88% of all variations. Single base conversion is the main type of SBS, and the average ratio of transition and transversion is 1.29, and more than half of the InDels are short-segmented mutation (1-2 bp). A total of 69.2% of the SBSs and InDels induced by CIBs occurred in intergenic regions on the genome. Surprisingly, the average mutation frequency in our study is 9.8 × 10-5/bp and much higher than that of the previous studies, which may result from the relatively high irradiation dosage and the dehulling of seeds for irradiation. By analyzing the mutation of every 1 Mb in the genome of each mutant strain, we found some unusual high-frequency (HF) mutation regions, where SBSs and InDels colocalized. This study revealed the mutation mechanism of dehulled rice seeds by CIB irradiation on the genome level, which will enrich our understanding of the mutation mechanism of CIB radiation and improve mutagenesis efficiency.

Whole genome resequencing unveils low-temperature stress tolerance specific genomic variations in jute (Corchorus sp.).

Jute (Corchorus sp.), a commercially important and eco-friendly crop, is widely cultivated in Bangladesh, India, and China. Some varieties of this tropical plant such as the Corchorus olitorius. Variety accession no. 2015 (acc. 2015) has been found to be low-temperature tolerant. The current study was designed to explore the genome-wide variations present in the tolerant plant acc. 2015 in comparison to the sensitive farmer popular variety Corchorus olitorius var. O9897 using the whole genome resequencing technique. Among different variations, intergenic Single Nucleotide Polymorphism (SNPs) and Insertion-Deletion (InDels) were found in the highest percentage whereas approximately 3% SNPs and 2% InDels were found in exonic regions in both plants. Gene enrichment analysis indicated the presence of acc. 2015 specific SNPs in the genes encoding peroxidase, ER lumen protein retaining receptor, and hexosyltransferase involved in stress response (GO:0006950) which were not present in sensitive variety O9897. Besides, distinctive copy number variation regions (CNVRs) comprising 120 gene loci were found in acc. 2015 with a gain of function from multiple copy numbers but absent in O9897. Gene ontology analysis revealed these gene loci to possess different receptors like kinases, helicases, phosphatases, transcription factors especially Myb transcription factors, regulatory proteins containing different binding domains, annexin, laccase, acyl carrier protein, potassium transporter, and vesicular transporter proteins that are responsible for low temperature induced adaptation pathways in plants. This work of identifying genomic variations linked to cold stress tolerance traits will help to develop successful markers that will pave the way to develop genetically modified cold-resistant jute lines for year-round cultivation to meet the demand for a sustainable fiber crop economy.

Genotyping Zebrafish Point Mutant by Allele-Specific Blocking PCR.

Genotyping zebrafish carrying wild-type, heterozygous, or homozygous copies of a mutant allele is often required for investigating gene specific functions, and is routinely performed to differentiate point mutants. In this study, we describe a modified allele-specific PCR method using an additional blocking primer to promote target sequence amplification while suppressing sequences with single mismatch. Using the tp53m214k point mutant as an example, we show that wild-type, heterozygous, and homozygous zebrafish can be easily distinguished using this simple PCR method, which could be widely adapted for genotyping zebrafish with point mutations or small nucleotide insertions/deletions.

Insertion-Deletion Events Are Depleted in Protein Regions with Predicted Secondary Structure.

A fundamental goal in evolutionary biology and population genetics is to understand how selection shapes the fate of new mutations. Here, we test the null hypothesis that insertion-deletion (indel) events in protein-coding regions occur randomly with respect to secondary structures. We identified indels across 11,444 sequence alignments in mouse, rat, human, chimp, and dog genomes and then quantified their overlap with four different types of secondary structure-alpha helices, beta strands, protein bends, and protein turns-predicted by deep-learning methods of AlphaFold2. Indels overlapped secondary structures 54% as much as expected and were especially underrepresented over beta strands, which tend to form internal, stable regions of proteins. In contrast, indels were enriched by 155% over regions without any predicted secondary structures. These skews were stronger in the rodent lineages compared to the primate lineages, consistent with population genetic theory predicting that natural selection will be more efficient in species with larger effective population sizes. Nonsynonymous substitutions were also less common in regions of protein secondary structure, although not as strongly reduced as in indels. In a complementary analysis of thousands of human genomes, we showed that indels overlapping secondary structure segregated at significantly lower frequency than indels outside of secondary structure. Taken together, our study shows that indels are selected against if they overlap secondary structure, presumably because they disrupt the tertiary structure and function of a protein.

Gamma-rays induced genome wide stable mutations in cowpea deciphered through whole genome sequencing.

PURPOSE: Gamma rays are the most widely exploited physical mutagen in plant mutation breeding. They are known to be involved in the development of more than 60% of global cowpea (Vigna unguiculata (L.) Walp.) mutant varieties. Nevertheless, the nature and type of genome-wide mutations induced by gamma rays have not been studied in cowpea and therefore, the present investigation was undertaken. MATERIALS AND METHODS: Genomic DNAs from three stable gamma rays-induced mutants (large seed size, small seed size and disease resistant mutant) of cowpea cultivar 'CPD103' in M6 generation along with its progenitor were used for Illumina-based whole-genome resequencing. RESULTS: Gamma rays induced a relatively higher frequency (88.9%) of single base substitutions (SBSs) with an average transition to transversion ratio (Ti/Tv) of 3.51 in M6 generation. A > G transitions, including its complementary T > C transitions, predominated the transition mutations, while all four types of transversion mutations were detected with frequencies over 6.5%. Indels (small insertions and deletions) constituted about 11% of the total induced variations, wherein small insertions (6.3%) were relatively more prominent than small deletions (4.8%). Among the indels, single-base indels and, in particular, those involving A/T bases showed a preponderance, albeit indels of up to three bases were detected in low proportions. Distributed across all 11 chromosomes, only a fraction of SBSs (19.45%) and indels (20.2%) potentially altered the encoded amino acids/peptides. The inherent mutation rate induced by gamma rays in cowpea was observed to be in the order of 1.4 × 10-7 per base pair in M6 generation. CONCLUSION: Gamma-rays with a greater tendency to induce SBSs and, to a lesser extent, indels could be efficiently and effectively exploited in cowpea mutation breeding.

InDEL instability in two different tumoral tissues and its forensic significance.

There may be cases where malignant tumor samples can be used for forensic DNA profiling studies. STRs are the first systems preferred in forensic science laboratories for identification purposes. However, genetic instability in tumoral tissues causes STR polymorphism to change, leading to erroneous results. On the other hand, insertion/deletion polymorphism (InDels) are used as genetic markers in forensic science, as they have features that make both STR and SNPs preferable. Although previous studies approved that STR instability is observed in many different tumors, there are only a few studies that have displayed the instability of InDels in tumoral tissues before. In this study, it was aimed to determine whether instability is observed in formalin-fixed paraffin-embedded breast and thyroid tumoral tissues at 36plex InDel Panel. A total of 47 cases, 26 of which were diagnosed as breast cancer and 21 as thyroid cancer, were included in the study. In 21 of 26 (80.76%) breast cancers mutational changes were observed, however only 6 of 21 (28.57%) thyroid carcinoma cases displayed instability.Moreover, in these six cases, mutations were detected at only 1 or 2 loci. The most common change in both tissues was loss of heterozygosity. These findings suggest that paraffin embedded tissues of thyroid tumor can be used in cases of forensic genetic identification, however paraffin embedded breast cancer tissues should be examined with care. In conclusion, low InDel mutation rates compared to STR instability, make InDel analysis from paraffin blocks suitable for forensic genetic identification. However, researchers should keep in mind that there may be differences between the profiles of the tumoral tissues taken as reference and the actual case. In addition, by incorporating additional markers such as SNPs and microhaplotypes with low mutation rates into the study alongside Indels, researchers can significantly enhance the discrimination power in identification processes.

New Genera and Species of Caulobacter and Brevundimonas Bacteriophages Provide Insights into Phage Genome Evolution.

Previous studies have identified diverse bacteriophages that infect Caulobacter vibrioides strain CB15 ranging from small RNA phages to four genera of jumbo phages. In this study, we focus on 20 bacteriophages whose genomes range from 40 to 60 kb in length. Genome comparisons indicated that these diverse phages represent six Caulobacter phage genera and one additional genus that includes both Caulobacter and Brevundimonas phages. Within species, comparisons revealed that both single base changes and inserted or deleted genetic material cause the genomes of closely related phages to diverge. Among genera, the basic gene order and the orientation of key genes were retained with most of the observed variation occurring at ends of the genomes. We hypothesize that the nucleotide sequences of the ends of these phage genomes are less important than the need to maintain the size of the genome and the stability of the corresponding mRNAs.

Phylogenomic and molecular markers based studies on clarifying the evolutionary relationships among Peptoniphilus species. Identification of several Genus-Level clades of Peptoniphilus species and transfer of some Peptoniphilus species to the genus Aedoeadaptatus.

To clarify the evolutionary relationships among Peptoniphilus species, whose members show association with increased risk for prostate cancer, detailed phylogenomic and comparative analyses were conducted on their genome sequences. In phylogenetic trees based on core genome proteins and 16S rRNA gene sequences, Peptoniphilus species formed eight distinct clades, with Aedoeadaptatus and Anaerosphaera species branching between them. The observed clades designated as Peptoniphilus sensu stricto (encompassing its type species), Harei, Lacrimalis, Duerdenii, Mikwangii, Stercorisuis, Catoniae and Aedoeadaptatus, show genus level divergence based on 16S rRNA similarity and average amino acid identity (AAI). The Genome Taxonomy Database also assigns most of these clades to distinct taxa. Several Peptoniphilus species (viz. P. coxii, P. ivorii, P. nemausensis and some non-validly published species) grouped reliably with the type species of Aedoeadaptatus (A. acetigenes) and are affiliated to this genus based on 16S rRNA similarity, AAI, and multiple uniquely shared molecular signatures. Hence, we are proposing the transfer of these species into the emended genus Aedoeadaptatus. Our analyses on protein sequences from Peptoniphilus genomes have also identified 54 novel molecular markers consisting of conserved signature indels (CSIs), which are specific for different Peptoniphilus species clades and provide reliable means for their demarcation in molecular terms. Lastly, we also show that based on the shared presence of these CSIs in the genomes of uncharacterized Peptoniphilus spp. (cultured and uncultured), their affiliations to the specific Peptoniphilus clades can be accurately predicted. These results should prove useful in understanding the potential involvement of Peptoniphilus-related species in diseases.

Genomic and Epigenomic Changes in the Progeny of Cold-Stressed Arabidopsis thaliana Plants.

Plants are continuously exposed to various environmental stresses. Because they can not escape stress, they have to develop mechanisms of remembering stress exposures somatically and passing it to the progeny. We studied the Arabidopsis thaliana ecotype Columbia plants exposed to cold stress for 25 continuous generations. Our study revealed that multigenerational exposure to cold stress resulted in the changes in the genome and epigenome (DNA methylation) across generations. Main changes in the progeny were due to the high frequency of genetic mutations rather than epigenetic changes; the difference was primarily in single nucleotide substitutions and deletions. The progeny of cold-stressed plants exhibited the higher rate of missense non-synonymous mutations as compared to the progeny of control plants. At the same time, epigenetic changes were more common in the CHG (C = cytosine, H = cytosine, adenine or thymine, G = guanine) and CHH contexts and favored hypomethylation. There was an increase in the frequency of C to T (thymine) transitions at the CHH positions in the progeny of cold stressed plants; because this type of mutations is often due to the deamination of the methylated cytosines, it can be hypothesized that environment-induced changes in methylation contribute to mutagenesis and may be to microevolution processes and that RNA-dependent DNA methylation plays a crucial role. Our work supports the existence of heritable stress response in plants and demonstrates that genetic changes prevail.

Phylogenomics and plastomics offer new evolutionary perspectives on Kalanchoideae (Crassulaceae).

BACKGROUND AND AIMS: Kalanchoideae is one of three subfamilies within Crassulaceae and contains four genera. Despite previous efforts, the phylogeny of Kalanchoideae remains inadequately resolved with persistent issues including low support, unstructured topologies and polytomies. This study aimed to address two central objectives: (1) resolving the pending phylogenetic questions within Kalanchoideae by using organelle-scale 'barcodes' (plastomes) and nuclear data; and (2) investigating interspecific diversity patterns among Kalanchoideae plastomes. METHODS: To explore the plastome evolution in Kalanchoideae, we newly sequenced 38 plastomes representing all four constituent genera (Adromischus, Cotyledon, Kalanchoe and Tylecodon). We performed comparative analyses of plastomic features, including GC and gene contents, gene distributions at the IR (inverted repeat) boundaries, nucleotide divergence, plastomic tRNA (pttRNA) structures and codon aversions. Additionally, phylogenetic inferences were inferred using both the plastomic dataset (79 genes) and nuclear dataset (1054 genes). KEY RESULTS: Significant heterogeneities were observed in plastome lengths among Kalanchoideae, strongly correlated with LSC (large single copy) lengths. Informative diversities existed in the gene content at SSC/IRa (small single copy/inverted repeat a), with unique patterns individually identified in Adromischus leucophyllus and one major Kalanchoe clade. The ycf1 gene was assessed as a shared hypervariable region among all four genera, containing nine lineage-specific indels. Three pttRNAs exhibited unique structures specific to Kalanchoideae and the genera Adromischus and Kalanchoe. Moreover, 24 coding sequences revealed a total of 41 lineage-specific unused codons across all four constituent genera. The phyloplastomic inferences clearly depicted internal branching patterns in Kalanchoideae. Most notably, by both plastid- and nuclear-based phylogenies, our research offers the first evidence that Kalanchoe section Eukalanchoe is not monophyletic. CONCLUSIONS: This study conducted comprehensive analyses on 38 newly reported Kalanchoideae plastomes. Importantly, our results not only reconstructed well-resolved phylogenies within Kalanchoideae, but also identified highly informative unique markers at the subfamily, genus and species levels. These findings significantly enhance our understanding of the evolutionary history of Kalanchoideae.

Polymorphisms in drug-metabolizing genes and urinary bladder cancer susceptibility and prognosis: Possible impacts and future management.

Epidemiological studies have shown the association of genetic variants with risks of occupational and environmentally induced cancers, including bladder (BC). The current review summarizes the effects of variants in genes encoding phase I and II enzymes in well-designed studies to highlight their contribution to BC susceptibility and prognosis. Polymorphisms in genes codifying drug-metabolizing proteins are of particular interest because of their involvement in the metabolism of exogenous genotoxic compounds, such as tobacco and agrochemicals. The prognosis between muscle-invasive and non-muscle-invasive diseases is very different, and it is difficult to predict which will progress worse. Web of Science, PubMed, and Medline were searched to identify studies published between January 1, 2010, and February 2023. We included 73 eligible studies, more than 300 polymorphisms, and 46 genes/loci. The most studied candidate genes/loci of phase I metabolism were CYP1B1, CYP1A1, CYP1A2, CYP3A4, CYP2D6, CYP2A6, CYP3E1, and ALDH2, and those in phase II were GSTM1, GSTT1, NAT2, GSTP1, GSTA1, GSTO1, and UGT1A1. We used the 46 genes to construct a network of proteins and to evaluate their biological functions based on the Reactome and KEGG databases. Lastly, we assessed their expression in different tissues, including normal bladder and BC samples. The drug-metabolizing pathway plays a relevant role in BC, and our review discusses a list of genes that could provide clues for further exploration of susceptibility and prognostic biomarkers.