Alternative splicing of the PECTINESTERASE gene encoding a cell wall-degrading enzyme affects postharvest softening in grape

The firmness of table grape berries is a crucial quality parameter. Despite extensive research on postharvest fruit softening, its precise molecular mechanisms remain elusive. To enhance our comprehension of the underlying molecular factors, we initially identified differentially expressed genes(DEGs) by comparing the transcriptomes of folic acid(FA)-treated and water-treated(CK) berries at different time points. We then analyzed the sequences to detect alternatively spliced(AS) genes associated with postharvest softening. A total of 2,559 DEGs were identified and categorized into four subclusters based on their expression patterns, with subcluster-4 genes exhibiting higher expression in the CK group compared with the FA treatment group. There were 1,045 AS-associated genes specific to FA-treated berries and 1,042 in the CK-treated berries, respectively. Gene Ontology(GO) annotation indicated that the AS-associated genes in CK-treated berries were predominantly enriched in cell wall metabolic processes,particularly cell wall degradation processes. Through a comparison between treatment-associated AS genes and subcluster-4 DEGs, we identified eight genes, including Pectinesterase 2(VvPE2, Vitvi15g00704), which encodes a cell wall-degrading enzyme and was predicted to undergo an A3SS event. The reverse transcription polymerase chain reaction further confirmed the presence of a truncated transcript variant of VvPE2 in the FA-treated berries.Our study provides a comprehensive analysis of AS events in postharvest grape berries using transcriptome sequencing and underscores the pivotal role of VvPE2 during the postharvest storage of grape berries.

Improved genome annotation of Brassica oleracea highlights the importance of alternative splicing

Brassica oleracea has been developed into many important crops,including cabbage,kale,cauliflower,broccoli and so on.The genome and gene annotation of cabbage(cultivar JZS),a representative morphotype of B.oleracea,has been widely used as a common reference in biological research.Although its genome assembly has been updated twice,the current gene annotation still lacks information on untranslated regions(UTRs)and alternative splicing(AS).Here,we constructed a high-quality gene annotation(JZSv3)using a full-length transcriptome acquired by nanopore sequencing,yielding a total of 59452 genes and 75684 transcripts.Additionally,we re-analyzed the previously reported transcriptome data related to the development of different tissues and cold response using JZSv3 as a reference,and found that 3843 out of 11908 differentially expressed genes(DEGs)underwent AS during the development of different tissues and 309 out of 903 cold-related genes underwent AS in response to cold stress.Meanwhile,we also identified many AS genes,including BolLHCB5 and BolHSP70,that displayed distinct expression patterns within variant transcripts of the same gene,highlighting the importance of JZSv3 as a pivotal reference for AS analysis.Overall,JZSv3 provides a valuable resource for exploring gene function,especially for obtaining a deeper understanding of AS regulation mechanisms.

2,3,5,4’-Tetrahydroxystilbene-2-O-b-D-Glucoside modulates CHEK2 and CCND1 alternative splicing to inhibit MCF-7 cells proliferation

Background:In our previous study,we observed a synergistic effect of 2,3,5,4’-Tetrahydroxystilbene-2-O-b-D-glucoside combined with adriamycin to induce apoptosis in MCF-7 breast cancer cells.However,the underlying mechanisms of epigenetic modifications,such as alternative splicing,have not been explored.In this study,we aimed to investigate the mechanism by which THSG inhibits MCF-7 cell proliferation using full-length transcriptome sequencing.Methods:First,cell viability was examined using the methyl thiazolyl tetrazolium method and full-length transcriptome sequencing was performed to identify genes and pathways.Gene Ontology and Kyoto Encyclopedia of Genes and Genomes were used to identify the principal pathways and targets of THSG.Flow cytometry analysis of cell cycle distribution was performed.Meanwhile,the analysis of alternative splicing and domains of the key proteins was conducted.Quantitative polymerase chain reaction and western blotting were performed for verification.Results:THSG showed significant cytotoxic activity in MCF-7 cells.Full-length transcriptome sequencing revealed differential alternative splicing with 173 upregulated and 263 downregulated genes.Further analysis identified distinct differential expression of genes(CHEK2-211 and CCND1-201)involved in the cell cycle in the THSG-treated group.Subsequently,alternative splicing types of CHEK2(mutually exclusive exon)and CCND1(intron retention).We found that THSG downregulated mRNA expression,as confirmed by quantitative polymerase chain reaction analysis.Interestingly,protein structural analysis revealed that THSG treatment led to the generation of CHK2-211,which was the result of a mutation in the amino acid residues(GLU-150,ASN-151)of the CHEK2 domain(VAL-150,GLY-151).and CyclinD1-201 were obtained when an amino acid(ASP-267)in the domain was lost in CyclinD1.Moreover,molecular docking analysis demonstrated that the domains of key proteins could bind THSG more effectively,with no difference in affinity.Western blotting confirmed that THSG inhibited the expression of CHK2 and CyclinD1.Conclusion:THSG modulated the alternative splicing of CHEK2 and CCND1 by inducing G0/G1 cell cycle arrest,consequently suppressing MCF-7 cell proliferation.

Molecular targets and mechanisms of different aberrant alternative splicing in metastatic liver cancer

Metastasis remains a major challenge in the successful management of malignant diseases.The liver is a major site of metastatic disease and a leading cause of death from gastrointestinal malignancies such as colon,stomach,and pancreatic cancers,as well as melanoma,breast cancer,and sarcoma.As an important factor that influences the development of metastatic liver cancer,alternative splicing drives the diversity of RNA transcripts and protein subtypes,which may provide potential to broaden the target space.In particular,the dysfunction of splicing factors and abnormal expression of splicing variants are associated with the occurrence,progression,aggressiveness,and drug resistance of cancers caused by the selective splicing of specific genes.This review is the first to provide a detailed summary of the normal splicing process and alterations that occur during metastatic liver cancer.It will cover the role of alternative splicing in the mechanisms of metastatic liver cancer by examining splicing factor changes,abnormal splicing,and the contribution of hypoxia to these changes during metastasis.

A geraniol synthase regulates plant defense via alternative splicing in tea plants

Geraniol is an important contributor to the pleasant floral scent of tea products and one of the most abundant aroma compounds in tea plants;however,its biosynthesis and physiological function in response to stress in tea plants remain unclear.The proteins encoded by the full-length terpene synthase(CsTPS1)and its alternative splicing isoform(CsTPS1-AS)could catalyze the formation of geraniol when GPP was used as a substrate in vitro,whereas the expression of CsTPS1-AS was only significantly induced by Colletotrichum gloeosporioides and Neopestalotiopsis sp.infection.Silencing of CsTPS1 and CsTPS1-AS resulted in a significant decrease of geraniol content in tea plants.The geraniol content and disease resistance of tea plants were compared when CsTPS1 and CsTPS1-AS were silenced.Down-regulation of the expression of CsTPS1-AS reduced the accumulation of geraniol,and the silenced tea plants exhibited greater susceptibility to pathogen infection than control plants.However,there was no significant difference observed in the geraniol content and pathogen resistance between CsTPS1-silenced plants and control plants in the tea plants infected with two pathogens.Further analysis showed that silencing of CsTPS1-AS led to a decrease in the expression of the defense-related genes PR1 and PR2 and SA pathway-related genes in tea plants,which increased the susceptibility of tea plants to pathogens infections.Both in vitro and in vivo results indicated that CsTPS1 is involved in the regulation of geraniol formation and plant defense via alternative splicing in tea plants.The results of this study provide new insights into geraniol biosynthesis and highlight the role of monoterpene synthases in modulating plant disease resistance via alternative splicing.

A Neural-network-based Alternative Scheme to Include Nonhydrostatic Processes in an Atmospheric Dynamical Core

Here,a nonhydrostatic alternative scheme(NAS)is proposed for the grey zone where the nonhydrostatic impact on the atmosphere is evident but not large enough to justify the necessity to include an implicit nonhydrostatic solver in an atmospheric dynamical core.The NAS is designed to replace this solver,which can be incorporated into any hydrostatic models so that existing well-developed hydrostatic models can effectively serve for a longer time.Recent advances in machine learning(ML)provide a potential tool for capturing the main complicated nonlinear-nonhydrostatic relationship.In this study,an ML approach called a neural network(NN)was adopted to select leading input features and develop the NAS.The NNs were trained and evaluated with 12-day simulation results of dry baroclinic-wave tests by the Weather Research and Forecasting(WRF)model.The forward time difference of the nonhydrostatic tendency was used as the target variable,and the five selected features were the nonhydrostatic tendency at the last time step,and four hydrostatic variables at the current step including geopotential height,pressure in two different forms,and potential temperature,respectively.Finally,a practical NAS was developed with these features and trained layer by layer at a 20-km horizontal resolution,which can accurately reproduce the temporal variation and vertical distribution of the nonhydrostatic tendency.Corrected by the NN-based NAS,the improved hydrostatic solver at different horizontal resolutions can run stably for at least one month and effectively reduce most of the nonhydrostatic errors in terms of system bias,anomaly root-mean-square error,and the error of the wave spatial pattern,which proves the feasibility and superiority of this scheme.

Comprehensive analysis of the full-length transcripts and alternative splicing involved in clubroot resistance in Chinese cabbage

Chinese cabbage is an economically important Brassica vegetable worldwide, and clubroot, which is caused by the soilborne protist plant pathogen Plasmodiophora brassicae is regarded as a destructive disease to Brassica crops. Previous studies on the gene transcripts related to Chinese cabbage resistance to clubroot mainly employed RNA-seq technology,although it cannot provide accurate transcript assembly and structural information. In this study, PacBio RS II SMRT sequencing was used to generate full-length transcriptomes of mixed roots at 0, 2, 5, 8, 13, and 22 days after P. brassicae infection in the clubroot-resistant line DH40R. Overall, 39 376 high-quality isoforms and 26 270 open reading frames(ORFs) were identified from the SMRT sequencing data. Additionally, 426 annotated long noncoding RNAs(lncRNAs),56 transcription factor(TF) families, 1 883 genes with poly(A) sites and 1 691 alternative splicing(AS) events were identified. Furthermore, 1 201 of the genes had at least one AS event in DH40R. A comparison with RNA-seq data revealed six differentially expressed AS genes(one for disease resistance and five for defensive response) that are potentially involved in P. brassicae resistance. The results of this study provide valuable resources for basic research on clubroot resistance in Chinese cabbage.

Workshop “Navigating the Transition: Standards Powering the Journey of Alternative Fuel Infrastructure”

April 18, Brussels, Belgium In light of recent significant regulatory changes within the European Union impacting the transport sector, such as the Alternative Fuels Infrastructure Regulation(AFIR), Re Fuel EU and Fuel EU Maritime, the role of European standardization becomes increasingly critical in facilitating the transition towards the deployment of alternative fuels.

Use of Topical Complementary and Alternative Medicines (CAM) in Diabetic Septic Foot Disease in Makkah Region, Western of Saudi Arabia, an Exploratory Study

Diabetes is a chronic condition that is significantly impacted daily by environment. At least one in five Saudis, suffer from diabetes. “Diabetes is a pandemic of unprecedented magnitude spiraling out of control” (Boulton). Saudi Arabia ranks the second highest in the Middle East, and is the seventh in the world for the prevalence of diabetes (WHO). Diabetes mellitus (DM) has been found to be related to high mortality, morbidity accompanied by poor general health and lower quality of life. Current Saudi diabetes patients’ behaviors, practices, and beliefs regarding foot health problems such as ulcers, are not well investigated. Diabetic patients frequently utilize natural remedies and Herbs for self-medication as a part of complementary and alternative Medicine. Aim of study: To explore the widespread use of the alternative remedies in the local communities of the Western Saudi Arabia, the drives behind using and to focus on the potential adverse effects following their usage. Method: A descriptive, cross-sectional study of 386 diabetic patients with feet problems in Makkah Region, West of Saudi Arabia during the period April - June 2023. Results: Out of the 531 participants, only 386 participants reported diabetic foot disorder (DFD). Most of the participant were above 65 years of age group (24.61 %). Male (52.08%) more than female (47.92%) in the research group. All of the participants had diabetic foot disorders at least once. The prevalence of complementary and alternative medicines (CAM) use among them was 67.88%, as solitary or in combination with hospital medicines. Honey headed the list followed by myrrh and black seeds. Acquaintances advice (59.54%) was the most reason for using the alternative medicine. 60.30% of Diabetic foot patient complained of adverse effects during CAM remedies use. Conclusion: Consumption of CAM remedies among diabetic patients for Diabetic Foot Disorder (DFD) is common. Honey headed the list followed by myrrh and black seeds. Additional wide-scale research is required to establish their actual efficacy, safety and potential adverse out-comes and to generalize the outcomes.

Progression of Corneal Astigmatism after Phaco-Alternative at the University Hospital Center of the Oueme-Plateau Department in Porto-Novo

Objective: To study the postoperative evolution of corneal astigmatism following phaco-alternative at the University Hospital Center of Ouémé-Plateau (UHC-OP) in Porto-Novo. Method: This was a prospective, cross-sectional study with a descriptive and analytical purpose conducted from April 19 to September 20, 2021;a period of 6 months. The study included all patients presenting with senile or non-senile cataracts without a history of corneal trauma and who underwent phaco-alternative during the study period. Results: A total of 62 eyes underwent phaco-alternative. The average age was 63 ± 12 years with a male predominance of 59.7%. Phaco-alternative was performed with 74.2% linear incision and 25.8% smile incision. Perioperative incidents were mainly capsular ruptures with vitreous loss occurring in 8.1% of cases. The mean preoperative astigmatism was 1.29 ± 1.5 D with an axis of 180˚ ± 20˚, indicating with-the-rule astigmatism. The mean postoperative astigmatism was 2.2 ± 1.5 D with an axis of 90˚ ± 20˚, indicating against-the-rule astigmatism. The mean induced astigmatism at Day 30 was 1.4 ± 1.2 D for smile incision and 1.8 ± 1.2 D for linear incision. Conclusion: Phaco-alternative yields good results with few complications but remains astigmatogenic. The smile incision appears to be less astigmatogenic.

Transformer 2β regulates the alternative splicing of cell cycle regulatory genes to promote the malignant phenotype of ovarian cancer

Late-stage ovarian cancer(OC)has a poor prognosis and a high metastasis rate,but the underlying molecular mechanism is unclear.RNA binding proteins(RBPs)play important roles in posttranscriptional regulation in the contexts of neoplasia and tumor metastasis.In this study,we explored the molecular functions of a canonical RBP,Transformer 2βhomolog(TRA2B),in cancer cells.TRA2B knockdown in HeLa cells and subsequent wholetranscriptome RNA sequencing(RNA-seq)analysis revealed the TRA2B-regulated alternative splicing(AS)profile.We disrupted TRA2B expression in epithelial OC cells and performed a series of experiments to confirm the resulting effects on OC cell proliferation,apoptosis and invasion.TRA2B-regulated AS was tightly associated with the mitotic cell cycle,apoptosis and several cancer pathways.Moreover,the expression of hundreds of genes was regulated by TRA2B,and these genes were enriched in the functions of cell proliferation,cell adhesion and angiogenesis,which are related to the malignant phenotype of OC.By integrating the alternatively spliced and differentially expressed genes,we found that AS events and gene expression were regulated independently.We then explored and validated the oncogenic functions of TRA2B by knocking down its expression in OC cells.The high TRA2B expression was associated with poor prognosis in patients with OC.In ovarian tissue,TRA2B expression showed a gradual increasing trend with increasing malignancy.We demonstrated the important roles of TRA2B in ovarian neoplasia and aggressive OC behaviors and identified the underlying molecular mechanisms,facilitating the targeted treatment of OC.

Dynamic regulation of alternative splicing and chromatin structure in Drosophila gonads revealed by RNA-seq

Both transcription and post-transcriptional processes, such as alternative splicing, play crucial roles in controlling developmental programs in metazoans. Recently emerged RNA-seq method has brought our understanding of eukaryotic transcriptomes to a new level, because it can resolve both gene expression level and alternative splicing events simultaneously. To gain a better understanding of cellular differentiation in gonads, we analyzed mRNA profiles from Drosophila testes and ovaries using RNA-seq. We identified a set of genes that have sex-specific isoforms in wild-type （WT） gonads, including several transcription factors. We found that differentiation of sperms from undifferentiated germ cells induced a dramatic downregulation of RNA splicing factors. Our data confirmed that RNA splicing events are significantly more frequent in the undifferentiated cell-enriched bag of marbles （barn） mutant testis, but downregulated upon differentiation in WT testis. Consistent with this, we showed that genes required for meiosis and terminal differentiation in WT testis were mainly regulated at the transcriptional level, but not by alternative splicing. Unexpectedly, we observed an increase in expression of all families of chromatin remodeling factors and histone modifying enzymes in the undifferentiated cell-enriched bam testis. More interestingly, chromatin regulators and histone modifying enzymes with opposite enzymatic activities are coenriched in undifferentiated cells in testis, suggesting that these cells may possess dynamic chromatin architecture. Finally, our data revealed many new features of the Drosophila gonadal transcriptomes, and will lead to a more comprehensive understanding of how differential gene expression and splicing regulate gametogenesis in Drosophila. Our data provided a foundation for the systematic study of gene expression and alternative splicing in many interesting areas of germ cell biology in Droso- phila, such as the molecular basis for sexual dimorphism and the regulation of the proliferation vs terminal differentiation programs in germline stem cell lineages. The GEO accession number for the raw and analyzed RNA-seq data is GSE16960.

Alterations of Alternative Splicing Patterns of Ser/Arg-Rich (SR) Genes in Response to Hormones and Stresses Treatments in Different Ecotypes of Rice (Oryza sativa)

Ser/Arg-rich （SR） genes encode proteins that play pivotal roles in both constitutive and alternative splicing of pre-mRNA. However, not much effort has been made to investigate the alternative splicing of their own pre-mRNA. In this study, we conducted comprehensive analyses of pre-mRNA splicing for 22 SR genes in three rice （Oryza sativa L.） ecotypes indica, japonica andjavanica. Using different ecotypes we characterized the variations in expression and splicing patterns of rice SR genes in different tissues and at different developmental stages. In addition, we compared the divergence in expression and splicing patterns of SR genes from seedlings of different rice ecotypes in response to hormones application and environmental stresses. Our results revealed the complexity of alternative splicing of SR genes in rice. The splicing varies in different tissues, in different ecotypes, in response to stresses and hormones. Thus, our study suggested that SR genes were subjected to sophisticated alternative splicing although their encoding proteins were involved in the splicing process.

Genome-wide Detection and Analysis of Alternative Splicing for Nucleotide Binding Site-Leucine-Rich Repeats Sequences in Rice

Alternative splicing is a major contributor to genomic complexity and proteome diversity, yet the analysis of alternative splicing for the sequence containing nucleotide binding site and leucine-rich repeats （NBS-LRR） domain has not been explored in rice （Oryza sativa L.）. Hidden Markov model （HMM） searches were performed for NBS-LRR domain. 875 NBS-LRR-encoding sequences were obtained from the Institute for Genomic Research （TIGR）. All of them were used to blast Knowledge-based Oryza Molecular Biological Encyclopaedia （KOME）, TIGR rice gene index （TGI）, and Universal Protein Resource （UniProt） to obtain homologous full-length cDNAs （FL-cDNAs）, tentative consensus sequences, and protein sequences. Alternative splicing events were detected from genomic alignment of FL-cDNAs, tentative consensus sequences, and protein sequences, which provide valuable information on splice variants of genes. These sequences were aligned to the corresponding BAC sequences using the Spidey and Sim4 programs and each of the proteins was aligned by tBLASTn. Of the 875 NBS-LRR sequences, 119 （13.6%） sequences had alternative splicing where multiple FL-cDNAs, TGI sequences and proteins corresponded to the same gene. 71 intron retention events, 20 exon skipping events, 16 alternative termination events, 25 alternative initiation events, 12 alternative 5＇ splicing events, and 16 alternative 3＇ splicing events were identified. Most of these alternative splices were supported by two or more transcripts. The data sets are available at http：//www.bioinfor.org. Furthermore, the bioinformatics analysis of splice boundaries showed that exon skipping and intron retention did not exhibit strong consensus. This implies a different regulation mechanism that guides the expression of splice isoforms. This article also presents the analysis of the effects of intron retention on proteins. The C-terminal regions of alternative proteins turned out to be more variable than the N-terminal regions. Finally, tissue distribution and protein localization of alternative splicing were explored. The largest categories of tissue distributions for alternative splicing were shoot and callus. More than one-thirds of protein localization for splice forms was plasma membrane and cytoplasm. All the NBS-LRR proteins for splice forms may have important function in disease resistance and activate downstream signaling pathways.

Magnesium alloys as extremely promising alternatives for temporary orthopedic implants-A review

Mg alloys are emerging as potential and very promising alternatives for replacing permanent metallic implant materials such as steels and titanium in applications where the implants need to be removed following healing through revision surgery.Use of Mg alloys for implant application is seen as a game changer and Mg alloys are almost perfect materials for the future in both engineering and biomedical applications.Present review therefore focuses on highlighting significance of Mg alloys in biomedical field and risks of using permanent metallic implants particularly when the implants are no longer required after the injury is healed.In this review,importance of orthopedic implants in present scenario,serious concern related to accidents that are causing permanent disabilities,demand in orthopedic implant market worldwide,potential applications of Mg based materials and their compatibility in biological environment is presented and discussed.In addition,degradation rate,major reactions associated with Mg based materials and effect of alloying elements on implant performance are also discussed based on in-vivo results.Recent advances in development of Mg alloys through various techniques and their performance in in-vitro conditions are also outlined.Possible ways to eliminate the limitations of Mg alloys include alloying,melt purification,surface alterations,surface modifications,chemical treatment,secondary processing etc.are discussed.Challenges and opportunities for Mg alloys to become ideal implant material is also addressed.

Three new alternative splicing variants of human cytochrome P450 2D6 mRNA in human extratumoral liver tissue

AIM: To identify the new alternative splicing variants of human CYP2D6 in human extratumoral liver tissue with RT-PCR and sequencing. METHODS: Full length of human CYP2D6 cDNAs was amplificated by reverse transcription-polymerase chain reaction (RT-PCR) from a human extratumoral liver tissue and cloned into pGEM-T vector. The cDNA was sequenced. Exons from 1 to 4 of human CYP2D6 cDNAs were also amplificated by RT-PCR from extratumoral liver tissues of 17 human hepatocellular carcinomas. Some RT-PCR products were sequenced. Exons 1 to 4 of CYP2D6 gene were amplified by PCR from extratumoral liver tissue DNA. Two PCR products from extratumoral liver tissues expressing skipped mRNA were partially sequenced. RESULTS: One of the CYP2D6 cDNAs had 470 nucleotides from 79 to 548 (3' portion of exons 1 to 5' portion of exon 4), and was skipped. Exons 1 to 4 of CYP2D6 cDNA were assayed with RT-PCR in 17 extratumoral liver tissues. Both wild type and skipped mRNAs were expressed in 4 samples, only wild type mRNA was expressed in 5 samples, and only skipped mRNA was expressed in 8 samples. Two more variants were identified by sequencing the RT-PCR products of exons 1 to 4 of CYP2D6 cDNA. The second variant skipped 411 nucleotides from 175 to 585. This variant was identified in 4 different liver tissues by sequencing the RT-PCR products. We sequenced partially 2 of the PCR products amplified of CYP2D6 exon 1 to exon 4 from extratumoral liver tissue genomic DNA that only expressed skipped mRNA by RT-PCR. No point mutations around exon 1, intron 1, and exon 4, and no deletion in CYP2D6 gene were detected. The third variant was the skipped exon 3, and 153 bp was lost. CONCLUSION: Three new alternative splicing variants of CYP2D6 mRNA have been identified. They may not be caused by gene mutation and may lose CYP2D6 activity and act as a down-regulator of CYP2D6.

Allele-specific expression and alternative splicing in horse×donkey and cattle×yak hybrids

Divergence of gene expression and alter native splicing is a crucial driving force in the evolution of species;to date, however the molecular mechanism remains unclear. Hybrids of closely related species provide a suitable model to analyze allele-specific expressi on (ASE) and allele-specific alter native splicing (ASS). Analysis of ASE and ASS can uncover the differences in cis-regulatory elements between closely related species, while eliminating interferenee of trans-regulatory elements. Here, we provide a detailed characterization of ASE and ASS from 19 and 10 transcriptome datasets across five tissues from reciprocal-cross hybrids of horsex don key (mule/hi nny) and cattlexyak (dzo), respectively. Results showed that 4.8%-8.7% and 10.8%-16.7% of genes exhibited ASE and ASS, respectively. Notably, IncRNAs and pseudogenes were more likely to show ASE than protein-coding genes. In addition, genes showing ASE and ASS in mule/hinny were found to be involved in the regulation of muscle strength, whereas those of dzo were involved in high-altitude adaptati on. In con clusi on, our study dem on strated that explorati on of genes showing ASE and ASS in hybrids of closely related species is feasible for species evolution research.

Attitudes and understanding of complementary and alternative medicine in cancer care:An exploratory study of patients’perspectives in Karachi,Pakistan

Objective:To evaluate the prevalence and types of complementary and alternative medicine(CAM)modalities among patients with cancer in Karachi,Pakistan.Methods:This descriptive cross-sectional study was conducted from March 2021 to December 2021.Five hundred patients with cancer were invited to participate in the study.Electronic databases,namely,Google scholar,Publons,EMBASE,PubMed,Chinese National Knowledge Infrastructure Database,and ResearchGate was used for questionnaire designed.The self-administered survey included questions on demographic characteristics,education level,socio-economic conditions and information about CAM therapies,prevalence,effectiveness,and common CAM modalities.Statistical analysis was conducted using SPSS software version 22.Results:Out of the 500 invited patients,433(86.6%)successfully completed and returned the questionnaires.In contrast to patients who were with younger,highly educated,professionally active,higher income,and had advanced cancer,time since diagnosis,type of treatment,cancer types and family history are significantly associated with CAM use.The results showed that 59.8%of the participants were acquainted with complementary and/or alternative medicine and considered safe owing to its natural ingredients.The prevalence of CAM usage among cancer patients was 40.9%and the most widely used CAM modality was herbal medicine(27.7%)and dietary supplements(28.8%).Patients used CAM as a complementary therapy to improve the morphological parameter(28.2%),strengthen the immune system(6.8%),and to decrease the side effects of conventional treatment(18.1%).Most of the respondents get the information regarding CAM therapy from the electronic media(43.2%)and the family members(48%)rather than healthcare personnel.Conclusions:Participants used CAM modalities along with the conventional health care practices.Further multicentre studies should be conducted to provide information regarding the usage of CAM therapies and their eventual benefits in patients with cancer.

Alternative splicing of VEGFA,APP and NUMB genes in colorectal cancer

AIM:To investigate alternative splicing in vascular endothelial growth factor A(VEGFA),amyloid beta precursor protein(APP),and Numb homolog(NUMB) in colorectal cancer(CRC).METHODS:Real-time quantitative reverse transcriptase polymerase chain reaction(q RT-PCR) and PCRrestriction fragment length polymorphism analyses were performed to detect the expression of VEGFA,APP,and NUMB mR NA in 20 CRC tissues and matched adjacent normal tissues,as well as their alternative splicing variants.RESULTS:q RT-PCR analysis revealed that the expression of APP,NUMB,and VEGFA 165 b m RNA were significantly downregulated,while VEGFA m RNA was upregulated,in CRC tissues(all P < 0.05).PCRrestriction fragment length polymorphism analysis revealed that the expression of VEGFA 165a/b in CRC tissues was significantly higher than in adjacent normal tissues(P < 0.05).Compared with adjacent normal tissues,the expression of NUMB-PRRS in CRC tissues was significantly decreased(P < 0.05),and the expression of NUMB-PRRL was increased(P < 0.05).CONCLUSION:Alternative splicing of VEGFA,APP,and NUMB may regulate the development of CRC,and represent new targets for its diagnosis,prognosis,and treatment.

Regulation of alternative splicing of Bcl-x by IL-6,GM-CSF and TPA

The splicing of many alternative exons in the precursor messenger RNA (pre-mRNA) is regulated by extracellular factors but the underlying molecular bases remain unclear. Here we report the differential regulation of Bcl-x pre-mRNA splicing by extracellular factors and their distinct requirements for pre-mRNA elements. In K562 leukemia cells, treatment with interleukin-6 (IL-6) or granulocyte-macrophage colony stimulating factor (GM-CSF) reduced the proportion of the Bcl-xL variant mRNA while treatment with 12-O-tetradecanoylphorbol 13-acetate (TPA) had no effect. In U251 glioma cells, however, TPA efficiently increased the Bcl-xL level. These regulations were also seen for a transfected splicing reporter mini-gene. Further analyses of deletion mutants indicate that nucleotides 1-176 of the downstream intron are required for the IL-6 effect, whereas additional nucleotides 177-284 are essential for the GM-CSF effect. As for the TPA effect, only nucleotides 1-76 are required in the downstream intron. Thus, IL-6, GM-CSF and TPA differentially regulate Bcl-x splicing and require specific intronic pre-mRNA sequences for their respective effects.