Frameshift Research Articles

Analysis of a Chinese pedigree with female infertility due to WEE2 gene c.495del homozygous frameshifting variant induced fertilization disorder

To explore the genetic basis for a patient with repeated fertilization failure during assisted reproductive therapy, and to identify the source and mode of mutation. A couple treated at the Center for Reproductive Medicine, Gansu Provincial Maternal and Child Health Care Hospital in January 2024 for infertility with incomplete left tube obstruction was selected as the study subject. Relevant clinical data was collected. The couple was subjected to whole exome sequencing (WES), and the candidate variant was verified by Sanger sequencing of their family members and bioinformatic analysis. WES has identified a homozygous c.495del frameshifting mutation of the WEE2 gene in the female partner, whilst no relevant variant was suspected in the male partner. The elder brother of the female partner was homozygous for the above variant, while her parents, maternal and paternal aunts, uncle, grandmother, and grandmother were heterozygous for it. Based on the guidelines from the American College of Medical Genetics and Genomics, above variant was rated to be pathogenic. The homozygous c.495del frameshifting mutation of the WEE2 gene probably underlay the oocyte fertilization disorder in this couple, which has conformed to an autosomal recessive inheritance.

A case of neonatal Mitochondrial DNA depletion syndrome type 13 caused by FBXL4 gene mutation

To explore the clinical phenotypes and genetic variant in a neonatal case of Mitochondrial DNA depletion syndrome type 13 (MTDPS13). Clinical data and results of genetic testing of a neonate admitted to the Children's Hospital of Zhejiang University School of Medicine in January 2023 was retrospectively analyzed. The study was approved by the Medical Ethics Committee of the Children's Hospital of Zhejiang University. The male infant was admitted to the NICU due to tachypnea and persistent lactic acidosis 6 hours after birth. At admission, distinctive facial features were noted. Laboratory tests showed elevated lactic acid (< 30 mmol/L). Whole-exome sequencing revealed that he has harbored homozygous c.141del frameshift mutation of FBXL4 gene, which was unreported previously. The mutation was inherited from both of his parents and classified as likely pathogenic based on the guidelines from the American College of Medical Genetics and Genomics (ACMG). The clinical phenotypes of this case of MTDPS13 is characterized by lactic acidosis, distinctive facial features, growth retardation and developmental delay, for which the homozygous c.141del variant of the FBXL4 gene may be accountable.

Review: Clinical findings and genetic characterization of children affected with X-linked retinoschisis in the Spanish population.

X-linked retinoschisis (XLRS) is an inherited retinal disorder due to mutations in retinoschisin 1, characterized by impaired central vision secondary to parafoveal cystic cavities and visual field loss by splitting through the retinal nerve fibre layer in the peripheral retina. It is the leading cause of juvenile macular degeneration in males, and currently there is no approved treatment but carbonic anhydrase inhibitors can be used. A retrospective review of the medical records of 17 children with confirmed XLRS seen in the Paediatric Ophthalmology Department of La Paz University Hospital from the 1st of January 2009 to the 1st of June of 2023 was conducted. Complete ophthalmological studies, genetic testing and full-field electroretinogram were performed. Topical brinzolamide was given to patients with foveoschisis, adding oral acetazolamide in those who did not improve with topical treatment alone or with very extensive foveoschisis at diagnosis. Surgical treatment was performed in retinal detahment (RD) or in no clearing hemovitreous cases. Mean age at diagnosis was 5,86 years and the most common reason for consultation was strabismus, followed by RD. The most frequently affected retinal later on Optic coherence tomography was the inner nuclear layer and throughout the follow-up we observed a decrease in central macular thickness. We found some genotype-phenotype correlation in our series and more severe phenotypes if the first amino acid of the protein is affected or in frameshift mutations. We found that medical treatment (topical and oral) improves foveoschisis, and that surgery shows poor outcomes, especially in younger patients.

Roles and regulatory patterns of protein isoforms in plant adaptation and development.

Protein isoforms (PIs) play pivotal roles in regulating plant growth and development that confer adaptability to diverse environmental conditions. PIs are widely present in plants and generated through alternative splicing (AS), alternative polyadenylation (APA), alternative initiation (AI), and ribosomal frameshifting (RF) events. The widespread presence of PIs not only significantly increases the complexity of genomic information but also greatly enriches regulatory networks and enhances their flexibility. PIs may also play important roles in phenotypic diversity, ecological niche differentiation, and speciation, thereby increasing the dimensions of research in molecular ecology. However, PIs pose new challenges for the quantitative analysis, annotation, and identification of genetic regulatory mechanisms. Thus, focus on PIs make genomic and epigenomic studies both more powerful and more challenging. This review summarizes the origins, functions, regulatory patterns of isoforms, and the challenges they present for future research in molecular ecology and molecular biology.

Molecular and Clinical Characterization of a Founder Mutation Causing G6PC3 Deficiency

G6PC3 deficiency is a monogenic immunometabolic disorder that causes severe congenital neutropenia type 4. Patients display heterogeneous extra-hematological manifestations, contributing to delayed diagnosis. Here, we investigated the origin and functional consequence of the G6PC3 c.210delC variant found in patients of Mexican descent. Based on the shared haplotypes amongst mutation carriers, we estimated that this variant originated from a founder effect in a common ancestor. Furthermore, by ancestry analysis, we concluded that it appeared in the indigenous Mexican population. At the protein level, we showed that this frameshift mutation leads to an aberrant protein expression in overexpression and patient-derived Epstein-Barr Virus-immortalized B (EBV-B) cells. The neutropenia observed in G6PC3-deficient patients is driven by the intracellular accumulation of the metabolite 1,5-anhydroglucitol-6-phosphate (1,5-AG6P) that inhibits glycolysis. We characterized how the c.210delC variant impacts glycolysis by performing extracellular flux assays on patient-derived EBV-B cells. When treated with 1,5-anhydroglucitol (1,5-AG), the precursor to 1,5-AG6P, patient cells exhibited markedly reduced engagement of glycolysis. Finally, we compared the clinical presentation of patients with the mutation c.210delC and all other G6PC3-deficient patients reported in the literature, and we found that the c.210delC carriers display all prominent clinical features observed in prior patients. In conclusion, G6PC3 c.210delC is a loss-of-function mutation that arose from a founder effect in the indigenous Mexican population. These findings may facilitate the diagnosis of additional patients in this geographical area. Moreover, the in vitro 1,5-AG-dependent functional assay used in our study could be employed to assess the pathogenicity of additional G6PC3 variants.

Novel low-frequency maternal mosaicism mutation of &lt;i&gt;CHRDL1&lt;/i&gt; gene resulted in the X-linked megalocornea in the offspring

X-linked megalocornea (MGC1) is an inherited disorder resulting from mutations of the CHRDL1 gene. Common symptoms of megalocornea include oversized cornea, deeper fluid-filled space between the cornea and iris, blurred or distorted vision at every distance, leading to mild vision issues. In this report, we present the first cases of megalocornea caused by the maternal low-frequency somatic mosaicism of the novel CHRDL1 mutation. Two Vietnamese siblings, 5 years old (MG001) and 13 years old (MG002) visited the hospital for regular check-ups with manifestations such as myopic, exophthalmia, and enlarged cornea. They were suspected of having inherited megalocornea disorder without glaucoma symptoms. The DNA genome of the proband (MG001) was applied for whole exome sequencing (WES) analysis, and the result revealed a hemizygous pathogenic variant c.943dupA in the CHRDL1 gene, which results in a frameshift mutation (p.Ile315fs) in protein. This mutation was also identified in the proband (MG002) by Sanger sequencing. Notably, the CHRDL1 c.943dupA mutation was recognized at a very low fraction in the maternal DNA after triplicate Sanger sequencing, indicating that the mother has a mosaicism mutation of CHRDL1 c.943dupA. This study reported a rare case of X-linked megalocornea in a family and provides a novel genetic factor relating to an eye disorder in Vietnam.

Experimental evolution reveals evolutionary bias and its causes

BackgroundSpecies generally exhibits evolutionary bias, adapting towards a specific direction rather than others, yet the underlying causes remains unknown.ResultsHere, we investigated evolutionary bias and its causes by conducting experimental evolution on Escherichia coli. We introduced an E. coli strain (lac-), initially unable to utilize lactose due to a frameshift mutation, into two different culture media: one medium (L) containing ample sodium acetate and lactose as carbon sources, and the other medium (G) containing abundant glucose and lactose as carbon sources. After 20 days of experimental evolution, our findings revealed that all L-populations underwent parallel evolution through reverse mutation to utilize lactose (lac+), resulting in a relatively higher fitness gain compared to utilizing sodium acetate. In contrast, all G-populations did not transition towards lactose utilization but instead continued to utilize glucose, which provides a higher fitness gain than utilizing lactose. These results demonstrate that our experimental populations in L and G media respectively exhibit biased evolution towards utilizing different carbon sources, yet all trajectories converge towards higher fitness gains. When lac+ (lactose-eater) and lac- (acetate-eater) were co-cultured in L medium, all lac- individuals were eventually eliminated, while lac + individuals were consistently selected and retained.ConclusionsOur findings indicate that species tend to evolve with a bias towards directions that offer higher fitness gains, partly because high-fitness-gain directions competitively exclude low-fitness-gain directions.

BSA-seq and transcriptome analysis reveal GhDSA05 as a candidate gene responsible for sensitivity to defoliants in Gossypium hirsutum L.

With the development of machine-harvested cotton, the application of defoliants is a key technology that can promote defoliation and reduce impurities in cotton fibers. Therefore, sensitivity to defoliants is a very important trait in machine-harvested cotton. In this study, to uncover the genetic basis of defoliant sensitivity in cotton, bulked segregant analysis by sequencing was used with a F2 population derived from Xinluzao 57 (XLZ57, defoliant sensitive) and Nan 2 (N2, defoliant insensitive) to mapped the quantitative trait locus (QTL) to a 3.66-Mb interval at chromosome A05. Then, 284 F2 individuals were used to validate and narrow the QTL to a 1.6-Mb interval and was named qSD-A05. To identify the candidate genes, dynamic transcriptome analysis of the abscission zone (AZ) in XLZ57 and N2 after defoliant treatment was performed. A total of 132 and 142 common differentially expressed genes (DEGs) were identified at all time points in N2 and XLZ57, respectively, and were enriched in the defense response pathway and abscisic acid-activated signaling pathway. More importantly, 15 DEGs were identified in the qSD-A05 interval, among which, Ghi_A05G23601 was significantly upregulated after defoliant treatment. There were multiple variants between N2 and XLZ57, including missense and frameshift mutations. Therefore, Ghi_A05G23601 was regarded as the key candidate gene and was named GhDSA05, which encodes a Facilitator superfamily transporter protein. To verify the function of GhDSA05, virus-induced gene silencing was performed, which showed that the GhDSA05-silenced plants exhibited decreased defoliant sensitivity. Additionally, the expression of organ abscission-related genes was downregulated in the GhDSA05-silenced plants. In summary, GhDSA05 positively regulates the formation of AZ under defoliant treatment and promotes leaf abscission in cotton. The results of this study not only enhance our knowledge of the defoliation mechanism of cotton but also provide a target for the genetic improvement of defoliant sensitivity.

Genetic dissection of flour whiteness through genome-wide association analysis in common wheat (Triticum aestivum L.)

The color of flour products has an important influence on consumer acceptance. Flour color is largely determined and measured by the index of flour whiteness (FW) in China. In this study, an association population comprising 207 wheat (Triticum aestivum) accessions originating from seven countries was used for dissection of FW-related genetic loci through genome-wide association analysis. Six quantitative trait loci (QTLs) significantly associated with FW were identified, accounting for 7.87%–16.53% of the total phenotypic variation. Four KASP markers were developed from single-nucleotide polymorphisms associated with the QTLs QFW.HAAS-1AS, QFW.HAAS-1BL, QFW.HAAS-5AL, and QFW.HAAS-7AL. The phytoene synthase-encoding gene TraesCS7A03G1357000 (TaPsyA1) was identified as a candidate gene for QFW.HAAS-7AL. Two allelic variants of TaPsyA1 (designated PsyA1-a and PsyA1-b) were differentiated on the basis of a 37bp insertion/deletion polymorphism in the second intron. PsyA1-b included the 37bp insertion, which led to a translational frameshift in the gene and was associated with higher FW. The PsyA1-a allele lacked the 37bp insertion and was classified into two haplotypes according to the number of repeated ‘TC’ units in a simple sequence repeat in the promoter region. Of the two PsyA1-a haplotypes, the Type 1 haplotype conferred higher FW, flour brightness, and flour redness, and lower yellow pigment content and flour yellowness. The KASP markers and PsyA1 polymorphic markers developed in the present study are suitable for use in molecular marker-assisted selection for improvement of wheat FW.

A single-base deletion in exon 2 of Hd1 delineates monogenic recessive photoperiod insensitivity in aromatic Joha rice: a novel allele for seasonal adaptability

BackgroundAssam's aromatic Joha rice is a unique rice class famous for its aroma, taste, and nutritional benefits, which fetch high market prices in domestic and international markets. Joha landraces are inherently poor yielders due to their strong aroma and predominantly photoperiod sensitivity. Hybridization involving non-aromatic HYVs improves yield with concomitant loss of quality. In this context, mutation breeding, a sustainable approach where genetic mutations are induced to create desirable traits, often provides useful allelic variation in specific morpho-agronomic traits. The present study delves into the genetic characterization of a photoperiod-insensitive mutant. As part of our mutation breeding programme, this mutant was isolated from a gamma ray-induced M2 population of a Joha rice landrace, Kon Joha.ResultsThe mutant was unique, and a single recessive gene conditions the induced photoperiod insensitivity. Mutant gene tagging involved 402 SSR and InDel markers, and later polymorphic markers were used for bulk segregant analysis (BSA) in the F2 population of ‘mutant × Kalijeera (distant parent)’. BSA revealed an association between the SSR marker RM527 and this mutant trait. This marker is present on chromosome 6 of the rice genome. Using chromosome 6-specific SSR markers in polymorphic screening and BSA revealed another associated marker, RM19725, for the mutant trait. The genomic interval between RM527 and RM19725 harbors a photoperiod-insensitive gene, Hd1, on chromosome 6. Cloning and sequencing of Hd1 genomic fragments from the parents and mutants revealed a single-base deletion in exon 2, leading to a frameshift mutation in the Hd1 protein. This mutation in exon 2 leads to severe structural abnormalities in the CCT domain of the Hd1 protein that is critical for the interaction of the repressing complex with conserved response elements in the florigen gene under long-day conditions, thereby causing photoperiod insensitivity.ConclusionsThe mutant's pleasant aroma and other quality characteristics, comparable to those of the parent cultivar, hold significant promise. They expand its potential use in a structured breeding programme aimed at developing high-value aromatic Joha rice. This rice, resilient to winter- and summer-growing environments and with broad seasonal adaptability, could revolutionize the rice market. The practical value of our research is underscored by this exciting possibility.

Chemotherapeutic agents and leucine deprivation induce codon-biased aberrant protein production in cancer.

Messenger RNA (mRNA) translation is a tightly controlled process frequently deregulated in cancer. Key to this deregulation are transfer RNAs (tRNAs), whose expression, processing and post-transcriptional modifications are often altered in cancer to support cellular transformation. In conditions of limiting levels of amino acids, this deregulated control of protein synthesis leads to aberrant protein production in the form of ribosomal frameshifting or misincorporation of non-cognate amino acids. Here, we studied leucine, an essential amino acid coded by six different codons. Surprisingly, we found that leucine deprivation leads to ribosomal stalling and aberrant protein production in various cancer cell types, predominantly at one codon, UUA. Similar effects were observed after treatment with chemotherapeutic agents, implying a shared mechanism controlling the downstream effects on mRNA translation. In both conditions, a limitation in the availability of tRNALeu(UAA) for protein production was shown to be the cause for this dominant effect on UUA codons. The induced aberrant proteins can be processed and immune-presented as neoepitopes and can direct T-cell killing. Altogether, we uncovered a novel mode of interplay between DNA damage, regulation of tRNA availability for mRNA translation and aberrant protein production in cancer that could be exploited for anti-cancer therapy.

Identification and analysis of drought-responsive F-box genes in upland rice and involvement of OsFBX148 in ABA response and ROS accumulation

BackgroundUpland rice varieties exhibit significant genetic diversity and broad environmental adaptability, making them ideal candidates for identifying consistently expressed stress-responsive genes. F-box proteins typically function as part of the SKP1-CUL1-F-box protein (SCF) ubiquitin ligase complexes to precisely regulate gene expression and protein level, playing essential roles in the modulation of abiotic stress responses. Therefore, utilizing upland rice varieties for screening stress-responsive F-box genes is a highly advantageous approach.ResultsThrough mRNA-seq analysis in the Brazilian upland rice (cv. IAPAR9), the research identified 29 drought-responsive F-box genes. Gene distribution and duplication analysis revealed these genes are distributed on 11 of the 12 chromosomes and 10 collinear gene pairs were identified on different chromosomes. 13 cis-elements or binding sites were identified in the promoters of the 29 drought-responsive F-box genes by analysis. Protein domain, stability and subcellular localization analysis results suggest that these F-box proteins possess F-box domain and several other domains, and they are mostly unstable proteins with subcellular localization in cytoplasm, nucleus, chloroplasts, mitochondria and endoplasmic reticulum. Most of drought-responsive F-box genes exhibited expression in various tissues such as root, stem, leaf, leaf sheath and panicle except for OsFBO10 and OsFBX283. These genes exhibited various responses to abiotic stresses such as osmotic, cold, heat, and salt stresses, along with ABA treatment. Importantly, a frame-shift mutation in OsFBX148 was created in the ZH11 variety, leading to altered ABA signal transduction and ROS accumulation. The study further elucidated the interaction of OsFBX148 with SKP1 family proteins OSK4/7/17 to form the SCF complex, dependent on the F-box domain.ConclusionsThe research identified and analyzed 29 drought-responsive F-box genes in upland rice and provides valuable insights into the role of OsFBX148 in ABA and ROS responses. It establishes a basis for future exploration of F-box genes in improving resistance to abiotic stresses, especially drought.

RMI1 is essential for maintaining rice genome stability at high temperature.

Heat is a critical environmental stress for plant survival.One of its harmful effects on the cells is the disruption of genome integrity. However, the mechanisms by which plants cope with heat-induced DNA damage remain largely unknown. RMI1, a component of the RTR (RECQ4-TOP3α-RMI1) complex, plays a pivotal role in maintaining genome stability. In this study, we identified the target gene RMI1by characterizing a high-temperature-sensitive mutant. The growth and development of rmi1-1 seedlings carrying a non-frameshift mutation in RMI1 were hindered at 38°C. Abnormal mitotic chromosome behaviours ultimately led to the cell death of root tips. Additionally, the presence of chromosome fragments during anaphase I caused pollen abortion and sterility in rmi1-1 plants. Yeast two-hybrid assays revealed that the interactions between RMI1-1 and RECQ4 or TOP3α were weakened with increasing temperature and entirely ceased at 36°C. In contrast, the functionalRMI1 maintained its interactions with RECQ4 or TOP3α under the same conditions. These results indicate that the non-frameshift mutation in RMI1 disrupts the formation of the RTR complex at high temperatures, leading to defects in DNA repair and increased sensitivity of rmi1-1 under heat stress. However, embryos of the rmi1-cr2 mutant with a frameshift mutation in RMI1 exhibited complete lethality. In addition, the overexpression of RMI1 enhanced the heat tolerance in rice. These findings provide insights into the molecular mechanisms that RMI1 responds to high temperatures by maintaining genome stability in rice.

Generation of Ext1 Gene-Edited Mice Model Via Dual sgRNAs/Cas9 System and Phenotypic Analyses.

Hereditary multiple exostoses (HME) is an autosomal dominant skeletal disease. Genetic linkage analyses have identified that mutations in the exostosin glycosyltransferase (EXT)1 and EXT2 genes are linked to HME pathogenesis, with EXT1 mutation being the most frequent. The aim of this study was to generate a mice model with Ext1 gene editing to simulate human EXT1 mutation and investigate the genetic pathogenicity of Ext1 through phenotypic analyses. We designed a pair of dual sgRNAs targeting exon 1 of the mice Ext1 gene for precise deletion of a 46bp DNA fragment, resulting in frameshift mutation of the Ext1 gene. The designed dual sgRNAs and Cas9 proteins were injected into mice zygotes cytoplasm. A total of 14 mice were obtained via embryo transfer, among which two genotypic chimera mice had a deletion of the 46bp DNA fragment in exon 1 of the Ext1 gene. By hybridization and breeding, we successfully generated heterozygous mice with edited Ext1 gene (Ext+/-). Off-target effect analysis did not reveal off-target mutations in Ext+/- mice caused by the two sgRNAs used. Compared to wild-type mice, Ext+/- mice exhibited lower body weights. X-ray imaging showed hyperplastic bone near caudal vertebrae only in male Ext+/- mice, with computed tomography values approximately at 200 HU for hyperplastic bone between ribs and spine regions. Furthermore, immunohistochemical analysis revealed fewer articular chondrocytes expressing EXT1 in edited mice compared to wild-type ones. Pathological section analysis demonstrated no structural or morphological abnormalities in heart, liver, lung, or kidney tissues from Ext+/- mice. In conclusion, we successfully generated an accurate DNA deletion model for studying Ext1 using dual sgRNAs/Cas9 systems. In conclusion, we successfully generated precise DNA deletions in the Ext1 mice model using the dual sgRNAs/Cas9 system. In conclusion, we observed significant phenotypic changes in Ext+/- mice, particularly bone hyperplasia in male individuals; however, no exostosis was detected in the gene-edited mice. The introduction of a frameshift mutation into the Ext1 gene through CRISPR/Cas9 technology resulted in novel phenotypic alterations, highlighting the genetic pathogenicity of Ext1. Therefore, our Ext+/- mice serve as a valuable model for further biomedical investigations related to the Ext1 gene.

Detecting transposable elements in long-read genomes using sTELLeR.

Repeat elements, such as transposable elements (TE), are highly repetitive DNA sequences that compose around 50% of the genome. TEs such as Alu, SVA, HERV, and L1 elements can cause disease through disrupting genes, causing frameshift mutations or altering splicing patters. These are elements challenging to characterize using short-read genome sequencing, due to its read length and TEs repetitive nature. Long-read genome sequencing (lrGS) enables bridging of TEs, allowing increased resolution across repetitive DNA sequences. lrGS therefore present an opportunity for improved TE detection and analysis not only from a research perspective but also for future clinical detection. When choosing an lrGS TE caller, parameters such as runtime, CPU hours, sensitivity, precision, and compatibility with inclusion into pipelines are crucial for efficient detection. We therefore developed sTELLeR, (s) Transposable ELement in Long (e) Read, for accurate, fast, and effective TE detection. Particularly, sTELLeR exhibit higher precision and sensitivity for calling of Alu elements than similar tools. The caller is 5-48× as fast and uses <2% of the CPU hours compared to competitive callers. The caller is haplotype aware and output results in a variant call format (VCF) file, enabling compatibility with other variant callers and downstream analysis. sTELLeR is a python-based tool and is available at https://github.com/kristinebilgrav/sTELLeR. Altogether, we show that sTELLeR is a fast, sensitive, and precise caller for detection of TE elements, and can easily be implemented into variant calling workflows.

Enhancing real-world far-field speech with supervised adversarial training

The generalization of speech enhancement models to real-world far-field speech encounters significant challenges, including low signal-to-noise ratio, high reverberation, and variable latency between far-field and near-field recordings. Additionally, using the non-ideal near-field recordings as the labeled desired output further reduces the effectiveness of commonly utilized predictive models. To tackle these challenges, we propose the Far-field to Near-field Speech Enhancement through Supervised Adversarial Training (FNSE-SAT) strategy. This approach leverages supervised adversarial learning via the Multi-Resolution Discriminator, leveraging diverse speech characteristics with different frequency resolutions. A temporal frame shift operation is also incorporated to mitigate alignment discrepancies observed in real-world data and its effectiveness is confirmed by counting the accuracy of Voice Activity Detection. Experimental validation in both causal and non-causal configurations demonstrates that FNSE-SAT significantly outperforms the state-of-the-art predictive model on real-world datasets. Furthermore, adopting the transfer learning strategy, where the model is initialized with a simulated dataset before fine-tuning with real-world data, strengthens the efficacy of FNSE-SAT, leading to superior outcomes. The results of character error rate show that FNSE-SAT generates fewer components that deviate from the textual content compared to the generative diffusion method. Reducing the Discriminator's resolution to a single version decreases the DNSMOS but has a slight effect on the character error rate.

Imaging and Quantifying Ribosomal Frameshifting Dynamics with Single-RNA Precision in Live Cells.

Recent advances in fluorescence microscopy have now made it possible to measure the translation dynamics of individual RNA in living cells and in multiple colors. Here we describe a protocol that exploits these recent advances to simultaneously image the translation of two open reading frames encoded on a single reporter RNA yet frameshifted with respect to each other. This enables precise measurements of frameshifting dynamics and efficiency from specific frameshift stimulatory sequences, all with single-RNA precision.

A novel frameshift mutation in the DIAPH1 gene causes a Chinese family autosomal dominant nonsyndromic hearing loss: Mutation in DIAPH1 causes hearing loss

ObjectiveThis study reports a novel heterozygous, likely truncating mutation in the diaphanous homolog 1 (DIAPH1) gene associated with non-syndromic hearing loss. MethodsFamily members underwent audiological and imaging assessments, whole-exome sequencing (WES), and Sanger sequencing. ResultsSensorineural hearing loss was observed in all five individuals, with severity ranging from mild to severe. None of the affected patients reported vestibular complaints, and routine blood tests showed normal platelet counts. Whole-exome sequencing (WES) revealed a novel frameshift variation, c.3555delA (p.Gln1185Hisfs*3), in exon 26 of the DIAPH1 gene. This variation co-segregated with the hearing-impaired phenotype in the family. The data collected support the classification of c.3555delA as a genetic etiology of hereditary hearing loss according to the American College of Medical Genetics and Genomics guidelines. ConclusionWe identified a novel pathogenic mutation in the DIAPH1 gene, thereby expanding the mutation spectrum associated with hearing loss.

A single upstream mutation of whiB7 underlies amikacin and clarithromycin resistance in Mycobacterium abscessus.

We aimed to investigate the molecular mechanisms underlying the survival of Mycobacterium abscessus when faced with antibiotic combination therapy. By conducting evolution experiments and whole genome sequencing (WGS), we sought to identify genetic variants associated with stress response mechanisms, with a particular focus on drug survival and resistance. We conducted evolution experiments on M. abscessus, exposing the bacteria to a combination therapy of amikacin and rifabutin. Genetic mutations associated with increased antibiotic survival and altered susceptibility were subsequently identified by WGS. We focused on mutations that contribute to stress response mechanisms and tolerance. Of particular interest was a novel frameshift mutation in MAB_3509c, a gene of unknown function within the upstream open reading frame of whiB7. A MAB_3509c knockout mutant was constructed, and expression of downstream drug resistance genes was assessed by RT-qPCR. Mutation of MAB_3509c results in increased RNA levels of whiB7 and downstream stress response genes such as eis2, which is responsible for aminoglycoside resistance. Our findings demonstrate the importance of whiB7 in the adaptive stress response in M. abscessus. Moreover, our results highlight the complexity of M. abscessus adapting to drug stress and underscore the need for further research.

Functional Analysis of CsWOX4 Gene Mutation Leading to Maple Leaf Type in Cucumber (Cucumis sativus L.).

The leaf morphology is an important agronomic trait in crop production. Our study identified a maple leaf type (mlt) cucumber mutant and located the regulatory gene for leaf shape changes through BSA results. Hybrid F1 and F2 populations were generated by F1 self-crossing, and the candidate mlt genes were identified within the 2.8 Mb region of chromosome 2 using map cloning. Through the sequencing and expression analysis of genes within the bulk segregant analysis (BSA) region, we identified the target gene for leaf shape regulation as CsWOX4 (CsaV3_2G026510). The change from base C to T in the original sequence led to frameshift mutations and the premature termination of translation, resulting in shortened encoded proteins and conserved WUSCHEL (WUS) box sequence loss. The specific expression analysis of the CsWOX4/Cswox4 genes in the roots, stems, leaves and other tissue types of wild-type (WT) and mutant plants revealed that CsWOX4 was higher in the root, but Cswox4 (mutant gene) was significantly higher in the leaf. Subcellular localization analysis revealed that CsWOX4 was localized in the nucleus. RNA-seq analysis revealed that the differentially expressed genes were mainly enriched in the mitochondrial cell cycle phase transition, nucleosome and microtubule binding pathways. Simultaneously, the quantitative analysis of the expression trends of 25 typical genes regulating the leaf types revealed the significant upregulation of CsPIN3. In our study, we found that the conserved domain of CsWOX4 was missing in the mutant, and the transcriptome data revealed that the expression of some genes, such as CsPIN3, changed simultaneously, thereby jointly regulating changes in the cucumber leaf type.