Mutations in Frataxin (FXN) cause Friedreich's ataxia (FRDA), a recessive neurodegenerative disorder. Previous studies have proposed that loss of FXN causes mitochondrial dysfunction, which triggers elevated reactive oxygen species (ROS) and leads to the demise of neurons. Here we describe a ROS independent mechanism that contributes to neurodegeneration in fly FXN mutants. We show that loss of frataxin homolog (fh) in Drosophila leads to iron toxicity, which in turn induces sphingolipid synthesis and ectopically activates 3-phosphoinositide dependent protein kinase-1 (Pdk1) and myocyte enhancer factor-2 (Mef2). Dampening iron toxicity, inhibiting sphingolipid synthesis by Myriocin, or reducing Pdk1 or Mef2 levels, all effectively suppress neurodegeneration in fh mutants. Moreover, increasing dihydrosphingosine activates Mef2 activity through PDK1 in mammalian neuronal cell line suggesting that the mechanisms are evolutionarily conserved. Our results indicate that an iron/sphingolipid/Pdk1/Mef2 pathway may play a role in FRDA.

Here, we document a collection of ∼7434 MiMIC (Minos Mediated Integration Cassette) insertions of which 2854 are inserted in coding introns. They allowed us to create a library of 400 GFP-tagged genes. We show that 72% of internally tagged proteins are functional, and that more than 90% can be imaged in unfixed tissues. Moreover, the tagged mRNAs can be knocked down by RNAi against GFP (iGFPi), and the tagged proteins can be efficiently knocked down by deGradFP technology. The phenotypes associated with RNA and protein knockdown typically correspond to severe loss of function or null mutant phenotypes. Finally, we demonstrate reversible, spatial, and temporal knockdown of tagged proteins in larvae and adult flies. This new strategy and collection of strains allows unprecedented in vivo manipulations in flies for many genes. These strategies will likely extend to vertebrates.

The findings that amyotrophic lateral sclerosis (ALS) patients almost universally display pathological mislocalization of the RNA-binding protein TDP-43 and that mutations in its gene cause familial ALS have nominated altered RNA metabolism as a disease mechanism. However, the RNAs regulated by TDP-43 in motor neurons and their connection to neuropathy remain to be identified. Here we report transcripts whose abundances in human motor neurons are sensitive to TDP-43 depletion. Notably, expression of STMN2, which encodes a microtubule regulator, declined after TDP-43 knockdown and TDP-43 mislocalization as well as in patient-specific motor neurons and postmortem patient spinal cord. STMN2 loss upon reduced TDP-43 function was due to altered splicing, which is functionally important, as we show STMN2 is necessary for normal axonal outgrowth and regeneration. Notably, post-translational stabilization of STMN2 rescued neurite outgrowth and axon regeneration deficits induced by TDP-43 depletion. We propose that restoring STMN2 expression warrants examination as a therapeutic strategy for ALS.

Mutations in PLA2G6 (PARK14) cause neurodegenerative disorders in humans, including autosomal recessive neuroaxonal dystrophy and early-onset parkinsonism. We show that loss of iPLA2-VIA, the fly homolog of PLA2G6, reduces lifespan, impairs synaptic transmission, and causes neurodegeneration. Phospholipases typically hydrolyze glycerol phospholipids, but loss of iPLA2-VIA does not affect the phospholipid composition of brain tissue but rather causes an elevation in ceramides. Reducing ceramides with drugs, including myriocin or desipramine, alleviates lysosomal stress and suppresses neurodegeneration. iPLA2-VIA binds the retromer subunits Vps35 and Vps26 and enhances retromer function to promote protein and lipid recycling. Loss of iPLA2-VIA impairs retromer function, leading to a progressive increase in ceramide. This induces a positive feedback loop that affects membrane fluidity and impairs retromer function and neuronal function. Similar defects are observed upon loss of vps26 or vps35 or overexpression of α-synuclein, indicating that these defects may be common in Parkinson disease.

Rhodopsin mistrafficking can cause photoreceptor (PR) degeneration. Upon light exposure, activated rhodopsin 1 (Rh1) in Drosophila PRs is internalized via endocytosis and degraded in lysosomes. Whether internalized Rh1 can be recycled is unknown. Here, we show that the retromer complex is expressed in PRs where it is required for recycling endocytosed Rh1 upon light stimulation. In the absence of subunits of the retromer, Rh1 is processed in the endolysosomal pathway, leading to a dramatic increase in late endosomes, lysosomes, and light-dependent PR degeneration. Reducing Rh1 endocytosis or Rh1 levels in retromer mutants alleviates PR degeneration. In addition, increasing retromer abundance suppresses degenerative phenotypes of mutations that affect the endolysosomal system. Finally, expressing human Vps26 suppresses PR degeneration in Vps26 mutant PRs. We propose that the retromer plays a conserved role in recycling rhodopsins to maintain PR function and integrity.

Friedreich's ataxia (FRDA) is an autosomal recessive neurodegenerative disease caused by mutations in Frataxin (FXN). Loss of FXN causes impaired mitochondrial function and iron homeostasis. An elevated production of reactive oxygen species (ROS) was previously proposed to contribute to the pathogenesis of FRDA. We recently showed that loss of frataxin homolog (fh), a Drosophila homolog of FXN, causes a ROS independent neurodegeneration in flies (Chen et al., 2016). In fh mutants, iron accumulation in the nervous system enhances the synthesis of sphingolipids, which in turn activates 3-phosphoinositide dependent protein kinase-1 (Pdk1) and myocyte enhancer factor-2 (Mef2) to trigger neurodegeneration of adult photoreceptors. Here, we show that loss of Fxn in the nervous system in mice also activates an iron/sphingolipid/PDK1/Mef2 pathway, indicating that the mechanism is evolutionarily conserved. Furthermore, sphingolipid levels and PDK1 activity are also increased in hearts of FRDA patients, suggesting that a similar pathway is affected in FRDA.

Recurrent de novo (DN) and likely gene-disruptive (LGD) mutations contribute significantly to autism spectrum disorders (ASDs) but have been primarily investigated in European cohorts. Here, we sequence 189 risk genes in 1,543 Chinese ASD probands (1,045 from trios). We report an 11-fold increase in the odds of DN LGD mutations compared with expectation under an exome-wide neutral model of mutation. In aggregate, ∼4% of ASD patients carry a DN mutation in one of just 29 autism risk genes. The most prevalent gene for recurrent DN mutations is SCN2A (1.1% of patients) followed by CHD8, DSCAM, MECP2, POGZ, WDFY3 and ASH1L. We identify novel DN LGD recurrences (GIGYF2, MYT1L, CUL3, DOCK8 and ZNF292) and DN mutations in previous ASD candidates (ARHGAP32, NCOR1, PHIP, STXBP1, CDKL5 and SHANK1). Phenotypic follow-up confirms potential subtypes and highlights how large global cohorts might be leveraged to prove the pathogenic significance of individually rare mutations.

The Cullin9 gene encodes a putative E3 ligase that serves a wide variety of biological functions in mitosis, whereas its roles in meiosis have not yet clearly defined. Here, we report that Cullin9 accumulates on the spindle apparatus and colocalizes with the microtubule fibers during mouse oocyte meiotic maturation. Depletion of Cullin9 by morpholino microinjection results in a remarkably higher rate of disorganized spindles and misaligned chromosomes in oocytes, which is coupled with the impaired kinetochore-microtubule attachments. Resultantly, the incidence of aneuploid eggs significantly increases in Cullin9-depleted oocytes. Moreover, we show that Cullin9 controls Survivin's protein level during meiotic maturation, and thus regulates microtubule stability in oocytes. Thus, our study assigns a new meiotic function to Cullin9 and reveals that it prevents mouse eggs from aneuploidy by regulating microtubule dynamics via Survivin.

Advanced maternal age is highly associated with a decline in oocyte quality, but effective approaches to improve it have still not been fully determined. Here, we report that in vivo supplementation of nicotinamide mononucleotide (NMN) efficaciously improves the quality of oocytes from naturally aged mice by recovering nicotinamide adenine dinucleotide (NAD+) levels. NMN supplementation not only increases ovulation of aged oocytes but also enhances their meiotic competency and fertilization ability by maintaining the normal spindle/chromosome structure and the dynamics of the cortical granule component ovastacin. Moreover, single-cell transcriptome analysis shows that the beneficial effect of NMN on aged oocytes is mediated by restoration of mitochondrial function, eliminating the accumulated ROS to suppress apoptosis. Collectively, our data reveal that NMN supplementation is a feasible approach to protect oocytes from advanced maternal age-related deterioration, contributing to the improvement of reproductive outcome of aged women and assisted reproductive technology.

Autism spectrum disorder (ASD) cases have increased rapidly in recent decades, which is associated with various genetic abnormalities. To provide a better understanding of the genetic factors in ASD, we assessed the global scientific output of the related studies. A total of 2944 studies published between 1997 and 2018 were included by systematic retrieval from the Web of Science (WoS) database, whose scientific landscapes were drawn and the tendencies and research frontiers were explored through bibliometric methods. The United States has been acting as a leading explorer of the field worldwide in recent years. The rapid development of high-throughput technologies and bioinformatics transferred the research method from the traditional classic method to a big data-based pipeline. As a consequence, the focused research area and tendency were also changed, as the contribution of de novo mutations in ASD has been a research hotspot in the past several years and probably will remain one into the near future, which is consistent with the current opinions of the major etiology of ASD. Therefore, more attention and financial support should be paid to the deciphering of the de novo mutations in ASD. Meanwhile, the effective cooperation of multi-research centers and scientists in different fields should be advocated in the next step of scientific research undertaken.

Direct analogs of chemically modified bases that carry important epigenetic information, such as 5-methylcytosine (m5C)/5-methyldeoxycytosine (5mC), 5-hydroxymethylcytosine (hm5C)/5-hydroxymethyldeoxycytosine (5hmC), and N6-methyladenosine (m6A)/N6-methyldeoxyadenosine (6mA), are detected in both RNA and DNA, respectively. The modified base N4-acetylcytosine (ac4C) is well studied in RNAs, but its presence and epigenetic roles in cellular DNA have not been explored.

Neuroblastoma is one of the most devastating forms of childhood cancer. Despite large amounts of attempts in precise survival prediction in neuroblastoma, the prediction efficacy remains to be improved.

Age-related decline in female fertility is a common feature that occurs in the fourth decade of women as a result of a reduction in both oocyte quality and quantity [1]. However, strategies to prevent the deterioration of maternal aged oocytes and relevant mechanisms are still underexplored. Here, we find that the reduced abundance of melatonin in the follicular fluid highly correlates with the advanced maternal age-related aneuploidy. Of note, we show that exposure of oocytes from aged mice both in vitro and in vivo to exogenous melatonin not only eliminates the accumulated reactive oxygen species-induced DNA damage and apoptosis, but also suppresses the occurrence of aneuploidy caused by spindle/chromosome defect that is frequently observed in aged oocytes. Importantly, we reveal that melatonin supplementation reverses the defective phenotypes in aged oocytes through a Sirt1/Sod2-dependent mechanism. Inhibition of Sirt1 activity abolishes the melatonin-mediated improvement of aged oocyte quality. Together our findings provide evidence that supplementation of melatonin is a feasible way to protect oocytes from advanced maternal age-related meiotic defects and aneuploidy, demonstrating the potential for improving the quality of oocytes from aged women and the efficiency of assisted reproductive technology.

Conflicting renal effects of nesiritide have been reported in patients with acute decompensated heart failure. To answer this controversy, we performed a meta-analysis of randomized controlled trials to evaluate the influence of nesiritide on renal function in patients with acute decompensated heart failure.

The comprehensive analysis of biological specimens brings about the demand for capturing the spatial, temporal and spectral dimensions of visual information together. However, such high-dimensional video acquisition faces major challenges in developing large data throughput and effective multiplexing techniques. Here, we report the snapshot hyperspectral volumetric microscopy that computationally reconstructs hyperspectral profiles for high-resolution volumes of ~1000 μm × 1000 μm × 500 μm at video rate by a novel four-dimensional (4D) deconvolution algorithm. We validated the proposed approach with both numerical simulations for quantitative evaluation and various real experimental results on the prototype system. Different applications such as biological component analysis in bright field and spectral unmixing of multiple fluorescence are demonstrated. The experiments on moving fluorescent beads and GFP labelled drosophila larvae indicate the great potential of our method for observing multiple fluorescent markers in dynamic specimens.

Intellectual disability (ID) and autism spectrum disorders (ASD) are genetically heterogeneous, and a significant number of genes have been associated with both conditions. A few mutations in POGZ have been reported in recent exome studies; however, these studies do not provide detailed clinical information. We collected the clinical and molecular data of 25 individuals with disruptive mutations in POGZ by diagnostic whole-exome, whole-genome, or targeted sequencing of 5,223 individuals with neurodevelopmental disorders (ID primarily) or by targeted resequencing of this locus in 12,041 individuals with ASD and/or ID. The rarity of disruptive mutations among unaffected individuals (2/49,401) highlights the significance (p = 4.19 × 10(-13); odds ratio = 35.8) and penetrance (65.9%) of this genetic subtype with respect to ASD and ID. By studying the entire cohort, we defined common phenotypic features of POGZ individuals, including variable levels of developmental delay (DD) and more severe speech and language delay in comparison to the severity of motor delay and coordination issues. We also identified significant associations with vision problems, microcephaly, hyperactivity, a tendency to obesity, and feeding difficulties. Some features might be explained by the high expression of POGZ, particularly in the cerebellum and pituitary, early in fetal brain development. We conducted parallel studies in Drosophila by inducing conditional knockdown of the POGZ ortholog row, further confirming that dosage of POGZ, specifically in neurons, is essential for normal learning in a habituation paradigm. Combined, the data underscore the pathogenicity of loss-of-function mutations in POGZ and define a POGZ-related phenotype enriched in specific features.

Forward genetic screens using chemical mutagens have been successful in defining the function of thousands of genes in eukaryotic model organisms. The main drawback of this strategy is the time-consuming identification of the molecular lesions causative of the phenotypes of interest. With whole-genome sequencing (WGS), it is now possible to sequence hundreds of strains, but determining which mutations are causative among thousands of polymorphisms remains challenging. We have sequenced 394 mutant strains, generated in a chemical mutagenesis screen, for essential genes on the Drosophila X chromosome and describe strategies to reduce the number of candidate mutations from an average of -3500 to 35 single-nucleotide variants per chromosome. By combining WGS with a rough mapping method based on large duplications, we were able to map 274 (-70%) mutations. We show that these mutations are causative, using small 80-kb duplications that rescue lethality. Hence, our findings demonstrate that combining rough mapping with WGS dramatically expands the toolkit necessary for assigning function to genes.

CK2 (casein kinase 2) is a serine/threonine-selective protein kinase that has been involved in a variety of cellular processes such as DNA repair, cell cycle control and circadian rhythm regulation. However, its functional roles in oocyte meiosis have not been fully determined.

Urine testing is an essential clinical diagnostic tool. The presence of urine sediments, typically analyzed through microscopic urinalysis or cell culture, can be indicative of many diseases, including bacterial, parasitic, and yeast infections, as well as more serious conditions like bladder cancer. Current urine analysis diagnostic methods are usually centralized and limited by high cost, inconvenience, and poor sensitivity. Here, we developed a lensless projection imaging optofluidic platform with motion-based particle analysis to rapidly detect urinary constituents without the need for concentration or amplification through culture. A removable microfluidics channel ensures that urine samples do not cross contaminate and the lens-free projection video is captured and processed by a low-cost integrated microcomputer. A motion tracking and analysis algorithm is developed to identify and track moving objects in the flow. Their motion characteristics are used as biomarkers to detect different urine species in near real-time. The results show that this technology is capable of detection of red and white blood cells, Trichomonas vaginalis, crystals, casts, yeast and bacteria. This cost-effective device has the potential to be implemented for timely, point-of-care detection of a wide range of disorders in hospitals, clinics, long-term care homes, and in resource-limited regions.

Citrus species are among the most economically important fruit crops. Physiological characteristics and molecular mechanisms associated with de-etiolation have been partially revealed. However, little is known about the mechanisms controlling the expression and function of genes associated with photosynthesis and chlorophyll biosynthesis in variegated citrus seedlings. The lower biomass, chlorophyll contents, and photosynthetic parameter values recorded for the variegated seedlings suggested that chlorophyll biosynthesis was partially inhibited. Additionally, roots of the variegated seedlings were longer than the roots of green seedlings. We obtained 567.07 million clean reads and 85.05 Gb of RNA-sequencing data, with more than 94.19% of the reads having a quality score of Q30 (sequencing error rate = 0.1%). Furthermore, we detected 4,786 and 7,007 differentially expressed genes (DEGs) between variegated and green Shiranuhi and Huangguogan seedlings. Thirty common pathways were differentially regulated, including pathways related to photosynthesis (GO: 0015979) and the chloroplast (GO: 0009507). Photosynthesis (44 and 63 DEGs), photosynthesis-antenna proteins (14 and 29 DEGs), and flavonoid biosynthesis (16 and 29 DEGs) pathways were the most common KEGG pathways detected in two analyzed libraries. Differences in the expression patterns of PsbQ, PetF, PetB, PsaA, PsaN, PsbP, PsaF, Cluster-2274.8338 (ZIP1), Cluster-2274.38688 (PTC52), and Cluster-2274.78784 might be responsible for the variegation in citrus seedlings. We completed a physiological- and transcriptome-level comparison of the Shiranuhi and Huangguogan cultivars that differ in terms of seedling variegation. We performed mRNA-seq analyses of variegated and green Shiranuhi and Huangguogan seedlings to explore the genes and regulatory pathways involved in the inhibition of chlorophyll biosynthesis and decreases in Chl a and Chl b contents. The candidate genes described herein should be investigated in greater detail to further characterize variegated citrus seedlings.