Identification and validation of seed dormancy loci and candidate genes and construction of regulatory networks by WGCNA in maize introgression lines.

KEY MESSAGE: Seventeen PHS-QTLs and candidate genes were obtained, including eleven major loci, three under multiple environments and two with co-localization by the other mapping methods; The functions of three candidate genes were validated using mutants; nine target proteins and five networks were filtered by joint analysis of GWAS and WGCNA. Seed dormancy (SD) and pre-harvest sprouting (PHS) affect yield, as well as grain and hybrid quality in seed production. Therefore, identification of genetic and regulatory pathways underlying PHS and SD is key to gene function analysis, allelic variation mining and genetic improvement. In this study, 78,360 SNPs by SLAF-seq of 230 maize chromosome segment introgression lines (ILs), PHS under five environments were used to conduct GWAS (genome wide association study) (a threshold of 1/n), and seventeen unreported PHS QTLs were obtained, including eleven QTLs with PVE > 10% and three QTLs under multiple environments. Two QTL loci were co-located between the other two genetic mapping methods. Using differential gene expression analyses at two stages of grain development, gene functional analysis of Arabidopsis mutants, and gene functional analysis in the QTL region, seventeen PHS QTL-linked candidate genes were identified, and their five molecular regulatory networks constructed. Based on the Arabidopsis T-DNA mutations, three candidate genes were shown to regulate for SD and PHS. Meanwhile, using RNA-seq of grain development, the weighted correlation network analysis (WGCNA) was performed, deducing five regulatory pathways and target genes that regulate PHS and SD. Based on the conjoint analysis of GWAS and WGCNA, four pathways, nine target proteins and target genes were revealed, most of which regulate cell wall metabolism, cell proliferation and seed dehydration tolerance. This has important theoretical and practical significance for elucidating the genetic basis of maize PHS and SD, as well as mining of genetic resources and genetic improvement of traits.

Integrated Transcriptomic and Proteomic Analyses of Low-Nitrogen-Stress Tolerance and Function Analysis of ZmGST42 Gene in Maize.

Maize (Zea mays L.) is one of the major staple crops providing human food, animal feed, and raw material support for biofuel production. For its growth and development, maize requires essential macronutrients. In particular, nitrogen (N) plays an important role in determining the final yield and quality of a maize crop. However, the excessive application of N fertilizer is causing serious pollution of land area and water bodies. Therefore, cultivating high-yield and low-N-tolerant maize varieties is crucial for minimizing the nitrate pollution of land and water bodies. Here, based on the analysis of the maize leaf transcriptome and proteome at the grain filling stage, we identified 3957 differentially expressed genes (DEGs) and 329 differentially abundant proteins (DAPs) from the two maize hybrids contrasting in N stress tolerance (low-N-tolerant XY335 and low-N-sensitive HN138) and screened four sets of low-N-responsive genes and proteins through Venn diagram analysis. We identified 761 DEGs (253 up- and 508 down-regulated) specific to XY335, whereas 259 DEGs (198 up- and 61 down-regulated) were specific to HN138, and 59 DEGs (41 up- and 18 down-regulated) were shared between the two cultivars under low-N-stress conditions. Meanwhile, among the low-N-responsive DAPs, thirty were unique to XY335, thirty were specific to HN138, and three DAPs were shared between the two cultivars under low-N treatment. Key among those genes/proteins were leucine-rich repeat protein, DEAD-box ATP-dependent RNA helicase family proteins, copper transport protein, and photosynthesis-related proteins. These genes/proteins were involved in the MAPK signaling pathway, regulating membrane lipid peroxidation, and photosynthesis. Our results may suggest that XY335 better tolerates low-N stress than HN138, possibly through robust low-N-stress sensing and signaling, amplified protein phosphorylation and stress response, and increased photosynthesis efficiency, as well as the down-regulation of 'lavish' or redundant proteins to minimize N demand. Additionally, we screened glutathione transferase 42 (ZmGST42) and performed physiological and biochemical characterizations of the wild-type (B73) and gst42 mutant at the seedling stage. Resultantly, the wild-type exhibited stronger tolerance to low N than the mutant line. Our findings provide a better understanding of the molecular mechanisms underlying low-N tolerance during the maize grain filling stage and reveal key candidate genes for low-N-tolerance breeding in maize.

NQO1 alleviates renal fibrosis by inhibiting the TLR4/NF-κB and TGF-ß/Smad signaling pathways in diabetic nephropathy.

OBJECTIVE: Diabetic nephropathy (DN) is one of the main complications of diabetes, and inflammation and fibrosis play an important role in its progression. NAD(P)H: quinone oxidoreductase 1 (NQO1) protects cells from oxidative stress and damage caused by toxic quinones. In the present study, we aimed to investigate the protective effects of NQO1 against diabetes-induced renal inflammation and fibrosis and the underlying mechanisms. METHODS: In vivo, the kidneys of type 2 diabetes model db/db mice were infected with adeno-associated virus vectors to induce NQO1 overexpression. In vitro, human renal tubular epithelial (HK-2) cells transfected with NQO1 pcDNA3.1(+) were cultured under high-glucose (HG) conditions. Gene and protein expression was assessed by quantitative real-time PCR, Western blotting, immunofluorescence, and immunohistochemical staining. Mitochondrial reactive oxygen species (ROS) were detected with MitoSOX Red. RESULT: Our study revealed that the expression of NQO1 was markedly downregulated and that Toll-like receptor (TLR)4 and TGF-ß1 expression was upregulated in vivo and in vitro under diabetic conditions. Overexpression of NQO1 suppressed proinflammatory cytokine (IL-6, TNF-α, MCP-1) secretion, extracellular matrix (ECM) (collagen IV, fibronectin) accumulation and epithelial-mesenchymal transition (EMT) (α-SMA, E-cadherin) in the db/db mouse kidneys and HG-cultured HK-2 cells. Furthermore, NQO1 overexpression ameliorated HG-induced TLR4/NF-κB and TGF-ß/Smad pathways activation. Mechanistic studies demonstrated that a TLR4 inhibitor (TAK-242) suppressed the TLR4/NF-κB signaling pathway, proinflammatory cytokine secretion, EMT and ECM-related protein expression in HG-exposed HK-2 cells. In addition, we found that the antioxidants N-acetylcysteine (NAC) and tempol increased the expression of NQO1 and decreased the expression of TLR4, TGF-ß1, Nox1, and Nox4 and ROS production in HK-2 cells cultured under HG conditions. CONCLUSIONS: These data suggest that NQO1 alleviates diabetes-induced renal inflammation and fibrosis by regulating the TLR4/NF-κB and TGF-ß/Smad signaling pathways.

Reproductive-Stage Heat Stress in Cereals: Impact, Plant Responses and Strategies for Tolerance Improvement.

Reproductive-stage heat stress (RSHS) poses a major constraint to cereal crop production by damaging main plant reproductive structures and hampering reproductive processes, including pollen and stigma viability, pollination, fertilization, grain setting and grain filling. Despite this well-recognized fact, research on crop heat stress (HS) is relatively recent compared to other abiotic stresses, such as drought and salinity, and in particular, RSHS studies in cereals are considerably few in comparison with seedling-stage and vegetative-stage-centered studies. Meanwhile, climate change-exacerbated HS, independently or synergistically with drought, will have huge implications on crop performance and future global food security. Fortunately, due to their sedentary nature, crop plants have evolved complex and diverse transient and long-term mechanisms to perceive, transduce, respond and adapt to HS at the molecular, cell, physiological and whole plant levels. Therefore, uncovering the molecular and physiological mechanisms governing plant response and tolerance to RSHS facilitates the designing of effective strategies to improve HS tolerance in cereal crops. In this review, we update our understanding of several aspects of RSHS in cereals, particularly impacts on physiological processes and yield; HS signal perception and transduction; and transcriptional regulation by heat shock factors and heat stress-responsive genes. We also discuss the epigenetic, post-translational modification and HS memory mechanisms modulating plant HS tolerance. Moreover, we offer a critical set of strategies (encompassing genomics and plant breeding, transgenesis, omics and agronomy) that could accelerate the development of RSHS-resilient cereal crop cultivars. We underline that a judicious combination of all of these strategies offers the best foot forward in RSHS tolerance improvement in cereals. Further, we highlight critical shortcomings to RSHS tolerance investigations in cereals and propositions for their circumvention, as well as some knowledge gaps, which should guide future research priorities. Overall, our review furthers our understanding of HS tolerance in plants and supports the rational designing of RSHS-tolerant cereal crop cultivars for the warming climate.

Sestrin2 remedies podocyte injury via orchestrating TSP-1/TGF-ß1/Smad3 axis in diabetic kidney disease.

Sestrin2 is identified as a stress-induced protein and could functionate in many aspects. In our study, we investigated the latent impact of Sestrin2 on podocyte injury and its molecular mechanism in vivo and in vitro in diabetic kidney disease (DKD). Sestrin2 was low-expressed in renal biopsies from individuals with DKD, the glomeruli from diabetic mice, and mouse podocytes exposed to high glucose (HG). Sestrin2 overexpression ameliorated HG-induced phenotypic alterations, apoptosis, and oxidative stress in conditionally immortalized mouse podocytes and modulated the activity of Thrombospondin-1 (TSP-1)/transforming growth factor (TGF-ß1)/Smad3 pathway in podocytes. Moreover, TSP-1 inhibitor LSKL or TGF-ß blocker Pirfenidone arrested podocyte injury induced by HG. Streptozotocin (STZ) was employed to render equivalent diabetes in B6-TgN (CMV-Sestrin2) (TgN) and wild-type (WT) control mice. Sestrin2 alleviated increased levels of 24-h urinary protein, blood urea nitrogen, serum creatinine and triglyceride, and urine 8-OHdG in diabetic mice. Podocyte phenotypic alterations, increased expression of apoptosis-associated proteins and podocyte loss were observed in WT but not in diabetic TgN mice, as well as oxidative stress. Additionally, TSP-1/TGF-ß1/Smad3 signaling pathway was also suppressed in glomeruli of diabetic TgN mice. Thus, Sestrin2 mitigates podocyte injury in DKD via orchestrating TSP-1/TGF-ß1/Smad3 pathway, underlining Sestrin2 as a promising therapeutic target for DKD.

Comparative Proteomic Analysis of Two Contrasting Maize Hybrids' Responses to Low Nitrogen Stress at the Twelve Leaf Stage and Function Verification of ZmTGA Gene.

Nitrogen is one of the essential nutrients for plant growth and development. However, large amounts of nitrogen fertilizer not only increase the production costs, but also lead to serious environmental problems. Therefore, it is particularly important to reduce the application of nitrogen fertilizer and develop maize varieties with low nitrogen tolerance. The aim of this study was to determine the phenotypic and proteomic alterations of maize affected by nitrogen deficiency and to elucidate the molecular and physiological mechanisms underpinning maize tolerance to low nitrogen. Two maize hybrids with contrasting low nitrogen tolerance were used as the experimental materials. Maize plants were grown under different nitrogen application levels (N0 and N240) and proteomic analysis performed to analyze leaf differentially abundant proteins (DAPs) under different nitrogen conditions. The results showed that under the nitrogen deficiency condition, the nitrogen content, leaf dry weight, leaf area, and leaf area index of XY335 decreased by 15.58%, 8.83%, 3.44%, and 3.44%, respectively. However, in the variety HN138, the same parameters decreased by 56.94%, 11.97%, 8.79%, and 8.79%, respectively. Through proteomic analysis, we found that the low nitrogen tolerance variety responded to low nitrogen stress through lignin biosynthesis, ubiquitin-mediated proteolysis, and stress defense proteins. Transmembrane transporters were differentially expressed in both hybrids after low nitrogen treatment, suggesting that this was a common response to low nitrogen stress. Using bioinformatics analysis, we selected the key candidate gene (ZmTGA) that was assumed to respond to low nitrogen stress, and its function was characterized by maize mutants. The results showed that when compared with normal nitrogen treatment, the root length of the mutants under low nitrogen treatment increased by 10.1%, while that of the wild-type increased by 14.8%; the root surface area of the wild type under low nitrogen treatment increased by 9.6%, while that of the mutants decreased by 5.2%; the root surface area of the wild type was higher than that of the mutant at both nitrogen levels; and the activities of glutathione and guaiacol peroxidase enzymes in the mutant were lower than those in the wild-type under low nitrogen treatment. In summary, the mutant was less adaptable to a low nitrogen environment than the wild type. Our results provide maize genetic resources and a new direction for a further understanding of maize response to low nitrogen stress.

Effects of transforming growth factor beta-activated kinase 1 (TAK1) on apoptosis of HK-2 cells in the high glucose environment.

To observe the role of transforming growth factor beta-activated kinase 1 (TAK1)/p38 MAPK/TGF-ß1 signal pathway plays in oxidative stress and apoptosis in human renal tubular epithelial cells (HK-2) under high glucose induction. HK-2 cells were cultured in high glucose medium with and without TAK1 inhibitor 5Z-7-oxozeaenol. TUNEL and flow cytometry were used to detect cell apoptosis. The protein expression of TAK1, TGF-ß1, Bax and Bcl-2 was detected by immunofluorescence. Meanwhile, flow cytometry was used to detect the production of reactive oxygen species (ROS), and MitoSOX staining was performed to detect the production of mitochondrial ROS. Moreover, real-time quantitative PCR and Western blotting was used to measure the expression of TAK1, TGF-ß1, NOX1, NOX4 and HO-1, Bax, Bcl-2, p38MAPK, p-p38MAPK and TGF-ß1. Results showed that high glucose up-regulated the protein expression of p-TAK1, p-p38 MAPK and TGF-ß1, which induced the aggravation of oxidative stress by promoting the production of ROS, thus promote the apoptosis in HK-2 cells. However, addition of 5z -7-oxozeaenol in HK-2 cells reversed all the above functions induced by high glucose. Another experimental result also showed that SB203580, a p38MAPK inhibitor can down-regulated TGF-ß1 expression and reduce ROS production, thus alleviate cell apoptosis in TAK1 overexpression group. In summary, high glucose intervention could activate TAK1 and promote apoptosis in HK-2 cells. Inhibition of TAK1 expression could block p38 MAPK/TGF-ß1 signaling pathway and reduce ROS production and oxidative stress, which may be one of the signal pathways of TAK1 to reduce apoptosis of HK-2 cells induced by high glucose.Abbreviations: DN, Diabetic nephropathy; TAK1, transforming growth factor ß-activated kinase-1; TGF-ß, transforming growth factor-ß; NG, normal glucose; HG, high glucose; p38 MAPK, p38 mitogen-activated protein kinase; ROS, reactive oxygen species.

NAD(P)H: quinone oxidoreductase 1 attenuates oxidative stress and apoptosis by regulating Sirt1 in diabetic nephropathy.

BACKGROUND: Diabetic nephropathy (DN) is one of the main complications of diabetes, and oxidative stress plays an important role in its progression. NAD(P)H: quinone oxidoreductase 1 (NQO1) protects cells from oxidative stress and toxic quinone damage. In the present study, we aimed to investigate the protective effects and underlying mechanisms of NQO1 on diabetes-induced renal tubular epithelial cell oxidative stress and apoptosis. METHODS: In vivo, the kidneys of db/db mice, which are a type 2 diabetes model, were infected with adeno-associated virus to induce NQO1 overexpression. In vitro, human renal tubular epithelial cells (HK-2 cells) were transfected with NQO1 pcDNA3.1(+) and cultured in high glucose (HG). Gene and protein expression was assessed by quantitative real-time PCR, western blotting, immunofluorescence analysis, and immunohistochemical staining. Reactive oxygen species (ROS) were examined by MitoSox red and flow cytometry. TUNEL assays were used to measure apoptosis. RESULT: In vivo, NQO1 overexpression reduced the urinary albumin/creatinine ratio (UACR) and blood urea nitrogen (BUN) level in db/db mice. Our results revealed that NQO1 overexpression could significantly increase the ratio of NAD+/NADH and silencing information regulator 1 (Sirt1) expression and block tubular oxidative stress and apoptosis in diabetic kidneys. In vitro, NQO1 overexpression reduced the generation of ROS, NADPH oxidase 1 (Nox1) and Nox4, the Bax/Bcl-2 ratio and the expression of Cleaved Caspase-3 and increased NAD+/NADH levels and Sirt1 expression in HK-2 cells under HG conditions. However, these effects were reversed by the Sirt1 inhibitor EX527. CONCLUSIONS: All these data suggest that NQO1 has a protective effect against oxidative stress and apoptosis in DN, which may be mediated by the regulation of Sirt1 through increasing intracellular NAD+/NADH levels. Therefore, NQO1 may be a new therapeutic target for DN.

Omics-Facilitated Crop Improvement for Climate Resilience and Superior Nutritive Value.

Novel crop improvement approaches, including those that facilitate for the exploitation of crop wild relatives and underutilized species harboring the much-needed natural allelic variation are indispensable if we are to develop climate-smart crops with enhanced abiotic and biotic stress tolerance, higher nutritive value, and superior traits of agronomic importance. Top among these approaches are the "omics" technologies, including genomics, transcriptomics, proteomics, metabolomics, phenomics, and their integration, whose deployment has been vital in revealing several key genes, proteins and metabolic pathways underlying numerous traits of agronomic importance, and aiding marker-assisted breeding in major crop species. Here, citing several relevant examples, we appraise our understanding on the recent developments in omics technologies and how they are driving our quest to breed climate resilient crops. Large-scale genome resequencing, pan-genomes and genome-wide association studies are aiding the identification and analysis of species-level genome variations, whilst RNA-sequencing driven transcriptomics has provided unprecedented opportunities for conducting crop abiotic and biotic stress response studies. Meanwhile, single cell transcriptomics is slowly becoming an indispensable tool for decoding cell-specific stress responses, although several technical and experimental design challenges still need to be resolved. Additionally, the refinement of the conventional techniques and advent of modern, high-resolution proteomics technologies necessitated a gradual shift from the general descriptive studies of plant protein abundances to large scale analysis of protein-metabolite interactions. Especially, metabolomics is currently receiving special attention, owing to the role metabolites play as metabolic intermediates and close links to the phenotypic expression. Further, high throughput phenomics applications are driving the targeting of new research domains such as root system architecture analysis, and exploration of plant root-associated microbes for improved crop health and climate resilience. Overall, coupling these multi-omics technologies to modern plant breeding and genetic engineering methods ensures an all-encompassing approach to developing nutritionally-rich and climate-smart crops whose productivity can sustainably and sufficiently meet the current and future food, nutrition and energy demands.

PP2 Ameliorates Renal Fibrosis by Regulating the NF-κB/COX-2 and PPARγ/UCP2 Pathway in Diabetic Mice.

Renal fibrosis is characterized by glomerulosclerosis and tubulointerstitial fibrosis in diabetic nephropathy (DN). We aimed to evaluate the effects of PP2 on renal fibrosis of DN. GSE33744 and GSE86300 were downloaded from the GEO database. Firstly, 839 DEGs were identified between nondiabetic and diabetic mice renal glomerular samples. COX-2 was selected to assess the effects of PP2 on renal glomerulosclerosis. In db/db mice, PP2 decreased the expression of COX-2, phosphorylated p65, and fibrotic proteins, accompanied with attenuated renal glomerulosclerosis. In cultured glomerular mesangial cells, high glucose- (HG-) induced p65 phosphorylation and COX-2 expression were attenuated by PP2 or NF-κB inhibitor PDTC. PP2, PDTC, or COX-2 inhibitor NS-398 ameliorated abnormal proliferation and expression of fibrotic proteins induced by HG. Secondly, 238 DEGs were identified between nondiabetic and diabetic mice renal cortex samples. UCP2 was selected to assess the effects of PP2 on renal tubulointerstitial fibrosis. In db/db mice, PP2 decreased the expression of PPARγ and UCP2, accompanied with attenuated renal tubulointerstitial fibrosis and EMT. In cultured proximal tubular cells, HG-induced PPARγ and UCP2 expression was inhibited by PP2 or PPARγ antagonist GW9662. PP2, GW9662, or UCP2 shRNA ameliorated HG-induced EMT. These results indicated that PP2 ameliorated renal fibrosis in diabetic mice.

Is it beneficial for patients with pT1-2N1M0 breast cancer to receive postmastectomy radiotherapy? An analysis based on RecurIndex assay.

The benefit of postmastectomy radiotherapy (PMRT) for pT1-2N1M0 breast cancer patients currently remains controversial. This study was conducted to investigate whether pT1-2N1M0 breast cancer patients could benefit from PMRT based on RecurIndex assay. The clinical data of 213 pT1-2N1M0 breast cancer patients were retrospectively analyzed. Through RecurIndex assay, 81 cases were assessed as the low risk, and 132 as the high risk. Compared to low-risk patients, high-risk patients especially those not receiving PMRT had a significantly increased risk of recurrence and metastasis, and worse 7-year local-regional recurrence-free interval (LRFI), distance recurrence-free interval (DRFI) and recurrence-free survival (RFS) rates. PMRT-based subgroup analysis indicated no significant differences between the low-risk patients with and without PMRT in 7-year LRFI, DRFI, RFS and overall survival (OS) rates, but apparent differences were all shown between the high-risk patients with and without PMRT in 7-year LRFI, DRFI, RFS and OS rates. Overall, for pT1-2N1M0 breast cancer patients at low risk of recurrence and metastasis stratified by RecurIndex assay, there may be a phenomenon of no PMRT benefits, while for those at high risk, use of PMRT may produce survival benefits.

Advances in Cereal Crop Genomics for Resilience under Climate Change.

Adapting to climate change, providing sufficient human food and nutritional needs, and securing sufficient energy supplies will call for a radical transformation from the current conventional adaptation approaches to more broad-based and transformative alternatives. This entails diversifying the agricultural system and boosting productivity of major cereal crops through development of climate-resilient cultivars that can sustainably maintain higher yields under climate change conditions, expanding our focus to crop wild relatives, and better exploitation of underutilized crop species. This is facilitated by the recent developments in plant genomics, such as advances in genome sequencing, assembly, and annotation, as well as gene editing technologies, which have increased the availability of high-quality reference genomes for various model and non-model plant species. This has necessitated genomics-assisted breeding of crops, including underutilized species, consequently broadening genetic variation of the available germplasm; improving the discovery of novel alleles controlling important agronomic traits; and enhancing creation of new crop cultivars with improved tolerance to biotic and abiotic stresses and superior nutritive quality. Here, therefore, we summarize these recent developments in plant genomics and their application, with particular reference to cereal crops (including underutilized species). Particularly, we discuss genome sequencing approaches, quantitative trait loci (QTL) mapping and genome-wide association (GWAS) studies, directed mutagenesis, plant non-coding RNAs, precise gene editing technologies such as CRISPR-Cas9, and complementation of crop genotyping by crop phenotyping. We then conclude by providing an outlook that, as we step into the future, high-throughput phenotyping, pan-genomics, transposable elements analysis, and machine learning hold much promise for crop improvements related to climate resilience and nutritional superiority.

Cotton GhSSI2 isoforms from the stearoyl acyl carrier protein fatty acid desaturase family regulate Verticillium wilt resistance.

Lipids are major and essential constituents of plant cells and provide energy for various metabolic processes. However, the function of the lipid signal in defence against Verticillium dahliae, a hemibiotrophic pathogen, remains unknown. Here, we characterized 19 conserved stearoyl-ACP desaturase family proteins from upland cotton (Gossypium hirsutum). We further confirmed that GhSSI2 isoforms, including GhSSI2-A, GhSSI2-B, and GhSSI2-C located on chromosomes A10, D10, and A12, respectively, played a dominant role to the cotton 18:1 (oleic acid) pool. Suppressing the expression of GhSSI2s reduced the 18:1 level, which autoactivated the hypersensitive response (HR) and enhanced cotton Verticillium wilt and Fusarium wilt resistance. We found that low 18:1 levels induced phenylalanine ammonia-lyase-mediated salicylic acid (SA) accumulation and activated a SA-independent defence response in GhSSI2s-silenced cotton, whereas suppressing expression of GhSSI2s affected PDF1.2-dependent jasmonic acid (JA) perception but not the biosynthesis and signalling cascade of JA. Further investigation showed that structurally divergent resistance-related genes and nitric oxide (NO) signal were activated in GhSSI2s-silenced cotton. Taken together, these results indicate that SA-independent defence response, multiple resistance-related proteins, and elevated NO level play an important role in GhSSI2s-regulated Verticillium wilt resistance. These findings broaden our knowledge regarding the lipid signal in disease resistance and provide novel insights into the molecular mechanism of cotton fungal disease resistance.

Global Transcriptome and Weighted Gene Co-expression Network Analyses of Growth-Stage-Specific Drought Stress Responses in Maize.

Drought is the major abiotic stress threatening maize (Zea mays L.) production globally. Despite recent scientific headway in deciphering maize drought stress responses, the overall picture of key genes, pathways, and co-expression networks regulating maize drought tolerance is still fragmented. Therefore, deciphering the molecular basis of maize drought tolerance remains pertinent. Here, through a comprehensive comparative leaf transcriptome analysis of drought-tolerant hybrid ND476 plants subjected to water-sufficient and water-deficit treatment conditions at flared (V12), tasseling (VT), the prophase of grain filling (R2), and the anaphase of grain filling (R4) crop growth stages, we report growth-stage-specific molecular mechanisms regulating maize drought stress responses. Based on the transcriptome analysis, a total of 3,451 differentially expressed genes (DEGs) were identified from the four experimental comparisons, with 2,403, 650, 397, and 313 DEGs observed at the V12, VT, R1, and R4 stages, respectively. Subsequently, 3,451 DEGs were divided into 12 modules by weighted gene co-expression network analysis (WGCNA), comprising 277 hub genes. Interestingly, the co-expressed genes that clustered into similar modules exhibited diverse expression tendencies and got annotated to different GO terms at different stages. MapMan analysis revealed that DEGs related to stress signal transduction, detoxification, transcription factor regulation, hormone signaling, and secondary metabolites biosynthesis were universal across the four growth stages. However, DEGs associated with photosynthesis and amino acid metabolism; protein degradation; transport; and RNA transcriptional regulation were uniquely enriched at the V12, VT, R2, and R4 stages, respectively. Our results affirmed that maize drought stress adaptation is a growth-stage-specific response process, and aid in clarifying the fundamental growth-stage-specific mechanisms regulating drought stress responses in maize. Moreover, genes and metabolic pathways identified here can serve as valuable genetic resources or selection targets for further functional validation experiments.

Metabolomic analysis to detect urinary molecular changes associated with bipolar depression.

Bipolar disorder (BD) is a debilitating mental disorder with complex clinical manifestations and low diagnostic accuracy. Depressive episodes are most common in the course of BD with high comorbidity and suicide rates, which present greater clinical challenges than mania and hypomania episodes. However, there are no objective biomarkers for bipolar depression. The aim of this study was to detect urinary metabolite biomarkers that could be useful for the diagnosis of bipolar depression. Nuclear magnetic resonance spectroscopy was used to profile urine samples of patients with bipolar depression (n = 37) and healthy volunteers (n = 48). Data were analyzed using Orthogonal Partial Least Square Discriminant Analysis and t-test. Differential metabolites were identified (VIP > 1 and p < 0.05), and further analyzed using Metabo Analyst 3.0 to identify associated metabolic pathways. In total, we identified seven metabolites differentially expressed in patients with BD and healthy controls. Compared with healthy group, the levels of betaine, glycerol, hippuric acid, indole sulfate, trimethylamine oxide, and urea in urine samples of BD patients were significantly higher, while the level of inositol was significantly lower. Most of these small molecules are related to lipid metabolism and gut microbiota metabolism. These differential metabolites could provide critical insight into the pathological mechanisms of bipolar depression. The results of this study provide a meaningful reference for similar and further studies in the future.

Cdk5-Mediated Phosphorylation of Sirt1 Contributes to Podocyte Mitochondrial Dysfunction in Diabetic Nephropathy.

Aims: Mitochondrial dysfunction contributes to podocyte injury, which is the leading cause of proteinuria in diabetic nephropathy (DN). In this study, we explored the role of cyclin-dependent kinase 5 (Cdk5) in mitochondrial dysfunction of podocytes under diabetic conditions. Results: Our results showed that the expression and activity of Cdk5 were significantly upregulated in vivo and in vitro under diabetic conditions, accompanied by the downregulation of synaptopodin and nephrin, as well as structural and functional mitochondrial dysfunction. Inhibition of Cdk5 with roscovitine or dominant-negative Cdk5 led to the attenuation of podocyte injury by upregulating synaptopodin and nephrin. The inhibition of Cdk5 also ameliorated mitochondrial dysfunction by decreasing reactive oxygen species levels and cytochrome c release, while increasing adenosine triphosphate production. Sirt1, an NAD+-dependent deacetylase, was decreased in podocytes with high glucose (HG) treatment; however, its phosphorylation level at S47 was significantly upregulated. We demonstrated that HG levels cause overactive Cdk5 to phosphorylate Sirt1 at S47. Suppression of Cdk5 reduced Sirt1 phosphorylation levels and mutation of S47 to nonphosphorable alanine (S47A), significantly attenuated podocyte injury and mitochondrial dysfunction in diabetic condition in vivo and in vitro. Innovation and Conclusion: Our study has demonstrated the role of Cdk5 in regulating mitochondrial function through Sirt1 phosphorylation and thus can potentially be a new therapeutic target for DN treatment. IRB number: 20190040. Antioxid. Redox Signal. 34, 171-190.

Comparative transcriptomic analysis of contrasting hybrid cultivars reveal key drought-responsive genes and metabolic pathways regulating drought stress tolerance in maize at various stages.

Drought stress is the primary environmental factor that negatively influences plant growth and yield in cereal grain crops such as maize (Zea mays L.). Crop breeding efforts for enhanced drought resistance require improved knowledge of plant drought stress responses. In this study, we applied a 12-day water-deficit stress treatment to maize plants of two contrasting (drought tolerant ND476 and drought sensitive ZX978) hybrid cultivars at four (V12, VT, R1, and R4) crop growth stages and we report key cultivar-specific and growth-stage-specific molecular mechanisms regulating drought stress responses in maize. Based on the transcriptome analysis, a total of 3451 and 4088 differentially expressed genes (DEGs) were identified in ND476 and ZX978 from the four experimental comparisons, respectively. These gene expression changes effected corresponding metabolic pathway responses related to drought tolerance in maize. In ND476, the DEGs associated with the ribosome, starch and sucrose metabolism, phenylpropanoid biosynthesis and phenylpropanoid metabolism pathways were predominant at the V12, VT, R2, and R4 stages, respectively, whereas those in ZX978 were related to ribosome, pentose and glucuronate interconversions (PGI), MAPK signaling and sulfur metabolism pathways, respectively. MapMan analysis revealed that DEGs related to secondary metabolism, lipid metabolism, and amino acid metabolism were universal across the four growth stages in ND476. Meanwhile, the DEGs involved in cell wall, photosynthesis and amino acid metabolism were universal across the four growth stages in ZX978. However, K-means analysis clustered those DEGs into clear and distinct expression profiles in ND476 and ZX978 at each stage. Several functional and regulatory genes were identified in the special clusters related to drought defense response. Our results affirmed that maize drought stress adaptation is a cultivar-specific response as well as a stage-specific response process. Additionally, our findings enrich the maize genetic resources and enhance our further understanding of the molecular mechanisms regulating drought stress tolerance in maize. Further, the DEGs screened in this study may provide a foundational basis for our future targeted cloning studies.

Comparative transcriptomic and physiological analyses of contrasting hybrid cultivars ND476 and ZX978 identify important differentially expressed genes and pathways regulating drought stress tolerance in maize.

BACKGROUND: Drought is the major abiotic stress factor that negatively influences growth and yield in cereal grain crops such as maize (Zea mays L.). A multitude of genes and pathways tightly modulate plant growth, development and responses to environmental stresses including drought. Therefore, crop breeding efforts for enhanced drought resistance require improved knowledge of plant drought responses. OBJECTIVE: Here, we sought to elucidate the molecular and physiological mechanisms underpinning maize drought stress tolerance. METHODS: We therefore applied a 12-day water-deficit stress treatment to maize plants of two contrasting (drought tolerant ND476 and drought sensitive ZX978) hybrid cultivars at the late vegetative (V12) growth stage and performed a large-scale RNA sequencing (RNA-seq) transcriptome analysis of the leaf tissues. RESULTS: A comparative analysis of the two genotypes leaf transcriptomes and physiological parameters revealed the key differentially expressed genes (DEGs) and metabolic pathways that respond to drought in a genotype-specific manner. A total of 3114 DEGs were identified, with 21 DEGs being specifically expressed in tolerant genotype ND476 in response to drought stress. Of these, genes involved in secondary metabolites biosynthesis, transcription factor regulation, detoxification and stress defense were highly expressed in ND476. Physiological analysis results substantiated our RNA-seq data, with ND476 exhibiting better cell water retention, higher soluble protein content and guaiacol peroxidase activity, along with low lipid peroxidation extent than the sensitive cultivar ZX978 under drought conditions. CONCLUSION: Our findings enrich the maize genetic resources and enhance our further understanding of the molecular mechanisms regulating drought stress tolerance in maize. Additionally, the DEGs screened in this study may provide a foundational basis for our future targeted cloning studies.