Graphene enhances artemisinin production in the traditional medicinal plant Artemisia annua via dynamic physiological processes and miRNA regulation.

We investigated the effects of graphene on the model herb Artemisia annua, which is renowned for producing artemisinin, a widely used pharmacological compound. Seedling growth and biomass were promoted when A. annua was cultivated with low concentrations of graphene, an effect which was attributed to a 1.4-fold increase in nitrogen uptake, a 15%-22% increase in chlorophyll fluorescence, and greater abundance of carbon cycling-related bacteria. Exposure to 10 or 20 mg/L graphene resulted in a âˆ¼60% increase in H2O2, and graphene could act as a catalyst accelerator, leading to a 9-fold increase in catalase (CAT) activity in vitro and thereby maintaining reactive oxygen species (ROS) homeostasis. Importantly, graphene exposure led to an 80% increase in the density of glandular secreting trichomes (GSTs), in which artemisinin is biosynthesized and stored. This contributed to a 5% increase in artemisinin content in mature leaves. Interestingly, expression of miR828 was reduced by both graphene and H2O2 treatments, resulting in induction of its target gene AaMYB17, a positive regulator of GST initiation. Subsequent molecular and genetic assays showed that graphene-induced H2O2 inhibits micro-RNA (miRNA) biogenesis through Dicers and regulates the miR828-AaMYB17 module, thus affecting GST density. Our results suggest that graphene may contribute to yield improvement in A. annua via dynamic physiological processes together with miRNA regulation, and it may thus represent a new cultivation strategy for increasing yield capacity through nanobiotechnology.

Comparative Genomics and Functional Studies of Putative m6A Methyltransferase (METTL) Genes in Cotton.

N6-methyladenosine (m6A) RNA modification plays important regulatory roles in plant development and adapting to the environment, which requires methyltransferases to achieve the methylation process. However, there has been no research regarding m6A RNA methyltransferases in cotton. Here, a systematic analysis of the m6A methyltransferase (METTL) gene family was performed on twelve cotton species, resulting in six METTLs identified in five allotetraploid cottons, respectively, and three to four METTLs in the seven diploid species. Phylogenetic analysis of protein-coding sequences revealed that METTL genes from cottons, Arabidopsis thaliana, and Homo sapiens could be classified into three clades (METTL3, METTL14, and METTL-like clades). Cis-element analysis predicated the possible functions of METTL genes in G. hirsutum. RNA-seq data revealed that GhMETTL14 (GH_A07G0817/GH_D07G0819) and GhMETTL3 (GH_A12G2586/GH_D12G2605) had high expressions in root, stem, leaf, torus, petal, stamen, pistil, and calycle tissues. GhMETTL14 also had the highest expression in 20 and 25 dpa fiber cells, implying a potential role at the cell wall thickening stage. Suppressing GhMETTL3 and GhMETTL14 by VIGS caused growth arrest and even death in G. hirsutum, along with decreased m6A abundance from the leaf tissues of VIGS plants. Overexpression of GhMETTL3 and GhMETTL14 produced distinct differentially expressed genes (DEGs) in A. thaliana, indicating their possible divergent functions after gene duplication. Overall, GhMETTLs play indispensable but divergent roles during the growth of cotton plants, which provides the basis for the systematic investigation of m6A in subsequent studies to improve the agronomic traits in cotton.

Comprehensive identification and expression analysis of CRY gene family in Gossypium.

BACKGROUND: The cryptochromes (CRY) are specific blue light receptors of plants and animals, which play crucial roles in physiological processes of plant growth, development, and stress tolerance. RESULTS: In the present work, a systematic analysis of the CRY gene family was performed on twelve cotton species, resulting in 18, 17, 17, 17, and 17 CRYs identified in five alloteraploid cottons (Gossypium hirsutum, G. barbadense, G. tomentosum, G. mustelinum and G. darwinii), respectively, and five to nine CRY genes in the seven diploid species. Phylogenetic analysis of protein-coding sequences revealed that CRY genes from cottons and Arabidopsis thaliana could be classified into seven clades. Synteny analysis suggested that the homoeolog of G. hirsutum Gh_A02G0384 has undergone an evolutionary loss event in the other four allotetraploid cotton species. Cis-element analysis predicated the possible functions of CRY genes in G. hirsutum. RNA-seq data revealed that Gh_D09G2225, Gh_A09G2012 and Gh_A11G1040 had high expressions in fiber cells of different developmental states. In addition, the expression levels of one (Gh_A03G0120), 15 and nine GhCRY genes were down-regulated following the PEG, NaCl and high-temperature treatments, respectively. For the low-temperature treatment, five GhCRY genes were induced, and five were repressed. These results indicated that most GhCRY genes negatively regulate the abiotic stress treatments. CONCLUSION: We report the structures, domains, divergence, synteny, and cis-elements analyses systematically of G. hirsutum CRY genes. Possible biological functions of GhCRY genes in differential tissues as well as in response to abiotic stress during the cotton plant life cycle were predicted.

Characterization of cotton ARF factors and the role of GhARF2b in fiber development.

BACKGROUND: Cotton fiber is a model system for studying plant cell development. At present, the functions of many transcription factors in cotton fiber development have been elucidated, however, the roles of auxin response factor (ARF) genes in cotton fiber development need be further explored. RESULTS: Here, we identify auxin response factor (ARF) genes in three cotton species: the tetraploid upland cotton G. hirsutum, which has 73 ARF genes, and its putative extent parental diploids G. arboreum and G. raimondii, which have 36 and 35 ARFs, respectively. Ka and Ks analyses revealed that in G. hirsutum ARF genes have undergone asymmetric evolution in the two subgenomes. The cotton ARFs can be classified into four phylogenetic clades and are actively expressed in young tissues. We demonstrate that GhARF2b, a homolog of the Arabidopsis AtARF2, was preferentially expressed in developing ovules and fibers. Overexpression of GhARF2b by a fiber specific promoter inhibited fiber cell elongation but promoted initiation and, conversely, its downregulation by RNAi resulted in fewer but longer fiber. We show that GhARF2b directly interacts with GhHOX3 and represses the transcriptional activity of GhHOX3 on target genes. CONCLUSION: Our results uncover an important role of the ARF factor in modulating cotton fiber development at the early stage.

Genome-wide characterization of the GRF family and their roles in response to salt stress in Gossypium.

BACKGROUND: Cotton (Gossypium spp.) is the most important world-wide fiber crop but salt stress limits cotton production in coastal and other areas. Growth regulation factors (GRFs) play regulatory roles in response to salt stress, but their roles have not been studied in cotton under salt stress. RESULTS: We identified 19 GRF genes in G. raimondii, 18 in G. arboreum, 34 in G. hirsutum and 45 in G. barbadense, respectively. These GRF genes were phylogenetically analyzed leading to the recognition of seven GRF clades. GRF genes from diploid cottons (G. raimondii and G. arboreum) were largely retained in allopolyploid cotton, with subsequent gene expansion in G. barbadense relative to G. hirsutum. Most G. hirsutum GRF (GhGRF) genes are preferentially expressed in young and growing tissues. To explore their possible role in salt stress, we used qRT-PCR to study expression responses to NaCl treatment, showing that five GhGRF genes were down-regulated in leaves. RNA-seq experiments showed that seven GhGRF genes exhibited decreased expression in leaves under NaCl treatment, three of which (GhGRF3, GhGRF4, and GhGRF16) were identified by both RNA-seq and qRT-PCR. We also identified six and three GRF genes that exhibit decreased expression under salt stress in G. arboreum and G. barbadense, respectively. Consistent with its lack of leaf withering or yellowing under the salt treatment conditions, G. arboreum had better salt tolerance than G. hirsutum and G. barbadense. Our results suggest that GRF genes are involved in salt stress responses in Gossypium. CONCLUSION: In summary, we identified candidate GRF genes that were involved in salt stress responses in cotton.

The miR319-Targeted GhTCP4 Promotes the Transition from Cell Elongation to Wall Thickening in Cotton Fiber.

Plant cell growth involves a complex interplay among cell-wall expansion, biosynthesis, and, in specific tissues, secondary cell wall (SCW) deposition, yet the coordination of these processes remains elusive. Cotton fiber cells are developmentally synchronous, highly elongated, and contain nearly pure cellulose when mature. Here, we report that the transcription factor GhTCP4 plays an important role in balancing cotton fiber cell elongation and wall synthesis. During fiber development the expression of miR319 declines while GhTCP4 transcript levels increase, with high levels of the latter promoting SCW deposition. GhTCP4 interacts with a homeobox-containing factor, GhHOX3, and repressing its transcriptional activity. GhTCP4 and GhHOX3 function antagonistically to regulate cell elongation, thereby establishing temporal control of fiber cell transition to the SCW stage. We found that overexpression of GhTCP4A upregulated and accelerated activation of the SCW biosynthetic pathway in fiber cells, as revealed by transcriptome and promoter activity analyses, resulting in shorter fibers with varied lengths and thicker walls. In contrast, GhTCP4 downregulation led to slightly longer fibers and thinner cell walls. The GhHOX3-GhTCP4 complex may represent a general mechanism of cellular development in plants since both are conserved factors in many species, thus providing us a potential molecular tool for the design of fiber traits.

Dexamethasone decreases cholestatic liver injury via inhibition of intrinsic pathway with simultaneous enhancement of mitochondrial biogenesis.

BACKGROUND: Mitochondria are known to be involved in cholestatic liver injury. We tested the hypothesis that glucocorticoids can modulate mitochondrial function to alleviate cholestatic liver injury. METHODS: A rat model of cholestasis was established by bile duct ligation (BDL), with a sham group receiving laparotomy without BDL, and a group receiving dexamethasone (DEX) treatment after BDL. RESULTS: The liver function including total bilirubin levels, alanine transaminase and aspartate transaminase activities was significantly improved in the DEX treatment group in comparison to the BDL group. There was a significant upregulation of liver peroxisome proliferator-activated receptor γ coactivator-1α and mitochondrial transcriptional factor A protein between 6 and 72 h was found in the DEX group. DEX treatment significantly down-regulated Bax, caspase 9 and caspase 3 expression induced by BDL at 24-72 h, but had little effect on the expression of caspase 8, Bcl(2,) Fas and Fas-FasL complex. Consequently, the number of apoptotic liver cells in the DEX group was significantly less than in the BDL group at 72 h. CONCLUSION: Our results indicate that glucocorticoids decreases cholestatic liver injury within hours after BDL. Early glucocorticoids treatment can enhance the mitochondrial biogenesis and modulate the intrinsic but not extrinsic pathway of apoptosis following BDL.

Cavernous malformations of the central nervous system combined with cutaneous vascular lesions due to KRIT1 mutation: a case report.

Cavernous malformations (CMs) of the central nervous system can occur in a sporadic condition or as a familial form with an autosomal-dominant inherited pattern. Apart from a family history, some clinical features may help to identify familial CMs. We demonstrate clinical, neuroradiological, pathological, and genetic data of a patient with cerebral and spinal CMs. The presence of multiple cerebral CMs and distinct cutaneous vascular lesions, including hyperkeratotic cutaneous capillary-venous malformations, in this patient suggested familial CMs. A genetic study confirmed a nonsense mutation (c.1708A>T) in the KRIT1 gene.

Clinicopathological characteristics and prognosis of patients with small intestinal stromal tumors.

BACKGROUND AND PURPOSE: Gastrointestinal stromal tumors (GISTs) involving the small intestine are less common than those involving the stomach, and data on small intestinal stromal tumors (SISTs) are more limited. This study investigated the clinicopathological characteristics and prognostic factors of SISTs and compared them with those of gastric stromal tumors (GSTs). METHODS: A total of 82 surgically resected and pathologically diagnosed smooth muscle tumors of small bowel in patients treated from January 1986 to December 2000 were included. Immunohistochemical studies were performed on these tumors with antibodies of CD117, CD34, smooth muscle actin (SMA), desmin and S-100. The results were analyzed and compared with 74 cases of GSTs diagnosed and treated from January 1986 to December 1997. RESULTS: Among the 82 small intestine tumors, 71 were CD117-positive (86.6%) and were classified as SISTs. Of the 71 SISTs, 70.4% were immunoreactive to CD34, 88.7% to SMA, 46.5% to S-100, but none to desmin. Survival analysis demonstrated that tumor size < 5 cm (p = 0.021), mitosis number < 5/50 high-power field (p < 0.001), SMA-positive (p = 0.027), non-epithelioid cell type (p = 0.005) and tumor with skeinoid fibers (p = 0.010) predicted longer disease-free survival after operation. Multivariate analysis revealed that mitotic number (p = 0.001), cell morphology (p = 0.031) and tumor size (p = 0.004) were independent prognostic factors. In comparison to GSTs, SISTs exhibited significantly lower rates of CD34, but significantly higher rates of SMA and S-100 immunoreactivity. CONCLUSIONS: SISTs exhibited a different immunophenotype from GSTs. SMA reactivity is a predictor of benign clinical behavior in SISTs. Tumor mitotic numbers, tumor size, and cell type were independent prognostic factors for patients with SISTs after operation.

Expression, by functional proteomics, of spontaneous tolerance in rat orthotopic liver transplantation.

Orthotopic liver transplants (OLT) performed in certain combinations of donor and recipient rat strains, such as DA (RT1a) to PVG (RT1c), without immunosuppressive drugs could completely overcome major histocompatibility complex barriers. Although other organs transplanted in a similar fashion within the same combination have been promptly rejected, 60 day post-OLT serum (POD 60) has been proven competent in rapidly reversing the established rejection in animal models. In order to understand the functional role of tolerogenic serum proteins and their involvement with immune response regulation, a comprehensive analysis surveying global changes in complex OLT systems by proteomic techniques was applied. The results display the varying protein expressions in sera extracted from naïve and transplanted animals on POD 60 with regard to immunosuppression. Among these proteins, haptoglobin (Hp) which is related to inhibition of T-cell proliferation was found to be up-regulated following OLT. In addition, the transcriptional expression level and intracellular localization of Hp correlated with the immune events. Hp also exhibited a strong in vitro immunosuppressive effect on the mixed lymphocyte reaction. In conclusion, the presence of Hp may play an important role in modulating the spontaneous tolerance of liver transplantation. Furthermore, the serum proteome map could provide guidance with respect to discovering potential protein targets in OLT tolerance and eventually prolong hepatic allograft survival in the future.