N6-甲基化腺嘌呤RNA甲基化修饰在中枢神经系统中的作用研究进展

mRNA存在多种转录后修饰,这些修饰调控mRNA的稳定和剪接、翻译、转运等多个过程,进而影响细胞发育、机体免疫、学习认知等重要生理功能。其中m⁶A修饰是转录后修饰中最丰富的一种,广泛存在于mRNA中,调控mRNA的代谢活动,影响基因表达。m⁶A修饰的稳态对神经系统的发育和功能维持至关重要。近年研究发现,在神经退行性疾病、精神疾病和脑肿瘤中均存在m⁶A修饰的身影。因此本文对近几年m⁶A甲基化修饰在中枢神经系统发育、功能及相关疾病中的作用进行总结,为神经系统疾病提供潜在的临床治疗靶点。     

Two calcium mobilizing pathways implicated within abscisic acid-induced stomatal closing in <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

The present study investigated whether, depending on the abscisic acid (ABA) concentration, phospholipase C (PLC) would be implicated within a Ca²⁺ mobilizing pathway that would regulate stomatal aperture under standard watering conditions. Among Al sensitive mutants the als1-1 mutant of Arabidopsis thaliana (L.) Heynh. (Columbia-4 ecotype) was selected for a pharmacological approach of stomatal closing in leaf epidermal peels induced by 3, 20 or 30 μM ABA. Comparison with the wild type (WT) revealed that, exclusively in the als1-1 mutant, the stomatal response to 3 or 20 μM ABA was inhibited by about 40 %, whereas the stomatal response to 30 μM ABA and the wilting response to drought were unaffected. In WT, the Ca²⁺ buffer EGTA and the PLC inhibitor, 1-[6-[[17β-3-methoxyestra-1,3,5(10)-trien-17-yl]amino]hexyl]-1H-pyrrole-2,5-dione (U73122), specifically inhibited by about 70 and 40 %, respectively, the response to 3 or 20 μM ABA, while the Ca²⁺ buffer 1,2-bis(o-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid (BAPTA) inhibited by about 70 % the response to 3, 20 or 30 μM ABA. EGTA, BAPTA and U73122 did not inhibit the part of the response to 3 or 20 μM ABA that was unaffected by the als1-1 mutation. Together, these results showed that ABA closes the stomata through two different Ca²⁺ mobilizing pathways. Since PLC could be indirectly deactivated in the als1-1 mutant, these results might suggest that, under sufficient water supply, PLC-mediated Ca²⁺ mobilization is needed for the regulation of stomatal aperture by endogenous ABA resting at concentrations below a drought-specific threshold value.     

Calcium is involved in the abscisic acid-induced ascorbate peroxidase, superoxide dismutase and chilling resistance in Stylosanthes guianensis

The objective of this work was to test whether Ca²⁺, a second messenger in stress response, is involved in ABA-induced antioxidant enzyme activities in Stylosanthes guianensis. Plants were sprayed with abscisic acid (ABA), calcium channel blocker, LaCl₃, calcium chelator, ethylene glycol-bis(β-amino ethyl ether)-N,N,N′, N′-tetraacetid acid (EGTA), and ABA in combination with LaCl₃ or EGTA. Their effects on superoxide dismutase (SOD) and ascorbate peroxidase (APX) activities and chilling resistance were compared. The results showed that ABA decreased electrolyte leakage and lipid peroxidation but increased maximum photochemical efficiency measured as variable to maximum fluorescence ratio (F_v/F_m) under chilling stress. Treatment with LaCl₃ or EGTA alone and in combination with ABA increased electrolyte leakage and lipid peroxidation, decreased Fv/Fm, suggesting that the block in Ca²⁺ signalling decreased chilling resistance of S. guianensis and the ABA-enhanced chilling resistance. ABA-induced SOD and APX activities were suppressed by LaCl₃ or EGTA. The results suggested that Ca²⁺ is involved in the ABA-enhanced chilling resistance and the ABA-induced SOD and APX activities in S. guianensis.     

Osmotic stress and abscisic acid reduce cytosolic calcium activities in roots of Arabidopsis thaliana*

Cytosolic Ca²⁺· ([Ca²⁺]_i, and elongation growth were measured in the roots of Arabidopsis thaliana. Exposure of plant tissues to high NaCl and abscisic acid (ABA) concentrations results in a reduction in the rate of growth, but the mechanism by which growth is inhibited is not understood. Both NaCl and ABA treatments are known to influence [Ca²⁺]_i, and in this study we measured the effects of salinity and ABA on [Ca²⁺]_i in cells from the meristematic region of Arabidopsis roots. The Ca²⁺-sensitive dye Fura-2 and ratiometric techniques were used to measure [Ca²⁺]_i in cells of the root meristem region. Resting [Ca²⁺]_i was found to be between 100 and 200 μmol m^?3 in roots of untreated plants. Resting [Ca²⁺]_i changed in response to changes in the [Ca²⁺] surrounding growing roots. An increase of external [Ca²⁺] increased [Ca²⁺]_i; conversely, a decrease of external [Ca²⁺] decreased [Ca²⁺]_i. Exposure of roots to NaCl caused a rapid reduction of [Ca²⁺]_i, a response that was proportional to the external NaCl concentration. Thus, as the NaCl concentration was increased, [Ca²⁺]_i in root meristematic cells decreased. Root elongation was also inhibited in proportion to the external NaCl concentration, with maximal inhibition occurring at 120 mol m^?3 NaCl. The [Ca²⁺]_i of root meristem cells also changed in response to ABA, and the magnitude of the effect of ABA was dependent upon ABA concentration. Treatment with 0.2 mmol m^?3 ABA caused a momentary increase in [Ca²⁺]_i followed by a decrease after 15 min, but 10 mmol m^?3 ABA caused an immediate decline in [Ca²⁺]_i. There was a strong positive correlation between [Ca²⁺]_i and root elongation rates. Experiments with the ABA-deficient Arabidopsis mutant aba-3 indicated that the reduction in [Ca²⁺]_i brought about by NaCl was unlikely to be mediated via changes in endogenous ABA. Experiments with solutes such as sorbitol, KCl and NaNO₃ indicated that the effects of NaCl could be mimicked by other solutes and was not specific for NaCl.     

脂肪含量和肥胖相关蛋白介导的mRNA m6A修饰对动物脂肪沉积的作用及其应用前景

在动物脂肪沉积过程中,前体脂肪细胞增殖、分化和脂滴甘油三酯水平的变化受到一系列转录因子和信号通路的调节。目前研究者虽对脂肪形成的转录调控机制进行了深入研究,但对转录后mRNA水平修饰的报道相对较少。甲基化转移酶、去甲基化酶和甲基化阅读蛋白共同调控的mRNA m⁶A修饰是动态可逆的且与脂肪沉积密切相关。脂肪含量和肥胖相关蛋白（fat mass and obesityassociated,FTO）作为RNA去甲基化酶,影响被修饰基因的表达,在脂肪沉积中起关键作用。文中系统分析并总结了FTO介导的mRNA m⁶A去甲基化对动物脂肪沉积的作用及分子调控机制的研究进展,提示FTO可能成为有效治疗肥胖症的靶点;还对近年来研发FTO抑制剂的情况进行了总结,并展望其在治疗肥胖症方面的研究前景。     

Genome-Wide Functional Analysis of Cotton (Gossypium hirsutum) in Response to Drought

Cotton is one of the most important crops for its natural textile fibers in the world. However, it often suffered from drought stress during its growth and development, resulting in a drastic reduction in cotton productivity. Therefore, study on molecular mechanism of cotton drought-tolerance is very important for increasing cotton production. To investigate molecular mechanism of cotton drought-resistance, we employed RNA-Seq technology to identify differentially expressed genes in the leaves of two different cultivars (drought-resistant cultivar J-13 and drought-sensitive cultivar Lu-6) of cotton. The results indicated that there are about 13.38% to 18.75% of all the unigenes differentially expressed in drought-resistant sample and drought-sensitive control, and the number of differentially expressed genes was increased along with prolonged drought treatment. DEG (differentially expression gene) analysis showed that the normal biophysical profiles of cotton (cultivar J-13) were affected by drought stress, and some cellular metabolic processes (including photosynthesis) were inhibited in cotton under drought conditions. Furthermore, the experimental data revealed that there were significant differences in expression levels of the genes related to abscisic acid signaling, ethylene signaling and jasmonic acid signaling pathways between drought-resistant cultivar J-13 and drought-sensitive cultivar Lu-6, implying that these signaling pathways may participate in cotton response and tolerance to drought stress.     

Abscisic acid concentrations and fluxes in droughted conifer saplings

We present the first study of abscisic acid (ABA) concentrations and fluxes in the xylem sap of conifers during a drought cycle. In both Pinus sylvestris and Picea sitchensis the concentration of ABA in the sap rose 11-fold as the drought progressed. There were clear diurnal trends in this concentration, which reached its maximum (6–8.ininol ABA m^?3) near the middle of the day. The fluxes of ABA were calculated by multiplying the xylem ABA concentration by the sap flow rate. The ABA fluxes in the droughted plants in the middle of the day were usually no higher than those of the controls, as a result of the very low sap flow in the droughted plants at that time. However, the ABA flux in the droughted plants was higher than in the controls in the morning, and we postulate that the stomata are responding to these ‘morning doses’ Stomatal conductance, g_s, could not be related statistically to leaf turgor or to the ABA flux. However, £s did display a negative exponential relationship with ABA concentration in the xylem. Pinus displayed more acclimation to drought than Picea, Its ABA concentration rose and its stomatal conductance fell at day 6 of the drought, as opposed to day 17 for Picea, and its osmotic potential fell during the drought treatment.     

Screening of abiotic stress‐responsive cotton genes using a cotton full‐length cDNA overexpressing Arabidopsis library

Cotton (Gossypium hirsutum L.) is a major crop and the main source of natural fiber worldwide. Because various abiotic and biotic stresses strongly influence cotton fiber yield and quality, improved stress resistance of this crop plant is urgently needed. In this study, we used Gateway technology to construct a normalized full‐length cDNA overexpressing (FOX) library from upland cotton cultivar ZM12 under various stress conditions. The library was transformed into Arabidopsis to produce a cotton‐FOX‐Arabidopsis library. Screening of this library yielded 6,830 transgenic Arabidopsis lines, of which 757 were selected for sequencing to ultimately obtain 659 cotton ESTs. GO and KEGG analyses mapped most of the cotton ESTs to plant biological process, cellular component, and molecular function categories. Next, 156 potential stress‐responsive cotton genes were identified from the cotton‐FOX‐Arabidopsis library under drought, salt, ABA, and other stress conditions. Four stress‐related genes identified from the library, designated as GhCAS, GhAPX, GhSDH, and GhPOD, were cloned from cotton complementary DNA, and their expression patterns under stress were analyzed. Phenotypic experiments indicated that overexpression of these cotton genes in Arabidopsis affected the response to abiotic stress. The method developed in this study lays a foundation for high‐throughput cloning and rapid identification of cotton functional genes.     

The role of N6‐methyladenosine modification on diapause in silkworm (Bombyx mori) strains that exhibit different voltinism

N6‐methyladenosine (m⁶A) plays a key role in regulating gene expression in myriad organisms. Diapause is an important plastic phenotype that allows insects to survive under specific environmental conditions. However, the diapause molecular mechanism remains unknown. In this study, we analyzed the phylogenetics of genes related to the m⁶A modification complex in the silkworm (Bombyx mori) based on identified sequences from other organisms. We detected the expression of these genes during different developmental phases from four strains with different voltinism. We also determined total m⁶A content in cells treated with different diapause hormone concentrations or eggs exposed to hydrochloric acid. Our data revealed that m⁶A‐modification‐related gene expression and m⁶A content were greater in diapause‐destinated compared to nondiapause‐destined strains. Our findings suggest that m⁶A modification may provide significant epigenetic regulation of diapause‐related genes in the silkworm.     

西葫芦NCED基因家族鉴定及其响应干旱胁迫分析

NCED基因家族成员在调节植物响应干旱胁迫中发挥着关键作用,该研究通过生物信息学技术分析NCED在西葫芦基因组中的分布、结构及进化,研究家族成员在不同组织中的表达特异性及其对10%PEG 6000模拟干旱、0.1 mmol·L-1ABA激素和自然干旱胁迫的响应,以解析NCED基因家族的生物学功能。结果表明:(1)从西葫芦全基因组中鉴定出6个NCED家族基因(CpNCED1~6),且6个基因均不含内含子、分别分布于西葫芦的1、10、12、14、19和20号共6条染色体上。(2)理化性质分析发现,CpNCED1~6蛋白长度为569~590 aa,理论分子量在62.64~65.54 kD之间。(3)蛋白保守元件分析显示,除CpNCED3蛋白在遗传进化过程中出现3个基序(motif 12、motif 13和motif 15)的缺失外,其余5个蛋白都有完整的16个motif保守基序,且分布在600个氨基酸以内,同时大部分NCED蛋白序列保守性较高。(4)顺式作用元件分析显示,西葫芦CpNCED1~6基因均含ABRE、W box、MBS、P-box、TCA-element、CGTCA-motif、TGA-element和TGA-box等潜在的干旱胁迫响应元件。(5)qRT-PCR分析表明,CpNCED1~6基因在西葫芦不同组织中的表达具有组织特异性,其中,CpNCED4和CpNCED1在茎中的表达量显著高于其他4个基因,CpNCED2、CpNCED4、CpNCED6在花中的表达显著高于其余3个基因且CpNCED2表达量最高,CpNCED1~6在果实和叶中的表达量均相对较低;与对照组相比,CpNCED1~6受模拟干旱、ABA激素和自然干旱胁迫均上调表达;伴随干旱胁迫的产生,叶片中脱落酸(ABA)含量逐渐升高,暗示CpNCEDs在西葫芦干旱胁迫响应与ABA的生物合成过程中发挥着正向调控作用。研究发现,6个CpNCED1~6基因与西葫芦干旱胁迫响应密切相关,且对西葫芦干旱胁迫的响应以及ABA生物合成具有重要作用,尤其以CpNCED2和CpNCED4基因的作用更为明显。