Extensive profiling of the expressions of tRNAs and tRNA-derived fragments (tRFs) reveals the complexities of tRNA and tRF populations in plants

Evidence is emerging that t RNA-derived fragments(t RFs) are regulatory molecules. Studies of t RFs in plants have been based on conventional small RNA sequencing, and focused on profiling of t RF-5 and t RF-3 species. A more comprehensive and quantitative analysis of the entire t RF population is highly necessary. Here, we employ t RNA-seq and YAMAT-seq, and develop a bioinformatics tool to comprehensively profile the expressions of t RNAs and t RFs in plants. We show that in Arabidopsis,approximately half of t RNA genes are extremely weakly expressed, accounting for only 1% of total t RNA abundance, while～12% of t RNA genes contribute to ～80% of t RNA abundance. Our t RNA sequencings in various plants reveal that t RNA expression profiles exhibit a cross-species conserved pattern. By characterizing the composition of a highly heterogeneous t RF population, we show that t RNA halves and previously unnoticed 10–16-nt tiny t RFs represent substantial portions. The highly accumulated 13-nt and 16-nt tiny t RFs in Arabidopsis indicate that tiny t RFs are not random t RNA degradation products. Finally,we provide a user-friendly database for displaying the dynamic spatiotemporal expressions of t RNAs and t RFs in the model plants Arabidopsis and rice.     

不稳定EGFP细胞模型的构建及其在基因编辑体系评价中的应用*

目的:为了更好地评价基因编辑效率,满足高通量筛选应用中快速、高效的检测要求,在细胞上建立一个原位检测方法具有重要的意义。通过检测荧光蛋白信号强度的变化可以评价CRISPR系统在细胞中的基因编辑情况,然而这一方法的效率受限于荧光蛋白较长的半衰期。方法:将鸟氨酸脱羧酶降解结构域(含PEST序列)与EGFP融合,通过慢病毒系统感染HEK-293T细胞,获得了表达单拷贝、EGFP-PEST报告基因的稳转细胞系。结果:与EGFP相比,EGFP-PEST在细胞内的降解速度明显加快,荧光水平在4 h内显著降低。利用该模型比较了3种商品化脂质体介导的CRISPR/Cas9基因编辑效率,能够在2~4 d实现定性和定量评价。结论:这一模型能够快速、灵敏地指示基因编辑效果,可以用于不同CRISPR系统或新递送工具的高通量筛选和评价。     

利用细菌人工染色体技术构建整合F基因的重组MDV疫苗株*

目的:预防马立克氏病病毒(MDV)和新城疫病毒(NDV)混合感染鸡引起的疾病,构建表达NDV F蛋白的MDV疫苗株CVI988 BAC重组载体,并包装成重组病毒,为疫苗免疫提供更多的重组疫苗选择。方法:首先利用PCR扩增带有卡那霉素(Kanamycin,Kana)抗性基因片段的F基因,采用同源重组的方法将其整合到CVI988 BAC上,进一步诱导I-SceI表达敲除Kana基因而获得重组质粒CVI988 BAC-F。通过磷酸钙法转染鸡胚成纤维细胞获得重组病毒。结果:Western blot和间接免疫荧光实验证实重组病毒能够表达F蛋白。病毒生长曲线和蚀斑大小测定结果表明,F基因的插入不影响病毒的体外增殖。结论:利用BAC技术成功构建了整合F基因的重组MDV病毒CVI988 BAC-F,为MDV重组疫苗研发,防控NDV与MDV共感染奠定了基础。     

中国亚热带锈菌补记

描述了无性型锈菌新种龙眼夏孢锈菌Uredo longan;对荔枝翼孢锈菌Skierka nephelii补述了冬孢子阶段并指定了附加模式;报道了中国新记录刚果盾锈菌Uredopeltis congensis;新组合了克莱门斯层锈菌Phakopsora clemensiae comb. nov.和马唐层锈菌Phakopsora digitariae comb. nov.。引证的标本采自我国台湾、广东、海南、四川等亚热带地区,均保存于中国科学院菌物标本馆（HMAS）。     

拮抗临床致病菌的一株地衣内生真菌

本研究运用组织分离法对采自贵州省都匀市斗篷山景区的地衣进行内生真菌分离,乙酸乙酯萃取菌株液体发酵物,通过打孔法筛选地衣内生真菌提取物并对9株临床致病菌株及耐药菌株进行抑菌活性实验。结果表明一株分离自星点梅衣属地衣Punctelia sp.的内生真菌DPS-165-9对其中的6株显示抑制活性。此6株菌为金黄色葡萄球菌Staphylococcus aureus 25923和6538、耐药表面葡萄球菌Staphylococcus epidermidis Z12、耐药布氏杆菌Brucella sp. B103、耐药枯草芽孢杆菌Bacillus subtilis 163和耐药藤黄微球菌Micrococcus luteus 261。经鉴定DPS-165-9为蔡氏轮层炭壳菌Daldinia childiae。本研究为下一步挖掘新的抗菌物质奠定基础。     

环磷酰胺辅助化疗对不同年龄乳腺癌患者卵巢功能的损伤情况及机制研究

目的:以月经变化情况及血清激素水平为观察指标,探讨环磷酰胺辅助化疗方案对绝经前不同年龄段乳腺癌患者卵巢功能的损伤情况及可能的机制研究。方法:收集2017年1月至2018年12月期间在我院接受环磷酰胺辅助化疗的绝经前乳腺癌患者77例。患者年龄范围为26-48岁,并按年龄分为两组:≤35岁组为16例;>35岁组为61例。调查研究两组患者化疗前后的月经情况,以及血清激素水平(E、FSH、AMH)。同时进行动物实验,采用单剂量环磷酰胺(200 mg/kg)处理不同月龄组(3月龄和6月龄)雌性C57BL/6J小鼠,每组小鼠12例。在环磷酰胺处理前及处理后一周,检测小鼠血清AMH水平,计数小鼠卵巢卵泡数量。结果:环磷酰胺化疗后,>35岁组与≤35岁组间比较,>35岁组闭经率更高,复经率更低,血清AMH和E2水平下降幅度更大(P<0.05),FSH水平升高幅度也大于≤35岁组,但差异无明显统计学意义(P>0.05)。不同年龄组内比较,≤35岁组患者化疗后的血清AMH和E2下降水平无明显差异(P>0.05);血清FSH水平明显升高(P<0.05)。而>35岁组的患者化疗后,血清AMH和E2水平明显下降,FSH水平明显升高,差异均有统计学意义(P<0.05)。不同月龄组小鼠环磷酰胺处理前后,血清AMH水平明显下降,卵巢原始卵泡和生长卵泡数明显较少,差异均有统计学意义(P<0.05),且6月龄组小鼠下降程度更明显(P<0.05)。结论:环磷酰胺辅助化疗方案明显损伤患者卵巢功能,且对年龄越大者损伤程度越严重,更易导致不可逆性卵巢损伤。     

线粒体蛋白质组及蛋白质量控制

线粒体是哺乳动物细胞内重要细胞器,不仅通过氧化磷酸化产生ATP为细胞提供能量,也参与调节钙离子稳态、活性氧(reactive oxygen species,ROS)的产生、细胞应激反应和细胞死亡等过程,其功能障碍不仅导致多种人类疾病的发生,而且也能降低动物卵母细胞质量和早期胚胎发育能力。大量证据表明,线粒体的功能依赖于线粒体蛋白质组完整性和稳态。基于此,该文综述了线粒体蛋白组、线粒体蛋白转运,聚焦蛋白酶、分子伴侣、线粒体囊泡、线粒体自噬和线粒体未折叠蛋白反应在帮助正确的蛋白质折叠,去除错误折叠或聚集的蛋白质和清除功能失调的线粒体方面的作用,为调控线粒体蛋白质量,从而维持线粒体健康、降低疾病发生提供理论依据。     

Comprehensive landscape and interference of clonal haematopoiesis mutations for liquid biopsy: A Chinese pan-cancer cohort

Tumour-derived DNA found in the plasma of cancer patients provides the probability to detect somatic mutations from circulating cell-free DNA (cfDNA) in plasma samples. However, clonal hematopoiesis (CH) mutations affect the accuracy of liquid biopsy for cancer diagnosis and treatment. Here, we integrated landscape of CH mutations in 11,725 pan-cancer patients of Chinese and explored effects of CH on liquid biopsies in real-world. We first identified 5933 CHs based on panel sequencing of matched DNA of white blood cell and cfDNA on 301 genes for 5100 patients, in which CH number of patients had positive correlation with their diagnosis age. We observed that canonical genes related to CH, including DNMT3A, TET2, ASXL1, TP53, ATM, CHEK2 and SF3B1, were dominant in the Chinese cohort and 13.29% of CH mutations only appeared in the Chinese cohort compared with the Western cohort. Analysis of CH gene distribution bias indicated that CH tended to appear in genes with functions of tyrosine kinase regulation, PI3K-Akt signalling and TP53 activity, suggesting unfavourable effects of CH mutations in cancer patients. We further confirmed effect of driver genes carried by CH on somatic mutations in liquid biopsy of cancer patients. Forty-eight actionable somatic mutations in 17 driver genes were considered CH genes in 92 patients (1.80%) of the Chinese cohort, implying potential impacts of CH on clinical decision-making. Taken together, this study exhibits strong evidence that gene mutations from CH interfere accuracy of liquid biopsies using cfDNA in cancer diagnosis and treatment in real-world.