The use of ejaculated boar semen after freezing in 2 or 6% glucerol for in vitro fertilization of porcine oocytes matured in vitro

Two concentrations of glycerol in a freezing diluent were tested with respect to the in vitro fertilizing capacity of frozen-thawed boar spermatozoa which, before exposure to oocytes, were subjected to 3 methods of fractionation. These were 1) the upper fraction, 2) the swim-up and 3) percoll gradinet-centrifugation. The highest proportions of motile spermatozoa were obtained by the swim-up procedure, while acrosomal integrity was best preserved by the upper fraction procedure. Raising the glycerol concentration from 2 to 6% (v/v) during freezing decreased the proportion of spermatozoa with a normal apical ridge. Spermatozoa separated by the upper fraction method showed the greatest penetration of oocytes and produced the highest indidence of polyspermy. The glycerol level affected penetration and polyspermy only with spermatozoa separated in a percoll gradient, where the higher level of glycerol increased oocytes penetration and polyspermy. Pronuclei formation was influenced by the separation procedure and by the glycerol concentration in the freezing diluent. The results indicate that frozen boar semen can be used for in vitro fertilization more successfully than fresh semen since penetration by frozen upper fraction spermatozoa was similar to, the degree of polyspermy was lower, and the formation of two pronuclei was greater (P<0.01) than in oocytes exposed to fresh semen.     

The determination of the pKa of histidine residues in proteins by Raman difference spectroscopy

Sensitive Raman difference spectroscopy was used to monitor the protonation and deprotonation of histidine residues in apo-transferrin. We have shown previously that the behavior of small molecules and/or small molecular groups bound to proteins or other large macromolecules can be studied by Raman difference spectroscopy (Yue, K.T. et al. (1989) J. Raman Spectrosc. 20, 541-545). Using this method, we have measured the Raman difference spectra of human transferrin at different pH values with respect to pH 8.9, titrating its various histidine residues. About 12 +/- 2 of the 19 residues were titrated. The pH difference spectrum of transferrin obtained is very similar to that of histidine in solution, but with clear differences in the 1200-1400 cm-1 region. A titration curve with pKa of 6.08 +/- 0.01 fit the data of histidine in solution and a value of 6.56 +/- 0.02 was found for the average value of the 12 histidine residues inside transferrin. The technique has enough sensitivity at present to monitor a single histidine residue in a 130 kDa molecule and to determine the titration curve of one residue in a 40 kDa protein.     

Functional analysis of Arg-308 mutants of Flp recombinase. Possible role of Arg-308 in coupling substrate binding to catalysis

The arginine residue at position 308 in the Flp recombinase corresponds to the only invariant arginine within the Int family of recombinases. Alterations of this residue result in Flp variants that retain substrate recognition, but form weaker protein-DNA complexes than wild type Flp. Furthermore, their DNA cleavage activity is significantly diminished. A conservative change of R308K results in a functional Flp variant; however, this protein has a lowered temperature optimum for recombination. The Arg-308 mutants can be stabilized on the DNA substrate through cooperativity with a partner Flp mutant that is tight binding. Thus, interactions between Flp monomers must be a relevant feature of the normal recombination reaction.     

The seminiferous growth factor induces proliferation of TM4 cells in serum-free medium

The seminiferous growth factor (SGF) of the mammalian testes induces DNA synthesis and cell proliferation of Balb/c 3T3 cells (Bellvé and Feig, 1984; Rec Prog Hormone Res 40:531-567). In this study, SGF was purified 80,000- to 100,000-fold from calf testes and used to examine the growth of TM4 cells in a chemically defined medium. Cells were seeded sparsely in Dulbecco's Modified Eagles/Ham's F12 medium (1:1;v:v) (DME/F12 degrees), containing epidermal growth factor (EGF; 1 ng/ml), insulin (1; 10 micrograms/ml), and transferrin (Tr; 5 micrograms/ml) (DME/F12). After 24 h, the medium was replaced with DME/F12 degrees supplemented with SGF, EGF, 1, or Tr, in two-, three- or four-way combinations. Cell numbers were quantified after another 48 h of culture. EGF, I, and Tr, alone or in two-way combinations, were not mitogenic for TM4 cells. By contrast, SGF (1 U) alone, or with any two of these factors, stimulated TM4 cell proliferation to commensurate levels, and to twofold greater numbers than occurred with the combination of EGF, I, and Tr. Synergisms or inhibitions were not measurable. Follicle-stimulating hormone, luteinizing hormone, prolactin, acidic fibroblast growth factor, or basic fibroblast growth factor was weakly or not mitogenic for TM4 cells. The effect of SGF on cell proliferation was inhibited by 1 microM - 1 nM retinoic acid, but not by retinol or retinyl acetate. SGF was mitogenic for bovine adrenal capillary endothelial cells, an effect that was potentiated by 10 micrograms heparin/ml. Thus, SGF can induce proliferation of TM4 cells and capillary endothelial cells. The former provides a sensitive, and selective, serum-free, bioassay system for SGF activity.     

污水处理厂生物气溶胶抗生素抗性污染特征、来源及潜在风险

污水处理厂是抗生素抗性基因（antibiotic resistance genes,ARGs）和抗生素抗性细菌（antibiotic resistant bacteria,ARB）重要的源和汇,生物气溶胶是ARGs和ARB自污水处理厂向周边环境释放的关键载体。目前缺乏对污水处理厂生物气溶胶抗生素抗性污染特征、来源及潜在风险的系统性总结。本文从采样方法、检测方法、逸散特征、来源、潜在危害和风险评估等方面对污水处理厂抗生素抗性污染研究现状进行综述。惯性采样法和过滤法是常用的污水处理厂抗生素抗性生物气溶胶主要采集方法,而宏基因组测序、组装和分箱为其ARGs组成、可移动性和宿主提供了有效的检测方法,抗多药类、抗杆菌肽类、抗氨基糖苷类、抗四环素类、抗β-内酰胺类、抗磺胺类、抗大环内酯类和抗糖肽类等抗性基因在污水处理厂PM₁₀、PM_2.5和PM_1.0颗粒物中广泛检出。格栅间、生化反应池和污泥处理单元是污水处理厂PM₁₀、PM_2.5和PM_1.0负载ARGs和ARB的主要释放单元。污水处理厂不同粒径生物气溶胶中致病性ARB的存在增加了抗生素治疗的难度,而污水和污泥对ARGs和ARB的释放起到了重要的源的贡献。本文在研究内容、研究技术和控制策略等方面也提出了相关展望,以期为污水厂生物气溶胶抗生素抗性污染的监测和防护提供参考和借鉴。     

基于BEPS模型的云南省碳源/汇时空特征及其适用性分析

净初级生产力(NPP)和净生态系统生产力(NEP)是估算陆地生态系统碳源/汇的重要指标,云南为我国碳汇的主要区域之一,开展云南NPP和NEP时空变化特征分析对科学评估陆地生态系统碳源/汇功能,以及开展碳排放交易具有重要意义。基于BEPS模型1981—2019年NPP和NEP产品,采用线性趋势分析、文献对比等方法,研究云南NPP和NEP时空变化特征及其在云南的适用性。结果表明：(1)1981—1999年云南NPP和NEP呈水平波动,2000年后云南NPP和NEP呈明显波动上升趋势,2000—2019年云南NPP高值区域主要分布在西部和南部,而NEP高值区则主要分布在东部和西部局部地区;(2)2000—2019年云南NPP和NEP除西北部部分地区为下降趋势外,其余大部地区为上升趋势;(3)云南NPP峰值出现在7、8月,谷值出现在2月,NEP峰值出现月份与NPP基本相同,但谷值出现月份较NPP滞后1—3个月,6—10月是云南碳汇的主要月份;(4)BEPS模型估算的NPP与目前广泛应用的CASA和遥感模型结果较为一致,时空变化特征与云南生态恢复措施和气候特征吻合,其估算的NEP与陆地生物圈模型...     

中国森林生态系统林冠层降雨截留特征

森林生态系统作为陆地生态系统的主体,其发达的林冠层通过调节降水量、改变降水强度等深刻影响着流域全过程水文通量及水分输出。以中国广泛开展的典型森林降雨再分配过程的年尺度监测数据为基础,揭示中国不同类型森林生态系统的降雨再分配及林冠层降雨截留特征,阐明森林生态系统林冠层截留特征与降雨、植被要素的关系。结果表明:我国不同森林生态系统年穿透雨量处于141.4-2450.0 mm之间,年穿透雨率为36.3%-92.3%。5种典型森林生态系统多年平均穿透雨量((445.3±252.9)-(1230.6±479.6) mm)占同期多年平均降雨量的(72.6±9.2)%-(77.4±8.9)%。不同森林生态系统年树干茎流量介于0-508.2 mm之间,占同期年降雨量的0-25.8%。5种典型森林生态系统树干茎流量多年平均值((9.8±17.3)-(87.8±81.6) mm)占同期多年平均降雨量的(1.4±1.9)%-(5.4±4.6)%。不同森林生态系统林冠层年降雨截留范围在25.7-812.9 mm之间,占年降雨量的4.2%-55.6%。5种典型森林生态系统多年平均林冠截留量((154.2±81.6)-(392.2±203.5) mm)占同期年平均降雨量的(18.7±7.4)%-(25.9±8.3)%。进一步分析表明,我国森林生态系统穿透雨量、树干茎流量和林冠层截留量随观测区年降雨量的增加而呈显著增大(P<0.05),年穿透雨率、年树干茎流率随年降雨量的增加呈显著线性上升趋势(P<0.05),而年林冠截留率与年降雨量呈显著的负相关关系(P<0.01),降雨量、叶面积指数是深刻影响森林生态系统林冠层降雨截留率等特征的重要因素。整体上,不同类型森林生态系统林冠截留降雨能力存在明显差异,林冠层截留率突出表现为:落叶林大于常绿林、针叶林大于阔叶林。     

Nontraditional translation is the key to UFMylation and beyond

The Ubiquitin-fold modifier 1 (Ufm1) is a ubiquitin-like protein that can also be conjugated to protein substrates and subsequently alter their fates. Both UFMylation and de-UFMylation are mediated by Ufm1-specific proteases (UFSPs). In humans, it is widely believed that UFSP2 is the only active Ufm1 protease involved in Ufm1 maturation and de-UFMylation, whereas UFSP1 is thought to be inactive. Here, Liang et al. provide strong evidence showing that human UFSP1 is also an active Ufm1 protease. These results solve an age-old mystery in the human Ufm1 conjugation system and could have a greater impact not only on Ufm1 biology but also on the translation of genes employing nontraditional start codons.