Normal expression of thymidine kinase and O6-methylguanine-DNA methyltransferase in cultured fibroblasts from individuals with hereditary galactokinase deficiency

Expression of the enzymes galactokinase, thymidine kinase, and O⁶-methylguanine-DNA methyltransferase is occasionally coordinately regulated in human cell lines. We have measured the activities of these three enzymes in extracts of fibroblasts from individuals with hereditary galactokinase deficiency. These cells do not express measurable galactokinase activity. The levels of O⁶-methylguanine-DNA methyltransferase were in the normal range in cells from three galactokinase-deficient individuals. The activity of thymidine kinase in the affected cells was in the normal range for two of the three individuals. The reduced thymidine kinase activity in the third individual reflected the extremely poor growth of the cells in culture. Immortalization of one galactokinase-deficient cell line resulted in loss of O⁶-methylguanine-DNA methyltransferase activity, but the galactokinase and thymidine kinase levels remained unchanged. The data indicate that the loss of galactokinase activity in these individuals is the consequence of an alteration of gene expression which does not involve coordinate silencing with the thymidine kinase and methyltransferase loci.     

Methylation of TMV RNA

Several plant and animal viral RNAs contain a tRNA like structure at their 3′ ends. In this communication we show that tobacco mosaic virus (TMV) RNA is an acceptable substrate for a specific tRNA methyltransferase. Using a crude preparation of $\text{E. coli}$ ribothymidine (rT) forming uracil methylase and (methyl ³H) S-adenosyl-L-methionine (SAM) as a methyl donor, 0.7 moles of methyl group is incorporated per mole of TMV RNA in 10 hours at 30°C. Upon T₂ RNAse digestion of the labeled RNA, all of the radioactivity was found to be in TMP. T₁ RNAse digestion of ³H methylated TMV RNA showed that all of the label was located in a tetranucleotide which co-migrated with authentic TpψpCpGp, an oligonucleotide characteristically found in normal cellular tRNA.The use of this specific methyl transferase reaction may provide a simple assay for the detection of tRNA like structures in large RNAs.     

A Comprehensive Analysis of Symmetric Arginine Dimethylation in Colorectal Cancer Tissues Using Immunoaffinity Enrichment and Mass Spectrometry

Protein arginine methyltransferase 5 (PRMT5) is a major enzyme responsible for generating monomethyl and symmetric dimethyl arginine in proteins. PRMT5 is essential for cell viability and development, and its overexpression is observed in a variety of cancers. In the present study, it is found that levels of PRMT5 protein and symmetric arginine dimethylation in colorectal cancer (CRC) tissues are increased compared to those in adjacent noncancerous tissues. Using immunoaffinity enrichment of methylated peptides combined with high‐resolution mass spectrometry, a total of 147 symmetric dimethyl‐arginine (SDMA) sites in 94 proteins are identified, many of which are RNA binding proteins and enzymes. Quantitative analysis comparing CRC and normal tissues reveals significant increase in the symmetric dimethylation of 70 arginine sites in 46 proteins and a decrease in that of four arginine sites in four proteins. Among the 94 proteins identified in this study, it is confirmed that KH‐type splicing regulatory protein is a target of PRMT5 and highly expressed in CRC tissues compared to noncancerous tissues. This study is the first comprehensive analysis of symmetric arginine dimethylation using clinical samples and extends the number of known in vivo SDMA sites. The data obtained are available via ProteomeXchange with the identifier PXD015653.     

Methyltransferase like 3 promotes colorectal cancer proliferation by stabilizing CCNE1 mRNA in an m6A-dependent manner

m6A modification is the most prevalent RNA modification in eukaryotes. As the critical N6-methyladenosine (m6A) methyltransferase, the roles of methyltransferase like 3 (METTL3) in colorectal cancer (CRC) are controversial. Here, we confirmed that METTL3, a critical m6A methyltransferase, could facilitate CRC progression in vitro and in vivo. Further, we found METTL3 promoted CRC cell proliferation by methylating the m6A site in 3′-untranslated region (UTR) of CCNE1 mRNA to stabilize it. Moreover, we found butyrate, a classical intestinal microbial metabolite, could down-regulate the expression of METTL3 and related cyclin E1 to inhibit CRC development. METTL3 promotes CRC proliferation by stabilizing CCNE1 mRNA in an m6A-dependent manner, representing a promising therapeutic strategy for the treatment of CRC.     

盐酸右美托咪定联合盐酸罗哌卡因胸椎旁神经阻滞对肺癌根治术患者血清炎性因子和免疫学指标的影响

摘要 目的：探讨盐酸右美托咪定联合盐酸罗哌卡因胸椎旁神经阻滞（TPVB）对肺癌根治术患者血清炎性因子和免疫学指标的影响。方法：选取2018年4月~2020年4月期间于我院行肺癌根治术的患者400例,根据信封抽签法分为对照组和观察组,各200例,对照组给予盐酸罗哌卡因TPVB,观察组在对照组基础上联合盐酸右美托咪定,对比两组疼痛、炎性因子、免疫学指标、血流动力学及不良反应。结果：观察组术后6 h（T5）~术后48h（T8）时间点视觉模拟评分法（VAS）评分均低于对照组（P<0.05）。对照组插管后5 min（T1）~术毕（T3）时间点心率（HR）及平均动脉压（MAP）较麻醉诱导前（T0）时间点升高（P<0.013）,观察组T1~T3时间点HR、MAP低于对照组（P<0.05）。两组T8时间点白介素-6（IL-6）、C反应蛋白（CRP）、肿瘤坏死因子-α（TNF-α）均较T0下降,且观察组低于对照组（P<0.05）。两组T8时间点CD4⁺、CD4⁺/CD8⁺均较T0时间点下降,但观察组高于对照组（P<0.05）,CD8⁺均较T0时间点升高,但观察组低于对照组（P<0.05）。观察组不良反应总发生率低于对照组（P<0.05）。结论：盐酸右美托咪定联合盐酸罗哌卡因TPVB用于肺癌根治术患者,可稳定血流动力学,且可获得较好的镇痛效果,降低不良反应发生率,减轻术后炎性损伤及免疫抑制。     

肺癌根治术后并发症危险因素分析及其生活质量变化研究

摘要 目的：探讨肺癌根治术后并发症的危险因素,并分析其生活质量的变化。方法：纳入我院2018年9月～2020年7月收治的行肺癌根治术患者104例,对患者的临床资料进行回顾性分析。根据患者术后3个月的并发症发生情况,分成并发症组（n=32）和非并发症组（n=72）,分析肺癌根治术后并发症发生的影响因素,利用癌症病人生活质量测定量表（QLQ-C30）评估患者术前及术后3个月的生活质量。结果：肺癌根治术后32例出现并发症,其中切口感染6例,肺炎7例,肺不张9例,心律失常6例,脓胸4例。并发症组年龄≥60岁、烟龄≥10年、传统开胸术、慢性阻塞性肺疾病史人数占比高于非并发症组（P＜0.05）。多因素Logistic回归分析显示,年龄≥60岁（OR=2.978,95%CI：1.415-6.267）、烟龄≥10年（OR=3.847,95%CI：1.869-7.918）、传统开胸术（OR=3.065,95%CI：1.544-6.084）、慢性阻塞性肺疾病史（OR=2.848,95%CI：1.481-5.477）是肺癌根治术后患者发生并发症的危险因素（P＜0.05）。非并发症组术后3个月的总体生活质量量表、角色功能、躯体功能、情绪功能、社会功能评分高于术前与并发症组,且恶心呕吐、疲乏、疼痛、便秘、呼吸困难、食欲下降、睡眠障碍、腹泻评分较术前与并发症组明显降低（P＜0.05）。结论：肺癌根治术后并发症的发生主要与患者年龄、烟龄、手术方式、慢性阻塞性肺疾病史有关,且并发症对患者术后生活质量影响较大。     

腹腔镜辅助远端胃癌根治术中十二指肠优先离断对胃癌患者应激反应、炎性因子和生活质量的影响

摘要 目的：探讨腹腔镜辅助远端胃癌根治术中十二指肠优先离断对胃癌患者应激反应、炎性因子和生活质量的影响。方法：回顾性选取2018年3月~2020年12月期间河北省邯郸市中心医院收治的行腹腔镜辅助远端胃癌根治术的胃癌患者93例,根据患者手术方式的不同将患者分为对照组45例和研究组48例,对照组给予左侧后入路的腹腔镜辅助远端胃癌根治术,研究组给予十二指肠优先离断腹腔镜辅助远端胃癌根治术,比较两组围术期指标、应激反应、炎性因子、生活质量以及术后并发症情况。结果：两组清除淋巴结数量对比差异无统计学意义（P>0.05）,研究组手术时间、胃肠道功能恢复时间、住院时间短于对照组,术中出血量少于对照组（P<0.05）。研究组术后3 d、术后5 d血糖、皮质醇、肾上腺素低于对照组（P<0.05）。研究组术后3 d、术后5 d白介素-6（IL-6）、C反应蛋白（CRP）、肿瘤坏死因子-?琢（TNF-?琢）低于对照组（P<0.05）。研究组术后1个月主观症状、生理功能状态、社会活动功能、心理情绪状态评分高于对照组（P<0.05）。两组术后并发症发生率对比无差异（P>0.05）。结论：与左侧后入路的腹腔镜辅助远端胃癌根治术相比,十二指肠优先离断可简化腹腔镜辅助远端胃癌根治术的手术步骤,减少胃癌患者术后应激反应和炎性反应,可有效促进患者术后恢复。     

The Radical SAM Superfamily

ABSTRACT

The radical S-adenosylmethionine (SAM) superfamily currently comprises more than 2800 proteins with the amino acid sequence motif CxxxCxxC unaccompanied by a fourth conserved cysteine. The charcteristic three-cysteine motif nucleates a [4Fe–4S] cluster, which binds SAM as a ligand to the unique Fe not ligated to a cysteine residue. The members participate in more than 40 distinct biochemical transformations, and most members have not been biochemically characterized. A handful of the members of this superfamily have been purified and at least partially characterized. Significant mechanistic and structural information is available for lysine 2,3-aminomutase, pyruvate formate-lyase, coproporphyrinogen III oxidase, and MoaA required for molybdopterin biosynthesis. Biochemical information is available for spore photoproduct lyase, anaerobic ribonucleotide reductase activation subunit, lipoyl synthase, and MiaB involved in methylthiolation of isopentenyladenine-37 in tRNA. The radical SAM enzymes biochemically characterized to date have in common the cleavage of the [4Fe–4S]^{1 +} –SAM complex to [4Fe–4S]^{2 +}–Met and the 5′ -deoxyadenosyl radical, which abstracts a hydrogen atom from the substrate to initiate a radical mechanism.