• 【BRCA1和BRCA2种系突变携带者的预防标本中的偶然癌,重点是输卵管病变:6例病例报告并复习文献。】 复制标题 收藏 收藏
    DOI:10.1097/01.pas.0000202161.80739.ac 复制DOI
    作者列表:Carcangiu ML,Peissel B,Pasini B,Spatti G,Radice P,Manoukian S
    BACKGROUND & AIMS: :The identification of germ-line mutations in 2 genes (BRCA1 and BRCA2) responsible for the majority of hereditary ovarian cancers has led an increasing number of women carriers of these mutations to undergo prophylactic oophorectomy (PO) to reduce their risk of subsequent ovarian carcinoma. A large number of unexpected, clinically occult neoplasms are thus being discovered. Up to December 2004, the Medical Genetics Service of the National Cancer Institute in Milan, Italy, has tested 756 probands from breast and/or ovarian cancer families for BRCA1 and BRCA2 germ-line mutations. Molecular screening of family members led to the identification of 344 female carriers of BRCA1 (239) or BRCA2 (105) germ-line mutations. Of the 186 potentially eligible women (37 of whom had tested positive for BRCA1 and 13 for BRCA2 mutation), 50 (26.8%) chose to undergo PO. Six clinically occult primary gynecologic malignancies (2 stage IIIC serous carcinomas of the ovary, 3 in situ serous carcinomas of the fallopian tube, and 1 stage IIB invasive serous carcinoma of the fallopian tube) and 1 occult ovarian metastasis from breast carcinoma were identified in the PO specimens of 7 women (all BRCA1 mutated). Four of the patients with occult primary gynecologic cancers are alive without disease 129, 87, 38, and 7 months after PO, respectively. One of the 2 patients with primary ovarian cancer and the single patient with tubal invasive carcinoma are alive with recurrent disease 83 and 20 months after PO, respectively. In addition, one of the patients whose PO specimen did not show any malignancy presented with stage IIIC tubal carcinoma 77 months after PO. The relatively high number of tubal neoplasms found at PO in this group of patients underlines the linkage between mutation and the risk of developing tubal cancer, and stresses the need to include removal of the entire tubes at the time of PO and of thoroughly evaluating the specimens at the microscopic level. The upstaging of all 3 invasive carcinomas after staging surgery, and the late recurrence and persistence of 2 of them despite treatment indicate that small size of the tumors should not preclude therapy.
    背景与目标: :对负责大多数遗传性卵巢癌的2个基因(BRCA1和BRCA2)的种系突变的鉴定已导致越来越多的女性携带这些突变的女性进行预防性卵巢切除术(PO),以降低其患上卵巢癌的风险。因此,发现了大量意想不到的临床隐匿性肿瘤。截至2004年12月,意大利米兰国家癌症研究所的医学遗传学服务已对756个来自乳腺癌和/或卵巢癌家族的先证者进行了BRCA1和BRCA2种系突变测试。家庭成员的分子筛查导致鉴定出344例BRCA1(239)或BRCA2(105)种系突变的女性携带者。在186名可能符合条件的妇女中(其中37人的BRCA1测试呈阳性,13人的BRCA2突变检测为阳性),其中50人(26.8%)选择接受PO。在临床中确定了6例临床隐匿的原发性妇科恶性肿瘤(2例卵巢IIIC浆液性癌,3例输卵管原位浆液性癌和1例IIB输卵管浸润性浆液性癌)和1例隐匿性卵巢癌卵巢转移。 7名妇女的PO标本(所有BRCA1突变)。分别在PO后129、87、38和7个月,有四名患有隐匿性原发性妇科癌症的患者还活着而没有疾病。 2例原发性卵巢癌患者中的1例和输卵管浸润性癌的单例患者在PO后分别存活83个月和20个月。此外,其中一名PO标本未显示任何恶性肿瘤的患者在PO后77个月出现IIIC期输卵管癌。该组患者在PO中发现的相对较多的输卵管肿瘤强调了突变与发生输卵管癌的风险之间的联系,并强调需要在PO时切除整个管并彻底评估标本在微观层面上。分期手术后所有3种浸润性癌的分期升级,并且尽管有治疗,但其中2种仍较晚复发和持续存在,这表明较小的肿瘤不应排除治疗的可能性。
  • 【人类疾病基因中异常的5'剪接位点:突变模式,核苷酸结构以及预测其利用率的计算工具的比较。】 复制标题 收藏 收藏
    DOI:10.1093/nar/gkm402 复制DOI
    作者列表:Buratti E,Chivers M,Královicová J,Romano M,Baralle M,Krainer AR,Vorechovsky I
    BACKGROUND & AIMS: :Despite a growing number of splicing mutations found in hereditary diseases, utilization of aberrant splice sites and their effects on gene expression remain challenging to predict. We compiled sequences of 346 aberrant 5'splice sites (5'ss) that were activated by mutations in 166 human disease genes. Mutations within the 5'ss consensus accounted for 254 cryptic 5'ss and mutations elsewhere activated 92 de novo 5'ss. Point mutations leading to cryptic 5'ss activation were most common in the first intron nucleotide, followed by the fifth nucleotide. Substitutions at position +5 were exclusively G>A transitions, which was largely attributable to high mutability rates of C/G>T/A. However, the frequency of point mutations at position +5 was significantly higher than that observed in the Human Gene Mutation Database, suggesting that alterations of this position are particularly prone to aberrant splicing, possibly due to a requirement for sequential interactions with U1 and U6 snRNAs. Cryptic 5'ss were best predicted by computational algorithms that accommodate nucleotide dependencies and not by weight-matrix models. Discrimination of intronic 5'ss from their authentic counterparts was less effective than for exonic sites, as the former were intrinsically stronger than the latter. Computational prediction of exonic de novo 5'ss was poor, suggesting that their activation critically depends on exonic splicing enhancers or silencers. The authentic counterparts of aberrant 5'ss were significantly weaker than the average human 5'ss. The development of an online database of aberrant 5'ss will be useful for studying basic mechanisms of splice-site selection, identifying splicing mutations and optimizing splice-site prediction algorithms.
    背景与目标: :尽管在遗传性疾病中发现了越来越多的剪接突变,但异常剪接位点的利用及其对基因表达的影响仍然难以预测。我们编辑了346个异常5'剪接位点(5个)的序列,这些序列被166个人类疾病基因的突变激活。 5位共识中的突变占254个隐性5位,而其他地方的突变则激活了92位从头5位。导致隐伏5激活的点突变最常见于第一个内含子核苷酸,其次是第五个核苷酸。位置5的替换仅是G> A转换,这主要归因于C / G> T / A的高变异率。但是,第5位的点突变频率显着高于人类基因突变数据库中观察到的频率,这表明该位置的改变特别容易出现异常剪接,这可能是由于需要与U1和U6 snRNA进行顺序相互作用所致。隐秘5最好通过适应核苷酸依赖性的计算算法进行预测,而不是通过权重矩阵模型进行预测。与前者相比,将内含子5与真正的异物区别开来的效果不佳,因为前者在本质上要强于后者。新外显子5's的计算预测很差,表明它们的激活关键取决于外显子剪接增强剂或沉默子。异常5的真实对应物显着弱于普通人类5的。异常5的在线数据库的开发将对研究剪接位点选择的基本机制,鉴定剪接突变和优化剪接位点预测算法非常有用。
  • 【烟曲霉中的新型环境偶氮抗性突变及其在性繁殖中的性繁殖的可能作用。】 复制标题 收藏 收藏
    影响因子 :
    发表时间:2017-06-27
    来源期刊:mBio
    DOI:10.1128/mBio.00791-17 复制DOI
    作者列表:Zhang J,Snelders E,Zwaan BJ,Schoustra SE,Meis JF,van Dijk K,Hagen F,van der Beek MT,Kampinga GA,Zoll J,Melchers WJG,Verweij PE,Debets AJM
    BACKGROUND & AIMS: :This study investigated the dynamics of Aspergillus fumigatus azole-resistant phenotypes in two compost heaps with contrasting azole exposures: azole free and azole exposed. After heat shock, to which sexual but not asexual spores are highly resistant, the azole-free compost yielded 98% (49/50) wild-type and 2% (1/50) azole-resistant isolates, whereas the azole-containing compost yielded 9% (4/45) wild-type and 91% (41/45) resistant isolates. From the latter compost, 80% (36/45) of the isolates contained the TR46/Y121F/T289A genotype, 2% (1/45) harbored the TR46/Y121F/M172I/T289A/G448S genotype, and 9% (4/45) had a novel pan-triazole-resistant mutation (TR463/Y121F/M172I/T289A/G448S) with a triple 46-bp promoter repeat. Subsequent screening of a representative set of clinical A. fumigatus isolates showed that the novel TR463 mutant was already present in samples from three Dutch medical centers collected since 2012. Furthermore, a second new resistance mutation was found in this set that harbored four TR46 repeats. Importantly, in the laboratory, we recovered the TR463 mutation from a sexual cross between two TR46 isolates from the same azole-containing compost, possibly through unequal crossing over between the double tandem repeats (TRs) during meiosis. This possible role of sexual reproduction in the emergence of the mutation was further implicated by the high level of genetic diversity of STR genotypes in the azole-containing compost. Our study confirms that azole resistance mutations continue to emerge in the environment and indicates compost containing azole residues as a possible hot spot. Better insight into the biology of environmental resistance selection is needed to retain the azole class for use in food production and treatment of Aspergillus diseases.IMPORTANCE Composting of organic matter containing azole residues might be important for resistance development and subsequent spread of resistance mutations in Aspergillus fumigatus In this article, we show the dominance of azole-resistant A. fumigatus in azole-exposed compost and the discovery of a new resistance mutation with clinical relevance. Furthermore, our study indicates that current fungicide application is not sustainable as new resistance mutations continue to emerge, thereby threatening the use of triazoles in medicine. We provide evidence that the sexual part of the fungal life cycle may play a role in the emergence of resistance mutations because under laboratory conditions, we reconstructed the resistance mutation through sexual crossing of two azole-resistant A. fumigatus isolates derived from the same compost heap. Understanding the mechanisms of resistance selection in the environment is needed to design strategies against the accumulation of resistance mutations in order to retain the azole class for crop protection and treatment of Aspergillus diseases.
    背景与目标: :这项研究调查了两个堆肥堆中烟熏曲霉的耐唑性表型的动态变化,这两种情况下与吡咯暴露的情况相反:无吡咯和暴露于吡咯。在热休克后,对性但非无性孢子具有很高的抵抗力,无唑堆肥产生98%(49/50)野生型和2%(1/50)对唑的抗药性,而含吡咯的堆肥产生9%(4/45)野生型和91%(41/45)抗性分离株。在后一种堆肥中,80%(36/45)的分离株含有TR46 / Y121F / T289A基因型,2%(1/45)具有TR46 / Y121F / M172I / T289A / G448S基因型,9%(4 / 45)具有新的泛三唑抗性突变(TR463 / Y121F / M172I / T289A / G448S),具有三重46 bp启动子重复序列。随后对代表性的临床烟曲霉菌株进行了筛选,结果显示自2012年以来从三个荷兰医学中心收集的样品中已经存在新的TR463突变体。此外,在该菌株中发现了第二个新的耐药突变,其中包含四个TR46重复序列。重要的是,在实验室中,我们可能是通过减数分裂过程中双串联重复序列(TR)之间的不相等交换,从同一个含唑的堆肥中的两个TR46分离株之间的有性杂交中恢复了TR463突变。在含唑的堆肥中,STR基因型的高遗传多样性进一步暗示了性繁殖在突变出现中的这种可能作用。我们的研究证实,环境中仍会继续出现唑类抗性突变,并表明含有唑类残留物的堆肥可能是热点。需要更好地了解环境抗性选择的生物学以保持唑类用于食品生产和治疗曲霉病。重要事项含吡咯残留物的有机物的堆肥对于烟曲霉的抗性发展和随后的抗性突变的传播可能很重要。在本文中,我们显示了对吡唑有抵抗力的烟曲霉在暴露有吡咯的堆肥中的优势地位,并发现了与临床相关的新的抗药性突变。此外,我们的研究表明,随着新的耐药性突变的不断出现,当前杀菌剂的应用是不可持续的,从而威胁到三唑类药物的使用。我们提供的证据表明,真菌生命周期的性部分可能在耐药性突变的出现中起作用,因为在实验室条件下,我们通过对两个具有唑类抗性的A进行有性杂交来重建耐药性突变。来自同一堆肥堆的烟熏分离株。需要了解环境中抗性选择的机制,以设计针对抗性突变积累的策略,以保留用于农作物保护和曲霉病治疗的唑类。
  • 【Src激酶的抑制作用可阻断肾小球系膜细胞中高葡萄糖诱导的EGFR反式激活和胶原合成,并预防小鼠的糖尿病性肾病。】 复制标题 收藏 收藏
    DOI:10.2337/db12-1010 复制DOI
    作者列表:Taniguchi K,Xia L,Goldberg HJ,Lee KW,Shah A,Stavar L,Masson EA,Momen A,Shikatani EA,John R,Husain M,Fantus IG
    BACKGROUND & AIMS: :Chronic exposure to high glucose leads to diabetic nephropathy characterized by increased mesangial matrix protein (e.g., collagen) accumulation. Altered cell signaling and gene expression accompanied by oxidative stress have been documented. The contribution of the tyrosine kinase, c-Src (Src), which is sensitive to oxidative stress, was examined. Cultured rat mesangial cells were exposed to high glucose (25 mmol/L) in the presence and absence of Src inhibitors (PP2, SU6656), Src small interfering RNA (siRNA), and the tumor necrosis factor-α-converting enzyme (TACE) inhibitor, TAPI-2. Src was investigated in vivo by administration of PP2 to streptozotocin (STZ)-induced diabetic DBA2/J mice. High glucose stimulated Src, TACE, epidermal growth factor receptor (EGFR), mitogen-activated protein kinases (MAPKs), extracellular signal-regulated kinase (ERK1/2, p38), and collagen IV accumulation in mesangial cells. PP2 and SU6656 blocked high glucose-stimulated phosphorylation of Src Tyr-416, EGFR, and MAPKs. These inhibitors and Src knockdown by siRNA, as well as TAPI-2, also abrogated high glucose-induced phosphorylation of these targets and collagen IV accumulation. In STZ-diabetic mice, albuminuria, increased Src pTyr-416, TACE activation, ERK and EGFR phosphorylation, glomerular collagen accumulation, and podocyte loss were inhibited by PP2. These data indicate a role for Src in a high glucose-Src-TACE-heparin-binding epidermal growth factor-EGFR-MAPK-signaling pathway to collagen accumulation. Thus, Src may provide a novel therapeutic target for diabetic nephropathy.
    背景与目标: :长期暴露于高葡萄糖会导致糖尿病肾病,其特征是肾小球系膜基质蛋白(例如胶原蛋白)蓄积增加。已经记录了改变的细胞信号传导和基因表达并伴有氧化应激。检查了对氧化应激敏感的酪氨酸激酶c-Src(Src)的贡献。在存在和不存在Src抑制剂(PP2,SU6656),Src小干扰RNA(siRNA)和肿瘤坏死因子-α转化酶(TACE)的情况下,将培养的大鼠肾小球系膜细胞暴露于高葡萄糖(25 mmol / L)抑制剂,TAPI-2。通过对链脲佐菌素(STZ)诱导的糖尿病DBA2 / J小鼠施用PP2在体内研究了Src。高葡萄糖刺激的Src,TACE,表皮生长因子受体(EGFR),有丝分裂原激活的蛋白激酶(MAPK),细胞外信号调节激酶(ERK1 / 2,p38)和胶原IV在肾小球膜细胞中的蓄积。 PP2和SU6656阻止了Src Tyr-416,EGFR和MAPK的高葡萄糖刺激磷酸化。这些抑制剂和siRNA以及SAPI敲除的Src敲除也废除了高葡萄糖诱导的这些靶标的磷酸化和胶原IV的积累。在STZ糖尿病小鼠中,白蛋白尿,Src pTyr-416升高,TACE活化,ERK和EGFR磷酸化,肾小球胶原蛋白积聚和足细胞丢失均被PP2抑制。这些数据表明Src在高葡萄糖-Src-TACE-肝素结合表皮生长因子-EGFR-MAPK信号通路中积累胶原蛋白的作用。因此,Src可以为糖尿病性肾病提供新的治疗靶标。
  • 【应激特异性p38 MAPK激活足以驱动EGFR内吞作用,但不足以驱动其核易位。】 复制标题 收藏 收藏
    DOI:10.1242/jcs.202358 复制DOI
    作者列表:Tomas A,Jones S,Vaughan SO,Hochhauser D,Futter CE
    BACKGROUND & AIMS: :EGF receptor (EGFR) endocytosis is induced by stress in a manner dependent on the p38 MAPK family. Ligand and stresses such as X-rays, reportedly promote nuclear trafficking of endocytosed EGFR for regulation of gene transcription and DNA repair. We fail to detect EGFR endocytosis or nuclear transport following X-ray treatment of HeLa or head and neck cancer cells, despite extensive DNA damage induction. Apparent nuclear staining with EGFR extracellular domain antibody remained present despite reduced/absent EGFR expression, and so did not represent nuclear EGFR. UVB and UVC, but not X-ray or UVA, treatment induced p38 activation and EGFR endocytosis, although all of these stresses induced DNA damage, indicating that DNA damage alone is not sufficient to induce EGFR endocytosis. Increased reactive oxygen species (ROS) levels following UVB treatment, compared to that seen with X-rays, do not alone explain differences in p38 activation. UVB, like UVC, induced EGFR accumulation predominantly in perinuclear endosomes, rather than in the nucleus. Our morphological techniques identifying major changes in receptor distribution do not exclude the possibility that small but biologically relevant amounts of EGFR enter the nucleus. This study highlights the importance and limitations of morphological analyses of receptor distribution in understanding signaling outcome.
    背景与目标: :EGF受体(EGFR)的内吞作用是由应激以依赖于p38 MAPK家族的方式诱导的。据报道,配体和压力(例如X射线)可促进内吞EGFR的核转运,从而调节基因转录和DNA修复。尽管广泛的DNA损伤诱导作用,但在X射线治疗HeLa或头颈部癌细胞后,我们仍未检测到EGFR的内吞作用或核转运。尽管EGFR表达减少/缺失,但仍存在EGFR胞外域抗体的明显核染色,因此并不代表核EGFR。 UVB和UVC处理可诱导p38活化和EGFR内吞作用,而不是X射线或UVA,尽管所有这些压力均可诱导DNA损伤,表明仅DNA损伤不足以诱导EGFR内吞作用。与X射线相比,UVB处理后增加的活性氧(ROS)水平不能单独解释p38活化的差异。与UVC一样,UVB主要诱导EGFR聚集在核周内体中,而不是在细胞核中。我们的鉴定受体分布主要变化的形态学技术并不排除少量但生物学上相关的EGFR进入细胞核的可能性。这项研究强调了受体分布形态分析在理解信号转导结果中的重要性和局限性。
  • 【针对携带罕见EGFR突变的非小细胞肺癌的第一代EGFR-TKIs的结果:BE阳性研究的事后分析。】 复制标题 收藏 收藏
    DOI:10.1016/j.cllc.2017.05.016 复制DOI
    作者列表:
    BACKGROUND & AIMS: BACKGROUND:Beyond progression after tyrosine kinase inhibitor in EGFR-positive non-small-cell lung cancer patients (BE-POSITIVE) was the first Italian multicenter observational study that reported the outcomes of first-generation epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitors (TKIs) in a "real-life" Caucasian EGFR-mutated non-small-cell lung cancer (NSCLC) population. The sharing of multi-institutional experiences represents a crucial strategy to enrich knowledge about uncommon EGFR mutations. Therefore, we performed a post hoc analysis of the BE-POSITIVE study. PATIENTS AND METHODS:Data of advanced NSCLC patients with uncommon EGFR mutations who received first-line first-generation EGFR-TKIs in 24 Italian Hospitals were collected. In this analysis we aimed to evaluate overall survival (OS), progression-free survival (PFS), and overall response rate (ORR) of EGFR-TKIs in NSCLC patients harboring uncommon EGFR mutations. RESULTS:Thirty-five patients harboring uncommon EGFR mutations (any mutation other than deletion 19 or substitution of leucine by arginine at codon 858) were included of the original 312 EGFR-mutated cases. Most of them were female (n = 20, 57.1%), former smokers (n = 23, 65.7%), with adenocarcinoma (n = 31, 88.6%). The most frequent EGFR mutations were G719X (n = 6, 17.2%) and L861Q (n = 5, 14.2%). The population presented an ORR of 25.7%, a median PFS of 5.19 months, and a median OS of 14.49 months. When stratified according to type of EGFR mutation, median OS ranged from 3.65 months for unspecified mutations to 21.29 for double EGFR mutations. Median PFS ranged from 1.77 months for unspecified mutations to 20.83 months for concomitant EGFR-anaplastic lymphoma kinase alteration. ORR varied from 0% in exon 18, 20 and double gene alteration to 66.6% in exon 19. CONCLUSION:Our study supports the existence of a strong outcome heterogeneity within patients harboring uncommon EGFR mutations, which needs to be clarified to achieve a real personalized treatment strategy.
    背景与目标: 背景:酪氨酸激酶抑制剂在EGFR阳性非小细胞肺癌患者中取得的突破性进展(BE-POSITIVE)是第一项意大利多中心观察性研究,该研究报告了第一代表皮生长因子受体(EGFR)-酪氨酸激酶的结果现实生活中的白种人EGFR突变的非小细胞肺癌(NSCLC)人群中存在多种抑制剂(TKIs)。共享多机构经验代表了一项重要策略,可以丰富有关罕见的EGFR突变的知识。因此,我们对正研究进行了事后分析。
    患者和方法:收集了意大利24所医院接受一线第一代EGFR-TKI的EGFR突变罕见的晚期NSCLC患者的数据。在这项分析中,我们旨在评估具有罕见EGFR突变的NSCLC患者的EGFR-TKIs的总生存期(OS),无进展生存期(PFS)和总缓解率(ORR)。
    结果:35例罕见的EGFR突变病例包括了不常见的EGFR突变(除19号缺失或亮氨酸在858位密码子处亮氨酸替代以外的任何突变)。其中大多数是女性(n = 20,57.1%),曾吸烟者(n = 23,65.7%),患有腺癌(n = 31,88.6%)。最常见的EGFR突变是G719X(n = 6,17.2%)和L861Q(n = 5,14.2%)。人群的ORR为25.7%,PFS中位数为5.19个月,OS中位数为14.49个月。根据EGFR突变的类型进行分层时,中位OS​​范围从3.65个月(未指定突变)到21.29倍(双重EGFR突变)不等。 PFS的中位值范围从未指定突变的1.77个月到伴随的EGFR-再生障碍性淋巴瘤激酶改变的20.83个月不等。 ORR从外显子18、20和双基因改变的0%到外显子19的66.6%不等。
    结论:我们的研究支持携带罕见EGFR突变的患者存在强大的预后异质性,需要阐明这一点以实现真正的个性化治疗策略。
  • 【IGF信号基因的复发突变和骨肉瘤中基因组重排的不同模式。】 复制标题 收藏 收藏
    DOI:10.1038/ncomms15936 复制DOI
    作者列表:
    BACKGROUND & AIMS: :Osteosarcoma is a primary malignancy of bone that affects children and adults. Here, we present the largest sequencing study of osteosarcoma to date, comprising 112 childhood and adult tumours encompassing all major histological subtypes. A key finding of our study is the identification of mutations in insulin-like growth factor (IGF) signalling genes in 8/112 (7%) of cases. We validate this observation using fluorescence in situ hybridization (FISH) in an additional 87 osteosarcomas, with IGF1 receptor (IGF1R) amplification observed in 14% of tumours. These findings may inform patient selection in future trials of IGF1R inhibitors in osteosarcoma. Analysing patterns of mutation, we identify distinct rearrangement profiles including a process characterized by chromothripsis and amplification. This process operates recurrently at discrete genomic regions and generates driver mutations. It may represent an age-independent mutational mechanism that contributes to the development of osteosarcoma in children and adults alike.
    背景与目标: :骨肉瘤是影响儿童和成人的原发性骨恶性肿瘤。在这里,我们介绍了迄今为止最大的骨肉瘤测序研究,包括112种儿童和成人肿瘤,涵盖了所有主要的组织学亚型。我们研究的关键发现是在8/112(7%)的病例中鉴定胰岛素样生长因子(IGF)信号基因的突变。我们在另外87个骨肉瘤中使用荧光原位杂交(FISH)验证了这一观察结果,并在14%的肿瘤中观察到了IGF1受体(IGF1R)扩增。这些发现可能会为将来在骨肉瘤中使用IGF1R抑制剂的患者选择药物提供依据。分析突变的模式,我们确定了独特的重排图谱,包括以色杆菌病和扩增为特征的过程。该过程在离散的基因组区域反复进行,并产生驱动突变。它可能代表了与年龄无关的突变机制,有助于儿童和成年人中骨肉瘤的发展。
  • 【定量共表达EGFR和HER2对HER激活和运输的影响。】 复制标题 收藏 收藏
    DOI:10.1016/j.bbrc.2008.04.043 复制DOI
    作者列表:Shankaran H,Zhang Y,Opresko L,Resat H
    BACKGROUND & AIMS: :The human epidermal growth factor receptor (HER) system is an intricately regulated system that plays critical roles in development and tumorigenesis. Here, we apply integrated experimentation and modeling to analyze HER receptor activation in a panel of cell lines expressing endogenous levels of EGFR/HER1 and different levels of HER2. A mathematical model that includes the fundamental processes involved in receptor activation and trafficking was used to fit the experimental data, and values of the independent parameters for active receptor dimer formation affinities, trafficking rates and relative phosphorylation levels were estimated. Obtained parameter values quantitatively support the existing ideas on the effect of HER2 on EGFR dynamics, and enable us to predict the abundances of various phosphorylated receptor dimers in the cell lines.
    背景与目标: :人类表皮生长因子受体(HER)系统是一个复杂的调控系统,在发育和肿瘤发生中起着至关重要的作用。在这里,我们应用综合实验和建模来分析一组表达内源性EGFR / HER1和不同水平HER2的细胞系中的HER受体激活。使用包括参与受体激活和运输的基本过程的数学模型来拟合实验数据,并估计活性受体二聚体形成亲和力,运输速率和相对磷酸化水平的独立参数的值。获得的参数值定量支持关于HER2对EGFR动力学影响的现有观点,并使我们能够预测细胞系中各种磷酸化受体二聚体的丰度。
  • 【携带TP63基因从头突变的患者的犁骨发育不良(3q27)。】 复制标题 收藏 收藏
    DOI:10.1016/j.ijporl.2013.06.027 复制DOI
    作者列表:Schindler A,Guazzarotti L,Mameli C,Urbani E,Mozzanica F,Guerrini L,Zuccotti GV
    BACKGROUND & AIMS: :The congenital vomer defect (CVD) is a rare and still partially unknown condition. Only few cases have been reported in the international literature and the large majority of them appeared to be isolated. We report a case of CVD detected in a 7-year-old girl affected by ectodermal dysplasia clefting syndrome caused by a mutation of the TP63 gene.
    背景与目标: :先天性犁骨缺损(CVD)是一种罕见且仍部分未知的疾病。国际文献中仅报道了少数病例,其中大多数似乎是孤立的。我们报告了一个在7岁的女孩中检测到的CVD病例,该女孩受TP63基因突变引起的外胚层发育不良裂口综合征的影响。
  • 【线粒体TIM10亚基中保守带电残基的突变排除了TIM10复合物的组装,但并未消除酵母细胞的生长。】 复制标题 收藏 收藏
    DOI:10.1016/j.jmb.2007.06.025 复制DOI
    作者列表:Vergnolle MA,Alcock FH,Petrakis N,Tokatlidis K
    BACKGROUND & AIMS: :The Saccharomyces cerevisiae TIM10 complex (TIM10c) is an ATP-independent chaperone of the mitochondrial intermembrane space, involved in transport of polytopic membrane proteins. The complex is an alpha(3)beta(3) hexamer of Tim9 and Tim10 subunits. We have generated specific mutations in charged residues in the central core domain of each subunit delineated by the characteristic twin CX(3)C motif, and investigated the effect of these mutations on subunit folding, complex assembly and TIM10 function in vitro and in vivo. Any combination of mutations that included a specific glutamate residue, conserved in all known Tim9 and Tim10 sequences, abolished assembly of the TIM10 complex. In vivo complementation analyses using a MET3-TIM10 strain that is selectively inactivated for the expression of wild-type Tim10 showed that (i) an N-terminal deleted version of Tim10 that was previously shown to be defective in substrate binding is lethal under all conditions, but (ii) the charged residues mutant of Tim10 that is defective in assembly with Tim9 can restore growth in glucose, but not in non-fermentable carbon sources. These data suggest that formation of the hexamer is beneficial but not vital for TIM10 function, whilst the N-terminal substrate-binding region of Tim10 is essential in vivo.
    背景与目标: :酿酒酵母TIM10复合物(TIM10c)是线粒体膜间空间的一个不依赖ATP的分子伴侣,参与多膜蛋白的转运。该复合物是Tim9和Tim10亚基的alpha(3)beta(3)六聚体。我们已经在每个亚基的中央核心域带电残基中生成特定的突变,这些突变由特征性双胞胎CX(3)C图案描绘,并研究了这些突变对亚基折叠,复杂装配和TIM10功能的影响。在所有已知的Tim9和Tim10序列中均保守的,包括特定谷氨酸残基的任何突变组合都可消除TIM10复合体的装配。使用针对野生型Tim10的表达而选择性失活的MET3-TIM10菌株的体内互补分析显示(i)先前被证明在底物结合方面有缺陷的Tim10的N末端缺失版本在所有情况下都是致死性的,但(ii)与Tim9组装有缺陷的Tim10的带电荷残基突变体可以恢复葡萄糖的生长,但不能恢复不可发酵的碳源。这些数据表明六聚体的形成对TIM10功能是有益的,但不是至关重要的,而Tim10的N端底物结合区在体内则是必不可少的。
  • 【ErbB3,EGFR和Erk的激活对于人类乳腺癌细胞株具有对氟维司群的抗性至关重要。】 复制标题 收藏 收藏
    DOI:10.1007/s10549-008-0011-8 复制DOI
    作者列表:Frogne T,Benjaminsen RV,Sonne-Hansen K,Sorensen BS,Nexo E,Laenkholm AV,Rasmussen LM,Riese DJ 2nd,de Cremoux P,Stenvang J,Lykkesfeldt AE
    BACKGROUND & AIMS: :Seven fulvestrant resistant cell lines derived from the estrogen receptor alpha positive MCF-7 human breast cancer cell line were used to investigate the importance of epidermal growth factor receptor (ErbB1-4) signaling. We found an increase in mRNA expression of EGFR and the ErbB3/ErbB4 ligand heregulin2 (hrg2) and a decrease of ErbB4 in all resistant cell lines. Western analyses confirmed the upregulation of EGFR and hrg2 and the downregulation of ErbB4. Elevated activation of EGFR and ErbB3 was seen in all resistant cell lines and the ErbB3 activation occurred by an autocrine mechanism. ErbB4 activation was observed only in the parental MCF-7 cells. The downstream kinases pAkt and pErk were increased in five of seven and in all seven resistant cell lines, respectively. Treatment with the EGFR inhibitor gefitinib preferentially inhibited growth and reduced the S phase fraction in the resistant cell lines concomitant with inhibition of Erk and unaltered Akt activation. In concert, inhibition of Erk with U0126 preferentially reduced growth of resistant cell lines. Treatment with ErbB3 neutralizing antibodies inhibited ErbB3 activation and resulted in a modest but statistically significant growth inhibition of two resistant cell lines. These data indicate that ligand activated ErbB3 and EGFR, and Erk signaling play important roles in fulvestrant resistant cell growth. Furthermore, the decreased level of ErbB4 in resistant cells may facilitate heterodimerization of ErbB3 with EGFR and ErbB2. Our data support that a concerted action against EGFR, ErbB2 and ErbB3 may be required to obtain complete growth suppression of fulvestrant resistant cells.
    背景与目标: :使用七种来自雌激素受体α阳性MCF-7人乳腺癌细胞的抗氟维司群抗性细胞系,调查表皮生长因子受体(ErbB1-4)信号传导的重要性。我们发现在所有耐药细胞系中EGFR和ErbB3 / ErbB4配体heregulin2(hrg2)的mRNA表达增加,而ErbB4的减少。 Western分析证实了EGFR和hrg2的上调以及ErbB4的下调。在所有耐药细胞系中均观察到EGFR和ErbB3的激活增强,并且ErbB3的激活是通过自分泌机制发生的。仅在亲本MCF-7细胞中观察到ErbB4激活。下游激酶pAkt和pErk在七个抗性细胞系中的五个中和在所有七个抗性细胞系中分别增加。用EGFR抑制剂吉非替尼治疗可优先抑制生长并降低耐药细胞系中的S期分数,同时抑制Erk和未改变的Akt激活。一致地,用U0126抑制Erk优先降低了抗性细胞系的生长。用ErbB3中和抗体处理可抑制ErbB3活化,并导致对两种抗性细胞系的抑制作用达到中等但统计学上显着的增长。这些数据表明配体激活的ErbB3和EGFR,以及Erk信号传导在耐氟司韦特的细胞生长中起重要作用。此外,耐药细胞中ErbB4水平降低可能有助于ErbB3与EGFR和ErbB2异源二聚化。我们的数据支持可能需要针对EGFR,ErbB2和ErbB3的协同作用才能获得对氟维司群抗性细胞的完全生长抑制。
  • 【TP63中的新突变与年龄相关的病理有关。】 复制标题 收藏 收藏
    DOI:10.1038/sj.ejhg.5201888 复制DOI
    作者列表:Holder-Espinasse M,Martin-Coignard D,Escande F,Manouvrier-Hanu S
    BACKGROUND & AIMS: :Increases in the number of allelic malformation syndromes have led to their classification according to their pathogenesis rather than their clinical specific phenotype. TP63 (also known as TP73L) mutations have been identified in several such syndromes characterized by autosomal dominant transmission and various combinations of ectodermal dysplasia, limb malformations and orofacial clefting. TP63 has not yet been implicated in early aging phenotype in humans, even though p63 activates a program of cellular senescence and p63-compromised mice display features of accelerated aging. We report on a family with four affected adult females presenting with Rapp-Hodgkin syndrome (RHS), an autosomal dominant clinical entity that associates anhidrotic ectodermal dysplasia with cleft lip and palate. Features between RHS and EEC syndrome (ectrodactyly, ectodermal dysplasia and cleft lip/palate) have led to the recent identification of mutations in the TP63 gene, located on 3q27, in this condition. Our patients present typical clinical features of RHS, but also ophthalmic anomalies such as corneal dystrophy and premature menopause (around 30 years). The latter findings have never been reported in this condition, and could be secondary to a new TP63 deletion that has been identified in this family.
    背景与目标: :等位基因畸形综合征数量的增加已导致根据其发病机理而不是根据临床特定表型对它们进行分类。 TP63(也称为TP73L)突变已在几种以常染色体显性遗传和外胚层发育不良,肢体畸形和口唇裂的各种组合为特征的综合症中得到鉴定。 TP63尚未涉及人类的早期衰老表型,即使p63激活了细胞衰老程序并且p63受损的小鼠也表现出加速衰老的特征。我们报告了一个家庭,该家庭有四名受影响的成年女性,表现出拉普霍奇金综合征(RHS),这是一种常染色体显性临床实体,将无角质外胚层发育不良与唇left裂相关。 RHS和EEC综合征之间的特征(实际上是外胚层发育异常和唇裂/唇pal裂)导致这种情况下最近鉴定出位于3q27的TP63基因突变。我们的患者表现出RHS的典型临床特征,还表现出眼科异常,例如角膜营养不良和更年期提前(约30年)。后者的发现从未在这种情况下报道过,可能是该家族中已发现的新的TP63缺失的继发者。
  • 【在人支气管上皮细胞中,TGF-β1介导的COX-2诱导需要EGFR信号传导。】 复制标题 收藏 收藏
    DOI:10.1165/rcmb.2007-0100OC 复制DOI
    作者列表:Liu M,Yang SC,Sharma S,Luo J,Cui X,Peebles KA,Huang M,Sato M,Ramirez RD,Shay JW,Minna JD,Dubinett SM
    BACKGROUND & AIMS: :Cyclooxygenase-2 (COX-2) is a key enzyme in the production of prostaglandins and thromboxanes from free arachidonic acid. Increasing evidence suggests that COX-2 plays a role in tumorigenesis. A variety of stimuli induce COX-2 and it is overexpressed in many tumors, including non-small cell lung cancer (NSCLC). We studied the regulation of COX-2 expression in immortalized human bronchial epithelial cells (HBECs) by transforming growth factor-beta1 (TGF-beta1) and epidermal growth factor (EGF) because these two growth factors are present in both the pulmonary milieu of those at risk for lung cancer as well as in the tumor microenvironment. EGF significantly enhanced TGF-beta1-mediated induction of COX-2 and corresponding prostaglandin E2 (PGE2) production. TGF-beta1 and EGF induced COX-2 at the transcriptional and post-transcriptional levels. EGF receptor (EGFR) inhibition, neutralizing antibody against amphiregulin, or mitogen-activated protein kinase kinase (MEK) inhibition blocked TGF-beta1-mediated COX-2 induction. COX-2 induction by TGF-beta1 depended upon Smad3 signaling and required the activity of EGFR or its downstream mediators. Autocrine amphiregulin signaling maintains EGFR in a constitutively active state in HBECs, allowing for COX-2 induction by TGF-beta1. Thus, EGFR ligands, which are abundant in the pulmonary microenvironment of those at risk for lung cancer, potentiate and are required for COX-2 induction by TGF-beta1 in HBEC. These findings emphasize the central role of EGFR signaling in COX-2 induction by TGF-beta1 and suggest that inhibition of EGFR signaling should be investigated further for lung cancer prevention.
    背景与目标: :环氧合酶2(COX-2)是由游离花生四烯酸生产前列腺素和血栓烷的关键酶。越来越多的证据表明,COX-2在肿瘤发生中起作用。多种刺激可诱导COX-2的表达,并在许多肿瘤中过度表达,包括非小细胞肺癌(NSCLC)。我们通过转化生长因子-beta1(TGF-beta1)和表皮生长因子(EGF)研究了永生化人支气管上皮细胞(HBECs)中COX-2表达的调节,因为这两个生长因子均存在于它们的肺环境中处于肺癌以及肿瘤微环境中的风险。 EGF显着增强了TGF-beta1介导的COX-2诱导和相应的前列腺素E2(PGE2)的产生。 TGF-beta1和EGF在转录和转录后水平诱导COX-2。 EGF受体(EGFR)抑制,抗双调蛋白的中和抗体或丝裂原激活的蛋白激酶激酶(MEK)抑制可阻止TGF-beta1介导的COX-2诱导。 TGF-beta1诱导COX-2依赖于Smad3信号传导,并需要EGFR或其下游介质的活性。自分泌两性调节蛋白信号转导使HBEC中的EGFR保持组成型活性状态,从而允许TGF-beta1诱导COX-2。因此,在有肺癌风险的人的肺微环境中丰富的EGFR配体增强了,并且是HBEC中TGF-beta1诱导COX-2所必需的。这些发现强调了EGFR信号传导在TGF-beta1诱导COX-2诱导中的核心作用,并建议应进一步研究EGFR信号传导的抑制作用以预防肺癌。
  • 【小麦胞质乙酰辅酶A羧化酶可补充酵母中的ACC1空突变。】 复制标题 收藏 收藏
    DOI:10.1073/pnas.94.18.9990 复制DOI
    作者列表:Joachimiak M,Tevzadze G,Podkowinski J,Haselkorn R,Gornicki P
    BACKGROUND & AIMS: :Spores harboring an ACC1 deletion derived from a diploid Saccharomyces cerevisiae strain, in which one copy of the entire ACC1 gene is replaced with a LEU2 cassette, fail to grow. A chimeric gene consisting of the yeast GAL10 promoter, yeast ACC1 leader, wheat cytosolic acetyl-CoA carboxylase (ACCase) cDNA, and yeast ACC1 3' tail was used to complement a yeast ACC1 mutation. The complementation demonstrates that active wheat ACCase can be produced in yeast. At low concentrations of galactose, the activity of the "wheat gene" driven by the GAL10 promoter is low and ACCase becomes limiting for growth, a condition expected to enhance transgenic yeast sensitivity to wheat ACCase-specific inhibitors. An aryloxyphenoxypropionate and two cyclohexanediones do not inhibit growth of haploid yeast strains containing the yeast ACC1 gene, but one cyclohexanedione inhibits growth of the gene-replacement strains at concentrations below 0.2 mM. In vitro, the activity of wheat cytosolic ACCase produced by the gene-replacement yeast strain is inhibited by haloxyfop and cethoxydim at concentrations above 0.02 mM. The activity of yeast ACCase is less affected. The wheat plastid ACCase in wheat germ extract is inhibited by all three herbicides at concentrations below 0.02 mM. Yeast gene-replacement strains will provide a convenient system for the study of plant ACCases.
    背景与目标: :带有源自二倍体酿酒酵母菌株的ACC1缺失的孢子无法生长,其中整个ACC1基因的一个拷贝被LEU2盒代替。由酵母GAL10启动子,酵母ACC1前导序列,小麦胞质乙酰辅酶A羧化酶(ACCase)cDNA和酵母ACC1 3'尾部组成的嵌合基因用于补充酵母ACC1突变。互补表明,可以在酵母中产生活性小麦ACCase。在低浓度的半乳糖下,由GAL10启动子驱动的“小麦基因”的活性低,ACCase成为限制生长的条件,该条件有望增强转基因酵母对小麦ACCase特异性抑制剂的敏感性。芳氧基苯氧基丙酸酯和两种环己二酮不抑制含有酵母ACC1基因的单倍体酵母菌株的生长,但是一种环己二酮在低于0.2 mM的浓度下抑制基因置换菌株的生长。在体外,由基因替代酵母菌株产生的小麦胞质ACCase的活性在0.02 mM以上的浓度下被haloxyfop和cethoxydim抑制。酵母ACCase的活性受到的影响较小。浓度低于0.02 mM的所有三种除草剂均能抑制小麦胚芽提取物中的小麦质体ACCase。酵母基因置换菌株将为植物ACCases的研究提供方便的系统。
  • 【域特定的模拟磷酸化突变允许解剖不同的蛋白激酶C(PKC)同型触发的RNA结合蛋白HuR的活动。】 复制标题 收藏 收藏
    DOI:10.1016/j.cellsig.2013.08.003 复制DOI
    作者列表:Schulz S,Doller A,Pendini NR,Wilce JA,Pfeilschifter J,Eberhardt W
    BACKGROUND & AIMS: :The ubiquitous mRNA binding protein human antigen R (HuR) participates in the post-transcriptional regulation of many AU-rich element (ARE)-bearing mRNAs. Previously, by using in vitro kinase assay, we have identified serines (Ser) 158, 221 and 318 as targets of protein kinase C (PKC)-triggered phosphorylation. In this study, we tested whether GFP- or GST-tagged HuR constructs bearing a phosphomimetic Ser (S)-to-Asp (D) substitution at the different PKC target sites, would affect different HuR functions including HuR nucleo-cytoplasmic redistribution and binding to different types of ARE-containing mRNAs. The phosphomimetic GFP-tagged HuR protein bearing a phosphomimetic substitution in the hinge region of HuR (HuR-S221D) showed an increased cytoplasmic abundance when compared to wild-type HuR. Conversely, data from in vitro kinase assay and electrophoretic mobility shift assay (EMSA), implicates that phosphorylation at Ser 221 is not relevant for mRNA binding of HuR. Quantification of in vitro binding affinities of GST-tagged wild-type HuR and corresponding HuR proteins bearing a phosphomimetic substitution in either RRM2 (HuR-S158D) or in RRM3 (HuR-S318D) by microscale thermophoresis (MST) indicates a specific binding of wild-type HuR to type I, II or type III-ARE-oligonucleotides in the high nanomolar range. Interestingly, phosphomimetic mutation at position 158 or 318 had a negative influence on HuR binding to type I- and type II-ARE-mRNAs whereas it significantly enhanced HuR affinity to a type III-ARE substrate. Our data suggest that differential phosphorylation of HuR by PKCs at different HuR domains coordinates subcellular HuR distribution and leads to a preferential binding to U-rich bearing target mRNA.
    背景与目标: :普遍存在的mRNA结合蛋白人类抗原R(HuR)参与许多富含AU元素(ARE)的mRNA的转录后调控。以前,通过使用体外激酶测定,我们已经将丝氨酸(Ser)158、221和318确定为蛋白激酶C(PKC)触发的磷酸化的靶标。在这项研究中,我们测试了在不同的PKC目标位点上带有磷酸仿Ser(S)-Asp(D)取代的GFP标记或GST标记的HuR构建体是否会影响不同的HuR功能,包括HuR核质重分布和结合对不同类型的含有ARE的mRNA的表达。与野生型HuR相比,在HuR的铰链区(HuR-S221D)带有磷酸模拟取代的磷酸化GFP标记的HuR蛋白显示出增加的细胞质丰度。相反,来自体外激酶测定和电泳迁移率变动测定(EMSA)的数据暗示Ser 221的磷酸化与HuR的mRNA结合无关。定量标记GST标签的野生型HuR和相应的在磷酸RRM2(HuR-S158D)或RRM3(HuR-S318D)中具有磷酸模拟取代作用的HuR蛋白的体外结合亲和力表明通过微型热电泳(MST) -在高纳摩尔范围内将HuR型转变为I,II或III型-ARE-寡核苷酸。有趣的是,位置158或318处的磷酸模拟突变对HuR与I型和II型ARE-mRNA的结合具有负面影响,而它显着增强了HuR对III型ARE底物的亲和力。我们的数据表明,PKCs在不同的HuR域上对HuR的磷酸化差异会协调亚细胞HuR的分布,并导致与富含U的靶标mRNA的优先结合。

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