后基因组时代的工业生物技术

简述了工业生物技术的发展背景和意义,分析了基因组学和功能基因组学发展对工业生物技术的推动作用,重点介绍了本期专刊发表的代谢工程、发酵工程以及工业酶与生物催化领域的17篇论文。     

DNA polymerase proofreading: Multiple roles maintain genome stability

DNA polymerase proofreading is a spell-checking activity that enables DNA polymerases to remove newly made nucleotide incorporation errors from the primer terminus before further primer extension and also prevents translesion synthesis. DNA polymerase proofreading improves replication fidelity ∼ 100-fold, which is required by many organisms to prevent unacceptably high, life threatening mutation loads. DNA polymerase proofreading has been studied by geneticists and biochemists for > 35 years. A historical perspective and the basic features of DNA polymerase proofreading are described here, but the goal of this review is to present recent advances in the elucidation of the proofreading pathway and to describe roles of DNA polymerase proofreading beyond mismatch correction that are also important for maintaining genome stability.     

Foetal hepatocyte transplantation in a vascularized AV-Loop transplantation model in the rat

The use of foetal liver cells (FLC) in the context of hepatic tissue engineering might permit efficient in vitro expansion and cryopreservation in a cell bank. A prerequisite for successful application of bioartificial liver tissue is sufficient initial vascularization. In this study, we evaluated the transplantation of fibrin gel-immobilized FLC in a vascularized arterio-veno-venous (AV)-loop model. FLC were isolated from embryonic/foetal (ED 16) rat livers and were enriched by using magnetic cell sorting (MACS). After cryopreservation, FLC were labelled by pkh-26. Cells were transplanted in a fibrin matrix into a subcutaneous chamber containing a microsurgically created AV-loop in the femoral region of the recipient rat. The chambers were explanted after 14 days. Subcutaneous implants without an AV-loop and cell-free implants served as controls. Fluorescence microscopy of the constructs was used to identify pkh-26⁺- donor cells. Characterization was performed by RT-PCR and immunhistology (IH) for CK-18 and CD31. Transplantation of FLC using the AV-loop permitted a neo -tissue formation in the fibrin matrix. A high-density vascularization was observed in the AV-loop constructs as shown by CD31 IH. Viable foetal donor cells were detected which expressed CK-18. FLC can be successfully used for heterotopic transplantation. Fibrin matrix permits rapid blood vessel ingrowth from the AV-loop and supports engraftment of FLC. It is therefore an appropriate environment for hepatocyte transplantation in combination with microsurgical vascularization strategies. Transplantation of fibrin gel-immobilized FLC may be a promising approach for the development of highly vascularized in vivo tissue-engineering-based liver support systems.     

Engineered heart tissue graft derived from somatic cell nuclear transferred embryonic stem cells improve myocardial performance in infarcted rat heart

The concept of regenerating diseased myocardium by implanting engineered heart tissue (EHT) is intriguing. Yet it was limited by immune rejection and difficulties to be generated at a size with contractile properties. Somatic cell nuclear transfer is proposed as a practical strategy for generating autologous histocompatible stem (nuclear transferred embryonic stem [NT‐ES]) cells to treat diseases. Nevertheless, it is controversial as NT‐ES cells may pose risks in their therapeutic application. EHT from NT‐ES cell‐derived cardiomyocytes was generated through a series of improved techniques in a self‐made mould to keep the EHTs from contraction and provide static stretch simultaneously. After 7 days of static and mechanical stretching, respectively, the EHTs were implanted to the infarcted rat heart. Four weeks after transplantation, the suitability of EHT in heart muscle repair after myocardial infarction was evaluated by histological examination, echocardiography and multielectrode array measurement. The results showed that large (thickness/diameter, 2–4 mm/10 mm) spontaneously contracting EHTs was generated successfully. The EHTs, which were derived from NT‐ES cells, inte grated and electrically coupled to host myocardium and exerted beneficial effects on the left ventricular function of infarcted rat heart. No teratoma formation was observed in the rat heart implanted with EHTs for 4 weeks. NT‐ES cells can be used as a source of seeding cells for cardiac tissue engineering. Large contractile EHT grafts can be constructed in vitro with the ability to survive after implantation and improve myocardial performance of infarcted rat hearts.     

类弹性蛋白多肽及其在生物医学材料的应用

类弹性蛋白多肽是一种人造基因工程蛋白质聚合物,其结构主要由五肽重复串连序列单元 (GVGXP) 的这一肽段单元重复组成。由于具有可逆相变特征,并可进行高通量生产,加之良好的生物相容性及生物可降解性,使其在新型生物医学材料方面展示了广阔的应用前景。概括了类弹性蛋白多肽的相变机理、合成方法及在生物医学材料上的应用,重点阐述了类弹性蛋白多肽在组织工程、靶向肿瘤、构造药物载体微粒的应用。     

Devil inside: does plant programmed cell death involve the endomembrane system?

Eukaryotic cells have to constantly cope with environmental cues and integrate developmental signals. Cell survival or death is the only possible outcome. In the field of animal biology, tremendous efforts have been put into the understanding of mechanisms underlying cell fate decision. Distinct organelles have been proven to sense a broad range of stimuli and, if necessary, engage cell death signalling pathway(s). Over the years, forward and reverse genetic screens have uncovered numerous regulators of programmed cell death (PCD) in plants. However, to date, molecular networks are far from being deciphered and, apart from the autophagic compartment, no organelles have been assigned a clear role in the regulation of cellular suicide. The endomembrane system (ES) seems, nevertheless, to harbour a significant number of cell death mediators. In this review, the involvement of this system in the control of plant PCD is discussed in‐depth, as well as compared and contrasted with what is known in animal and yeast systems.     

Enzyme activity of the phenylpropanoid pathway as a response to apple scab infection

The study was performed on apple trees, ‘Golden Delicious' cv., which is a scab-susceptible cultivar. The phenolic content of apple fruit was determined in different parts of the peel. The phenolic compounds were analysed in the scab spot, in the tissue around the spot and in the healthy tissue. We determined the concentration of various phenolic compounds and related enzyme activities. Infection with the Venturia inaequalis fungus enhanced the metabolism of phenolic compounds at the scab spot, around the spot and in healthy peel. Compared with the healthy tissue and the tissue around the spot, the scab spot showed higher enzyme activity for all tested enzymes, except for dihydrochalcone 2′-O-glucosyltransferase, which had lower activity in the scab spot. In comparison to the healthy peel, the scab spot showed up to 3.4 times more hydroxycinnamic acids, up to 1.1 times more dihydrochalcones and up to 1.4 times more flavan-3-ols. In contrast, the healthy peel showed up to 1.6 times more flavonols than the scab spot.     

Involvement of lectin pathway activation in the complement killing of Giardia intestinalis

Giardia intestinalis (syn. G. lamblia, G. duodenalis) is a flagellated unicellular eukaryotic microorganism that commonly causes diarrheal disease throughout the world. In humans, the clinical effects of Giardia infection range from the asymptomatic carrier state to a severe malabsorption syndrome possibly due to different virulence of the Giardia strain, the number of cysts ingested, the age of the host, and the state of the host immune system at the time of infection.The question about how G. intestinalis is controlled by the organism remains unanswered. Here, we investigated the role of the complement system and in particular, the lectin pathway during Giardia infections. We present the first evidence that G. intestinalis activate the complement lectin pathway and in doing so participate in eradication of the parasite. We detected rapid binding of mannan-binding lectin, H-ficolin and L-ficolin to the surface of G. intestinalis trophozoites and normal human serum depleted of these molecules failed to kill the parasites. Our finding provides insight into the role of lectin pathway in the control of G. intestinalis and about the nature of surface components of parasite.