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Cloning and expression of AmphiSRP19, a homologue of SRP19 from amphioxus Branchiostoma belcheri tsingtauense

Cloning and expression of AmphiSRP19, a homologue of SRP19 f...

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中国细胞生物学学会第八届会员代表大会暨学术大会

作者： Mei Liu, Shicui Zhang, Zhenhui Liu, Yongjun Wang, Hongyan Li and Anlong Xu Department of Marine Biology, Ocean University of China, Qingdao, 266003, P.R. China. College of Life Science, Zhongshan University, Guangzhou, 510275, P.R. China.

Signal recognition particle (SRP) plays a critical role in the co-translational translocation of membrane bound and secreted proteins. An essential component of SRP is the protein SRP 19, which is the initiator of SRP assembly. A cDNA clone, AmphiSRP19, encoding the protein SRP19 is isolated from the gut cDNA library of Branchiostoma belcheri tsingtaunese. The predicted amino acid sequence of AmphiSRP19 exhibits 52.8%, 54.9%, 42.3%, 35.3%, 28.2% and 19.7%-24.6% similarity to its known counterparts from human, mouse, Drosophila, rice, thale cress and five fungi species, respectively. Like other eukaryotic SRP19 proteins, AmphiSRP19 has the most conserved amino acid residues located at the sites required for binding to SRP RNA. This is the first SRP19 gene isolated from protochordates. Northern blotting and in situ hybridization analyses show AmphiSRP19 is expressed in all the tissues examined, including fully-grown ovaries and testes, muscle, notochord, gill and gut of adult amphioxus, and in all the embryos and larvae ranging from zygote to 2-day larva. These agree well with the expectation that SRP19 is an essential house-keeping gene.

关键词： SRP Cloning and expression of AmphiSRP19 a homologue of SRP19 from amphioxus Branchiostoma belcheri tsingtauense

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Development of Proteomic Patterns for Detecting Lung Cancer

Development of Proteomic Patterns for Detecting Lung Cancer

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中国细胞生物学学会第八届会员代表大会暨学术大会

作者： Dacheng He Xueyuan Xiao, Danhui Liu, Ying Tang, Fuzheng Guo, Liang Xia, Jin Liu, Key laboratory for Cell Proliferation and Regulation Biology Ministry of Education, Beijing Normal University, Universities' of Confederated Institute for Proteomics, Beijing 100875, China

Lung cancer is at present the number one cause of cancer death and no biomarker is available to detect lung cancer in serum samples so far. Blood serum is a complex body fluid that contains various proteins including secreting proteins from cell or tissues. It is clear that discovery of lung cancer soluble biomarkers in serum for early detection becomes an urgent task in clinic. The objective of this study is to find specific biomarkers for detection of lung cancer using Surface Enhanced Laser Desorption/Ionization (SELDI) technique. SELDI protein chips can capture the specific proteins in mixture by means of specific surface chemistries following binding and washing. In this study, serum samples from 30 lung cancer patients and 51 age- and sex-matched healthy peoples were analyzed by SELDI on a ProteinChip reader, PBSII-C. The spectra were generated on WCX2 chips and protein peaks clustering and classification analyses were performed utilizing Biomarker Wizard and Biomarker Patterns software packages, respectively. Three protein peaks were automatically chosen for the system training and the development of a decision classification tree. The constructed model was then used to test an independent set of masked serum samples from 30 lung cancer patients and 31 healthy individuals. The analysis yielded a sensitivity of 93.3%, and a specificity of 96.7%. These results suggest that the serum is a capable resource for detection of specific lung cancer biomarkers. SELDI technique combined with an artificial intelligence classification algorithm can both facilitate the discovery of better biomarkers for lung cancer and provide a useful tool for molecular diagnosis in future.

关键词： Development of Proteomic Patterns for Detecting Lung Cancer

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Stress Protein GRP170 Localizes Down-stream of c-Myc and Has Oncogenic Potential

Stress Protein GRP170 Localizes Down-stream of c-Myc and Has...

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中国细胞生物学学会第八届会员代表大会暨学术大会

作者： Chang Ming Fang and Yong Hua Xu (Laboratory of Molecular and Cellular Oncology, Shanghai Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, 320 Yue-yang Road, Shanghai, 200031, P.R. China)

Cells up-regulate the expression of molecular chaperones, such as glucose/oxygen-regulated proteins (GRPs/ORPs) and heat-shock proteins (HSPs), when subjected to environmental stress. GRP170/ORP150 is a novel endoplasmic reticulum-associated protein in the HSP70 family, which was first purified for its capacity to promote cell survival under the condition of hypoxia. We here reported that hamster grp170 was highly expressed in CHO-c-mycER cells when c-MycER was activated by the synthetic steroid, 4-hydroxy-tamoxifen (OHT). Moreover, the expression of its human homologue orp150 increased in some of human hepaitocellular carcinomas in which elevated c-myc expression was frequently detected. Our data indicated put GRP170/ORP150 down-steam of c-Myc signal pathway. The forced expression of grp170 in CHO cells stimulated the anchorage-dependent and anchorage-independent growth, reduced the serum dependency of cell growth in vitro. Serum starvation experiments showed that the over-expression of grp170 rescued the CHO cells frohi serum free induced DNA degradation and apoptosis of cells. Furthermore, the grp170 highly expressed CHO cells showed an increased potential of tumorigenicity in nude mice. The results suggest that grp170/orp150 is a down-stream target gene of oncogene c-Myc playing an active role in oncogenic transformation.

关键词： Differential display c-myc glucose/oxygen-regulated protein (GRP/ORP)

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High-frequency plant regeneration frbm Astragalus melilotoides via somatic embryogenesis or organogenesis

High-frequency plant regeneration frbm Astragalus melilotoid...

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中国细胞生物学学会第八届会员代表大会暨学术大会

作者：侯岁稳贾敬芬兰州大学生命科学学院西北大学生命科学学院

Astragalus melilotoides pall, is widely cultivated in many arid and desert regions of northChina for improving pasture because it is very nutritious to livestock. The plant can also be utilized as windbreaks or for crop rotation, sand-fixation and desalination because of its tolerance to drought, coldness, salinity and disease. Biotechnological approaches such as somatic hybridization and genetic transformation can be used for not only improving this crop, but also transferring its favorable resistant characters to other legumes. Here we reported a highly efficient and reproducible procedure developed for plant regeneration from hypocotyl or hypocotyl- and stem-derived calli of Astragalus melilotoides via somatic embryogenesis or organogenesis. High frequency of somatic embryos was induced directly from hypocotyl explants on MS supplemented with NAA (2.69 μM) and BA (4.44 μM) within 5 weeks. The development of somatic embryos was asynchronous and the globular, heart, torpedo and cotyledonary somatic embryos could be observed in the course. All of the somatic embryos developed in a single state and could be separated from hypocotyls easily. Some somatic embryos developed directly into whole plantlets with well-formed shoots and roots on the same medium. Three types of calli were induced when hypocotyl and stem segments were cultured on MS medium containing 2,4-D (9.05 μM) and BA (2.22 or 4.44 μM). Indirect somatic embryos or adventitious buds were initiated from hypocotyl-derived calli and only adventitious buds were formed from stem-derived calli on MS added 2.69 μM NAA with 4.44 or 8.89 μM BA. Many plantlets developed from somatic embryos or organogenetic buds when they were transferred on growth hormone-free MS medium or MS with 14.78 μM IBA after 3 weeks in culture, respectively. Somatic embryos induced directly from hypocotyls are more desirable because they can produce genetically stable plantlets and this is most suitable for plant genetic transformation. This

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A Single Amino Acid Change(Asp 53→Ala53) Converts Survivin from anti-apoptotic to proapoptotic

A Single Amino Acid Change(Asp 53→Ala53) Converts Survivin ...

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中国细胞生物学学会第八届会员代表大会暨学术大会

作者： Zhiyin Song, Shixin Liu, He He, Yi Wang, Shanshan Feng and Mian Wu Department of Molecular and Cell Biology, Key Laboratory of Structural Biology, School of Life Sciences, University of Science and Technology of China, Hefei, Anhui, PR China 230027

Survivin is a member of the inhibitor of apoptosis protein (IAP) family that has been implicated in both apoptosis inhibition and cell cycle control. Recently, growing attentions have been attracted due to its tumor-specific expression and potential applications in tumor therapy. However, its inhibitory mechanism and subcellular localization remain controversial. Here, we report a novel Survivin mutant Surv-D53A, which displays opposing function and distinctive subcellular distribution compared with its wild type counterpart. Surv-D53A was shown to induce apoptosis in a p53-independent fashion, indicating that tumor suppressor p53 is not involved in its apoptosis pathway. Surv-D53A was shown to markedly sensitize apoptosis induced by TRAIL, doxorubicin and RIP3. We also demonstrated that Surv-D53A localizes in cytoplasm in interphase and to midbody at telophase similar to wild-type Survivin, however, it fails to co-localize with Aurora B to centromere in metaphase as wt-Survivin. Surv-D53A mutant is less stable than wt-Survivin and is degraded by ubiquitin-proteasome pathway more rapidly. Additionally, we found that Surv-D53A could interact with wt-Survivin to form heterodimer, which may account for the loss of anti-apoptotic function of wt-Survivin. Finally, unlike wt-Survivin, Surv-D53A is unable to bind to Smac/DIABLO, which may explain the underlying mechanism for its abolishment of anti-apoptotic activity of Survivin.

关键词： Survivin A Single Amino Acid Change Ala53 Asp 53 Converts Survivin from anti-apoptotic to proapoptotic

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Roles of tissue plasminogen activator in epidermal stratification

Roles of tissue plasminogen activator in epidermal stratific...

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中国细胞生物学学会第八届会员代表大会暨学术大会

作者：连小华杨恬 Department of Cell Biology Third Military Medical University Chongqing 400038 Dept.of Cell Biology Third Military Medical Univ

Epidermal stratification has a primary role in the epidermal barrier formation and in wound reepithelialization, and abnormal keratinocyte stratification is a consistent feature of several cutaneous disorders. In the process of keratinocyte stratification, proteolysis plays a crucial role. The tissue plasminogen activator (tPA) expressed predominantly both in the lesional epidermis and in the epidermis during wound repair, and is localized focally in the suprabasal keratinocytes, so it suggests that tPA may be related to epidermal keratinocyte stratification, but roles of tPA in regulating epidermal stratification remain unknown. In present study, using synthetic tPA and monoclonal tPA antibody, we examined the importance and mechanism of tPA in modulating of keratinocyte stratification. Histological investigation of the stratified fetal skin revealed that after continually maintained at the interface of air-liquid for 7d, the addition of tPA resulted in a significant increase in epidermal stratification, more suprabasal layers with flattened and organized orderly keratinocytes were formed, and the stratum corneum was anucleated. The immunocytochemical localization of KIO by confocal laser scanning revealed that after 7d in high Ca 2+ medium , the addition of tPA severely enhanced high Ca2+-induced stratification, the epidermis piece of the keratinocyte stratification became thicker and wider, most of stratified keratinocytes expressed KIO intensely. The infrastructure of desmosomes in cultured keratinocytes was examined by electron microscopy. When cultures were incubated with high Ca2+ plus tPA antibodies, the structure of desmosomes became loosened and tended to be disassembled, however, some neighboring keratinocytes else showed no *** results indicated that in cultures in high Ca2+ medium, intense staining for E-cadherin antigen was detected in most keratinocytes and appeared concentrating on the cell surfaces. After incubation with tP

关键词： tissue plasminogen activator epidermal stratification epidermal differentiation

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利用我国栽培黑麦白粉病新抗源实现1RS/1BL易位系抗性改良的细胞遗传学研究

利用我国栽培黑麦白粉病新抗源实现1RS/1BL易位系抗性改良的细胞...

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中国细胞生物学学会第八届会员代表大会暨学术大会

作者：张怀渝任正隆四川农业大学植物遗传育种省级重点实验室四川农业大学植物遗传育种省级重点实验室

栽培黑麦（*** L.）是重要的抗性资源,在小麦遗传改良中起着重要的作用。在目前发现的32个抗白粉病基因中,有4个Pm基因是来自于栽培黑麦,但没有一个与中国栽培黑麦有关,而且,已丧失白粉病抗性,在育种上不能有效地发挥作用。中国威岭栽... 详细信息

栽培黑麦（*** L.）是重要的抗性资源,在小麦遗传改良中起着重要的作用。在目前发现的32个抗白粉病基因中,有4个Pm基因是来自于栽培黑麦,但没有一个与中国栽培黑麦有关,而且,已丧失白粉病抗性,在育种上不能有效地发挥作用。中国威岭栽培黑麦是发现的一个新的白粉病抗源,通过远缘杂交,已被转移到普通栽培小麦。本研究对白粉病免疫的中国威岭栽培黑麦与两个高感白粉病的普通小麦（Triticum aestivum L.）My8443（含1RS/1BL 染色体易位）和A42912（不含1RS/1BL染色体易位）的远缘杂交、回交后代（BCF）群体中的单株系进行了减数分裂细胞学行为和Giemsa C-带核型分析,及GISH鉴定,结果表明:黑麦染色体导入1RS/1BL染色体小麦背景后所引起的花粉母细胞减数分裂的紊乱程度显著高于在非1RS/1BL小麦背景下产生的减数分裂异常行为,表现在黑麦染色体在不同小麦背景下,花粉母细胞减数分裂时每细胞平均交叉数,单价体频率,微核率等的不同,以及发生单价体错分裂,后期桥、断片和染色体碎裂上的差异。在非1RS/1BL背景下,检测到大量的新1RS/1BL染色体易位,而在1RS/1BL背景中也发现1R二体附加植株的自交F1代产生较高频率的小麦/黑麦的染色体易位。实验表明1R染色体的着丝点较其它黑麦染色体更易断裂,它是造成小麦-黑麦染色体臂间易位形成的重要原因;而在黑麦染色体导入1RS/1BL小麦背景之后,1R的这种易断裂特性表现更突出,同时也诱导了已存在的1RS/1BL染色体易位的不稳定性,从而导致断裂;研究结果证明,对已丧失抗病特性的1RS/1BL小麦品种（系）,导入抗病性强的中国威岭栽培黑麦的染色体,利用1R的易断裂特性和1R诱导的1RS/1BL易位染色体的不稳定性,通过染色体断裂和重接,可以构建一个不同于原有的1RS/1BL的新的1RS/1易位染色体,使丧失抗病特性的1RS/1BL小麦品种（系）重新恢复抗病特性。研究不排除新导入的1RS与原1RS/1BL上的1RS配对和交叉,实现有利基因的相互交换,赋之小麦新的抗病遗传特性的可能性,本文对此进行了深入讨论。

关键词：黑麦小麦染色体易位减数分裂 C-带 GISH 小麦白粉病

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Lily (Lilium longlflorum L.) pollen protoplast adhesion is increased in the presence of the peptide SCA (stigma/stylar cysteine-rich adhesin)

Lily (Lilium longlflorum L.) pollen protoplast adhesion is i...

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中国细胞生物学学会第八届会员代表大会暨学术大会

作者： Jean-Claude Mollet E.M.Lord Laboratoire de Glycobiologie et Physiologie Vegetale Facultie Jean Perrin Universite d'Artois 62307 Lens Cedes France Center for Plant Cell Biology Department of Botany and Plant Sciences University of California Riverside California 92521 USA

The style of lily produces a specialized extracellular matrix (ECM) in the transmitting tract epidermis that functions to guide pollen tubes to the ovary. This adhesive ECM contains low esterified pectins and a peptide SCA (stigma/stylar cysteine-rich adhesin). Together they form a matrix to which pollen tubes adhere as they grow through the style. Adhesion also occurs between the pollen tubes themselves when they grow in vivo, an event that does not occur in vitro. Pollen does not produce SCA, but when SCA is added to an in vitro growth medium, it binds to pollen tubes which have esterified and low-esterified pectins in their walls. The pollen partner in the adhesive interaction is not known. Here, we use a pollen protoplast system to examine the effect of SCA on protoplast adhesion when it is added to the growth medium. We found that SCA induces a two-fold increase in protoplast adhesion when it is added at the start of protoplast culture. This adhesion is more extensive than that seen in the control as measured by the length of the adhesion zone. This effect is less when SCA is added to the medium after the cell wall on the protoplast has begun to regenerate. Reformation of the wall on the pollen protoplast at 6 hours of culture was detected by Calcofluor white staining for glucans. Using MAbs to arabinogalactan proteins (AGPs) (JIM 13) and to pectins (JIMs 5,7), we were able to show that among the first components deposited in the new wall were AGPs (patchy labeling at 1-3 hours and continuous at 6 hours) and highly esterified pectins (patchy labeling at 3-6 hours and continuous at 24 hours). We saw no labeling for low esterified pectins even after 3 days of culture. This is interesting since it is the low esterified pectins from the style that bind SCA and form the adhesive matrix. In the pollen protoplast culture, adhesion occurs in the absence of the low esterified pectin. The newly formed wall on the protoplast mirrors that of the pollen tube tip in lily tha

关键词： adhesion, AGPs, LTP, pectins, pollen protoplasts, SCA

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Changes of uPA and uPA-R expression in the cornea after alkali burn

Changes of uPA and uPA-R expression in the cornea after alka...

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中国细胞生物学学会第八届会员代表大会暨学术大会

作者：严军杨恬李国平张艺曾益军杨进 Department of Cell Biology Third Military Medical University Chongqing 400038 P R China Dept.of Cell Biology Third Military Medical Univ Department of Cell Biology Third Military Medical University Chongqing 400038 P R China Department of Cell Biology Third Military Medical University Chongqing 400038 P R China Department of Cell Biology Third Military Medical University Chongqing 400038 P R China Department of Cell Biology Third Military Medical University Chongqing 400038 P R China

In order to investigate the expression levels of uPA and uPA-R in corneal repair after alkali burn, we first established the corneal alkali burn models in vitro and in vivo, then we performed immunocytochemistry(ICC) of uPA /uPA-R and image analysis and statistical evaluation to determine their expression levels both in vitro and in vivo. Compared with control group, the expressions of uPA and uPA-R after alkali burn had no significant increases till 6h,then increased rapidly from 12h to 24h and reached their maxima at 24h. From 24h their expression levels decreased rapidly. In vivo they rebounded again after 48 hours and attained their second peaks at 96h respectively. After that, their expressions decreased again. TheuPA-positive reaction mainly distributed in the cytoplasm while that of uPA-R mostly distributed on the cellular membrane. Their expression changes were similar to each other, both in vivo and in vitro. In vivo, uPA and uPA-R expressed and gathereid in the basal layer of corneal epithelium. These results suggest that the time phase of 24h after wound is the typical stage associating with the expressional maximum of uPA and uPA-R both in vivo and in vitro. Furthermore, the results also imply that the expression changes of uPA and uPA-R correlate to the wound healing after the corneal alkali burn, uPA and uPA-R coordinate with each other to stimulate the wound healing.

关键词： uPA uPA-R cornea alkali burn

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Plant regeneration from callus protoplasts of the forage legume Astragalus melilotoides Pall.

Plant regeneration from callus protoplasts of the forage leg...

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中国细胞生物学学会第八届会员代表大会暨学术大会

作者：侯岁稳贾敬芬兰州大学生命科学学院西北大学生命科学学院

Astragalus melilotoides Pall, is one of important perennial forage legumes. It is widely cultivated in many arid regions of north China for improving pasture and in crop rotation because of its drought-, cold- and disease-resisting, soil- and water-holding, and sand-binding characteristics. Unfortunately, its palatability is not as good as many other legumes such as alfalfa and sainfoin. Genetic manipulations, such as somatic hybridization, direct gene transfer and mutant selection may provide useful approaches for the improvement of this species. In this study, we described a reproducible procedure for the regeneration of plantlets from protoplasts isolated from hypocotyl-derived friable embryogenic calli of the forage legume Astragalus melilotoides. The induction of soft, friable and yellow-green embryogenic calli was obtained on MS medium plus lower concentration of 2,4-D (1 mg/1) and BA (0.25 mg/1) and they were critical for protoplast yield and viability. The maximum yield of viable protoplasts was obtained with an enzymatic mixture containing 2% (w/v) cellulase onozuka R10, 0.5% (w/v) cellulase onozuka RS, 0.5% (w/v) macerozyme R10, 0,5% (w/v) hemicellulase and 1% (w/v) pectinase in CPW salts with 13% (w/v) sorbitol incubated for 10 h at 25±1℃. First divisions occurred after 3-7 days of initial culture. The division frequency and plating efficiency were highest in solid-liquid double layer KM8P medium supplemented with 0.2 M glucose, 0.3 M mannitol, 500 mg/1 CH, 1 mg/1 2,4-D, 0.5 mg/1 BA and 0.2 mg/1 KIN. Protoplast-derived microcalli were obtained after 8 weeks in culture and proliferated on MS medium containing 1 mg/12,4-D and 0.25 mg/1 BA. The plantlets were regenerated from protoplast-derived calli via somatic embryogenesis (56%) and organogenesis (22%) on MS medium with 0.5 mg/1 NAA and 1-2 mg/1 BA. Rooted plantlets developed from somatic embryos or adventitious buds on MS medium without any plant growth regulators or with 3 mg/1 IBA 3 weeks later. All th

关键词： Plant regeneration from callus protoplasts of the forage legume Astragalus melilotoides Pall

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