Dlmo基因编码一个32kDa的蛋白

为了获取研究Dlmo（果蝇LMO基因的简称）的功能信息,使用耦联网织红细胞体外翻译系统将Dlmo基因进行体外转录翻译得到32kDa的蛋白产物。该蛋白产物与抗人类LMO2蛋白的多抗抗体发生免疫沉淀,并得到了32kDa的阳性带。为了证实Dlmo基因在体外和体内翻译能得到同一大小蛋白,从2～4小时果蝇胚盘中分别抽提核和胞浆提取物进行Western印迹分析,免疫血清可以识别胞浆提取物中的32kDa蛋白,而在核提取物中未曾见到,证实体外和体内翻译产物相同。免疫组织化学的分析在0～4小时胚盘外周胞浆中有Dlmo基因的阳性染色信号,结果证实,Dlmo与人类LMO不同,其表达产物是一个胞浆蛋白。 Abstract：In order to gain some insight into a possible function of Dlmo gene,we used the TNT coupled reticulocyte lysate systems as an in vitro translation system to detect the Drosoplila protein. We found a 32kDa protein product.To demonstrate that the DLMO protein has the same size in vivo as in vitro we studied nuclear and cytoplasmic extracts from 2-4h embryos in Western blots. The immuno serum recognizes a 32kDa protein in the cytoplasm that is not present in the nucleus.The products were immune precipitated with the polyclonal anti-LMO2 antibody raised against the human protein and seen a positive band of 32kDa.Using immuno histochemical analysis was seen a positive staining in the basal cytoplasm of blastoderm embryos at 4 hours. This result confirms that the DLMO is a cytoplasmprotein, not like human LMO protein.     

Wild type and <Emphasis Type="Italic">H43Y</Emphasis> variant of human <Emphasis Type="Italic">TRIM5α</Emphasis> show similar anti-human immunodeficiency virus type 1 activity both in vivo and in vitro

Polymorphisms in human genes have been shown to affect the rate of disease progression to acquired immune deficiency syndrome in human immunodeficiency virus type 1 (HIV-1)-infected individuals. Recently, tripartite motif 5α (TRIM5α) was identified as a factor that confers resistance to HIV-1 infection in Old World monkey cells. Subsequently, Sawyer et al. (Curr Biol 16:95–100, 2006) reported a single nucleotide polymorphism (H43Y) in the human TRIM5α gene and TRIM5α protein with 43Y was found to lose its ability to restrict HIV-1. In the present study, we reevaluated effects of this allele on in vitro anti-HIV-1 activity as well as on HIV-1 disease progression in European and Asian cohorts of HIV-1-infected individuals. Our epidemiological and molecular biological findings clearly indicate H43Y has a very minor effect on anti-HIV-1 activity of TRIM5α, suggesting that this allele is immaterial, at least in HIV-1-infected Europeans and Asians.     

High-throughput fluorescent tagging of full-length Arabidopsis gene products in planta

We developed a high-throughput methodology, termed fluorescent tagging of full-length proteins (FTFLP), to analyze expression patterns and subcellular localization of Arabidopsis gene products in planta. Determination of these parameters is a logical first step in functional characterization of the approximately one-third of all known Arabidopsis genes that encode novel proteins of unknown function. Our FTFLP-based approach offers two significant advantages: first, it produces internally-tagged full-length proteins that are likely to exhibit native intracellular localization, and second, it yields information about the tissue specificity of gene expression by the use of native promoters. To demonstrate how FTFLP may be used for characterization of the Arabidopsis proteome, we tagged a series of known proteins with diverse subcellular targeting patterns as well as several proteins with unknown function and unassigned subcellular localization.     

Attributes of short linear motifs

Traditionally, protein-protein interactions were thought to be mediated by large, structured domains. However, it has become clear that the interactome comprises a wide range of binding interfaces with varying degrees of flexibility, ranging from rigid globular domains to disordered regions that natively lack structure. Enrichment for disorder in highly connected hub proteins and its correlation with organism complexity hint at the functional importance of disordered regions. Nevertheless, they have not yet been extensively characterised. Shifting the attention from globular domains to disordered regions of the proteome might bring us closer to elucidating the dense and complex connectivity of the interactome. An important class of disordered interfaces are the compact mono-partite, short linear motifs (SLiMs, or eukaryotic linear motifs (ELMs)). They are evolutionarily plastic and interact with relatively low affinity due to the limited number of residues that make direct contact with the binding partner. These features confer to SLiMs the ability to evolve convergently and mediate transient interactions, which is imperative to network evolution and to maintain robust cell signalling, respectively. The ability to discriminate biologically relevant SLiMs by means of different attributes will improve our understanding of the complexity of the interactome and aid development of bioinformatics tools for motif discovery. In this paper, the curated instances currently available in the Eukaryotic Linear Motif (ELM) database are analysed to provide a clear overview of the defining attributes of SLiMs. These analyses suggest that functional SLiMs have higher levels of conservation than their surrounding residues, frequently evolve convergently, preferentially occur in disordered regions and often form a secondary structure when bound to their interaction partner. These results advocate searching for small groupings of residues in disordered regions with higher relative conservation and a propensity to form the secondary structure. Finally, the most interesting conclusions are examined in regard to their functional consequences.     

Polychaete diversity in the Scotia Arc benthic realm: Are polychaetes tracers for faunal exchange?

The Scotia Arc is the only shallow-water and island bridge linking nowadays Patagonia and the Antarctic. The Antarctic Circumpolar Current as an oceanographic peculiarity makes this region an interesting biogeographic transition zone, because this frontal system traditionally is said to isolate the Antarctic fauna from that of the adjacent northern ecosystems. Based on benthos samples from three expeditions onboard R/V Polarstern, we studied distribution patterns of 200 polychaete species and 34 major benthic taxa in order to evaluate the role of polychaetes in the benthic realm of this part of the Southern Ocean. ANOSIM test distinguished three station groups: the central eastern Scotia Sea, the continental shelf off South America and stations at the tip of the Antarctic Peninsula. These station groups differed in organism densities and diversities with stations at the tip of the Antarctic Peninsula hosting the most diverse and dense community. The polychaete diversity patterns in the three assemblages evidenced closer connectivity between the tip of the Antarctic Peninsula and the central eastern Scotia Sea than between the continental shelf off South America with either the stations off the tip of the Peninsula or the central eastern Scotia Sea. This is probably supported by the Polar Front, which divides the island chain into two branches. Species distribution and community patterns of polychaetes appear to be associated with oceanographic and sediment conditions in this region. Most of the shared species showed the capability to tolerate differences in hydrostatic pressure. We suggest that the islands of the Scotia Sea may constitute a bridge for exchange of benthic species, particularly for polychaetes with eurybathic distribution and high dispersal capabilities.     

Identification and re-addressing of a transcriptionally permissive locus in the porcine genome

Recently, we established the Sleeping Beauty transposon system for germ line competent transgenesis in the pig. Here, we extend this approach to re-target a transposon-tagged locus for a site-specific gene knock-in, and generated a syngeneic cohort of piglets carrying either the original transposon or the re-targeted event. A Cre-loxP-mediated cassette exchange of the tagging transposon with a different reporter gene was performed, followed by flow cytometric sorting and somatic cell nuclear transfer of recombined cells. In parallel, the original cells were employed in somatic cell nuclear transfer to generate clone siblings, thereby resulting in a clone cohort of piglets carrying different reporter transposons at an identical chromosomal location. Importantly, this strategy supersedes the need for an antibiotic selection marker. This approach expands the arsenal of genome engineering technologies in domestic animals, and will facilitate the development of large animal models for human diseases. Potentially, the syngeneic cohort of pigs will be instrumental for vital tracking of transplanted cells in pre-clinical assessments of novel cell therapies.     

Mesenchymal stem cells show radioresistance in vivo

Irradiation impacts on the viability and differentiation capacity of tissue‐borne mesenchymal stem cells (MSC), which play a pivotal role in bone regeneration. As a consequence of radiotherapy, bones may develop osteoradionecrosis. When irradiating human bone‐derived MSC in vitro with increasing doses, the cells’ self‐renewal capabilities were greatly reduced. Mitotically stalled cells were still capable of differentiating into osteoblasts and pre‐adipocytes. As a large animal model comparable to the clinical situation, pig mandibles were subjected to fractionized radiation of 2 χ 9 Gy within 1 week. This treatment mimics that of a standardized clinical treatment regimen of head and neck cancer patients irradiated 30 χ 2 Gy. In the pig model, fractures which had been irradiated, showed delayed osseous healing. When isolating MSC at different time points post‐irradiation, no significant changes regarding proliferation capacity and osteogenic differentiation potential became apparent. Therefore, pig mandibles were irradiated with a single dose of either 9 or 18 Gy in vivo, and MSC were isolated immediately afterwards. No significant differences between the untreated and 9 Gy irradiated bone with respect to proliferation and osteogenic differentiation were unveiled. Yet, cells isolated from 18 Gy irradiated specimens exhibited a reduced osteogenic differentiation capacity, and during the first 2 weeks proliferation rates were greatly diminished. Thereafter, cells recovered and showed normal proliferation behaviour. These findings imply that MSC can effectively cope with irradiation up to high doses in vivo. This finding should thus be implemented in future therapeutic concepts to protect regenerating tissue from radiation consequences.     

Mapping of resistance to the potato cyst nematode Globodera rostochiensis from the wild potato species Solanum vernei

A population of diploid potato (Solanum tuberosum) was used for the genetic analysis and mapping of a locus for resistance to the potato cyst nematode Globodera rostochiensis, introgressed from the wild potato species Solanum vernei. Resistance tests of 108 genotypes of a F₁ population revealed the presence of a single locus with a dominant allele for resistance to G. rostochiensis pathotype Ro1. This locus, designated GroV1, was located on chromosome 5 with RFLP markers. Fine-mapping was performed with RAPD and SCAR markers. The GroV1 locus was found in the same region of the potato genome as the S. tuberosum ssp. andigena H1 nematode resistance locus. Both resistance loci could not excluded to be allelic. The identification of markers flanking the GroV1 locus offers a valuable strategy for marker-assisted selection for introgression of this nematode resistance.Abbreviations BSA bulked segregant analysis - RAPD random-amplified polymorphic DNA - RFLP restriction fragment length polymorphism - SCAR sequence-characterized amplified region