Adaptation to different growth temperatures modifies some mammalian cell survival responses

Chinese hamster (HA-1) cells that have been grown at 37 °C since explant several years ago can adapt themselves to grow at temperatures ranging from 32 to 41 °C. This growth adaptation is accompanied by major phenotypic changes in, for exampie, the cellular responses to 43 and 45 °C heat challenges and to ethanol challenges (0–10% in concentration). Cells grown at 39.5 °C are seen to acquire substantial heat resistance when compared with cells grown at 37 °C; resistance is even more pronounced if the growth temperature is at 41 °C. On the other hand, cells grown at 32 °C become more sensitive to heat than controls. Our results also indicate an increased resistance to ethanol of the 41 °C grown cells. By contrast the cells' X-ray survival response is affected only minimally. The changes seen are phenotypic; upon being returned to 37 °C, HA-1 cells within 34 h regain their ‘normal’ heat responses.     

Localization of the nucleolar organizer by computer-aided analysis of a variant no. 21 in a human isolate

Summary A variant chromosome no. 21 consisting of two stalks and two satellites in tandem was detected during a survey of a human isolate. The variant segregated in three generations of a large kindred. One male had the variant no. 21, a metacentric Y, and a 47, XXY complement; however, no other evidence of chromosomal nondisjunction was found. Computer-aided analysis of sequentially stained variant no. 21 chromosomes indicated that silver-stained material corresponded to the proximal stalk region (as defined defined by Giemsa). These data support the hypothesis that human nucleolar organizers are localized to the stalks of acrocentric chromosomes.     

Lung retinol storing cells synthesize and secrete retinoic acid,an inducer of alveolus formation

Retinoids play a key role in the formation of pulmonary alveoli. Lipid interstitial cells (LICs) of the alveolar wall store retinol and are concentrated at sites of alveolus formation, suggesting they are an endogenous source of retinoids for alveolus formation. We show in cultured rat lung cells that LICs synthesize and secrete all-trans retinoic acid (ATRA); its secretion is halved by dexamethasone, an inhibitor of alveolus formation. In a second alveolar wall cell, the pulmonary microvascular endothelial cell (PMVC), ATRA increases expression of the mRNA of cellular retinol binding protein-I (CRBP-I), a protein involved in ATRA synthesis. Serum-free, exogenous ATRA-free medium conditioned by LICs rich in retinol storage granules caused a 10-fold greater increase of CRBP-I mRNA in PMVCs than media conditioned by LICs with few retinol storage granules. This action of medium conditioned by retinol storage granule-rich LICs is decreased by a retinoic acid receptor pan-antagonist and by a retinoid X receptor pan-antagonist, suggesting the responsible molecule(s) is a retinoid and that retinoid signaling occurs in a paracrine fashion.     

Mapping eIF5A binding sites for Dys1 and Lia1: in vivo evidence for regulation of eIF5A hypusination

The evolutionarily conserved factor eIF5A is the only protein known to undergo hypusination, a unique posttranslational modification triggered by deoxyhypusine synthase (Dys1). Although eIF5A is essential for cell viability, the function of this putative translation initiation factor is still obscure. To identify eIF5A-binding proteins that could clarify its function, we screened a two-hybrid library and identified two eIF-5A partners in S. cerevisiae: Dys1 and the protein encoded by the gene YJR070C, named Lia1 (Ligand of eIF5A). The interactions were confirmed by GST pulldown. Mapping binding sites for these proteins revealed that both eIF5A domains can bind to Dys1, whereas the C-terminal domain is sufficient to bind Lia1. We demonstrate for the first time in vivo that the N-terminal alpha-helix of Dys1 can modulate enzyme activity by inhibiting eIF5A interaction. We suggest that this inhibition be abrogated in the cell when hypusinated and functional eIF5A is required.     

Effects of Plectin Depletion on Keratin Network Dynamics and Organization

The keratin intermediate filament cytoskeleton protects epithelial cells against various types of stress and is involved in fundamental cellular processes such as signaling, differentiation and organelle trafficking. These functions rely on the cell type-specific arrangement and plasticity of the keratin system. It has been suggested that these properties are regulated by a complex cycle of assembly and disassembly. The exact mechanisms responsible for the underlying molecular processes, however, have not been clarified. Accumulating evidence implicates the cytolinker plectin in various aspects of the keratin cycle, i.e., by acting as a stabilizing anchor at hemidesmosomal adhesion sites and the nucleus, by affecting keratin bundling and branching and by linkage of keratins to actin filament and microtubule dynamics. In the present study we tested these hypotheses. To this end, plectin was downregulated by shRNA in vulvar carcinoma-derived A431 cells. As expected, integrin β4- and BPAG-1-positive hemidesmosomal structures were strongly reduced and cytosolic actin stress fibers were increased. In addition, integrins α3 and β1 were reduced. The experiments furthermore showed that loss of plectin led to a reduction in keratin filament branch length but did not alter overall mechanical properties as assessed by indentation analyses using atomic force microscopy and by displacement analyses of cytoplasmic superparamagnetic beads using magnetic tweezers. An increase in keratin movement was observed in plectin-depleted cells as was the case in control cells lacking hemidesmosome-like structures. Yet, keratin turnover was not significantly affected. We conclude that plectin alone is not needed for keratin assembly and disassembly and that other mechanisms exist to guarantee proper keratin cycling under steady state conditions in cultured single cells.     

香港的生物多样性及其保育工作

香港位于热带,属海洋性气候。地势崎岖多山,山地约占全港总面积的3/4。城市发展多集中在沿海平坦地带。目前香港的城市和乡镇面积约占总面积的20%,农地约占5%(当中大部份已遭荒废),余下的均为郊野地区,这包括天然林和人工林(约占14%)、灌丛(约占36%)及草地(约占17%)。由于良好的气候和地理条件,形成了众多不同的生态环境,使总面积仅1090 km²的弹丸之地孕育出种类多样的动植物,生物多样性十分丰富。香港约有2500种原生植物,包括被子植物约1900种,裸子植物7种,蕨类植物220多种及苔藓植物300多种。动物方面,已记录的野生哺乳类动物有40多种,鸟类超过459种,两栖类23种,爬行类70多种。昆虫种类繁多,其中蜻蜓目100多种,鳞翅目2200多种(蝴蝶200多种,蛾类2000多种)。有很多是国家保护物种和特有种。植物方面属国家一级保护的有1种——刺桫椤(Alsophila spinulosa);国家二级保护的有6种,如四药门花(Tetrathryrium subcordatum);国家三级保护的有8种,如穗花杉(Amentotaxus argotaenia)。此外,香港特有种有16种,例如紫萁科(Osmundaceae)的粤紫萁(Osmunda mildei)、马兜铃科(Aristolochiaceae)的香港细辛(Asarum hongkongense)和兰科(Orchidaceae)的谢氏卷瓣兰(Bulbophyllum tseanum)。动物方面有9种属国家一级保护,例如中华白海豚(Sousa chinensis);79种属国家二级保护。特有种则有卢氏小树蛙(Philautus romeri)、包氏双足蜥(Dibamus bogadeki)及多种昆虫。为了保护丰富的野生动植物及其栖息的环境,香港特别行政区政府制定了一些法例并推行了不少保护措施,例如设立了21个郊野公园和14个特别地区,占全港陆地总面积约38%。此外,还成立了2个禁区、3个海岸公园和1个海洋保护区。另一方面,政府还设立了59个“具特殊科学价值地点”,以保护及研究各种动植物、生态系统和特殊的地质地貌。香港地少人多,总人口超过600万,是世界上人口最稠密的地区之一。多年来香港这个生物宝库不断地遭受人类活动的威胁,近年来由于人口急剧上升,对土地需求迫切,不少郊野地区被开发利用,环境污染亦日益严重。此外,一些野生植物因具有药用价值、观赏价值或其他用途而遭盗伐或采集。上述种种因素已使香港野生动植物及其生境受到严重损害,一些物种更濒于灭绝,进行生物多样性的保育工作刻不容缓。因此,香港确实需要制定整体的生物多样性保护策略。有鉴于此,香港大学生态及分类学系于1996年展开了一项为期3年的香港生物多样性调查,以增加对动植物资源现况的了解,为保护香港的珍稀濒危物种和日益恶化的自然环境提出补救方案,并为制订长远的保育策略奠定基础。     

Bile acids,FGF15/19 and liver regeneration: From mechanisms to clinical applications

The liver has an extraordinary regenerative capacity rapidly triggered upon injury or resection. This response is intrinsically adjusted in its initiation and termination, a property termed the “hepatostat”. Several molecules have been involved in liver regeneration, and among them bile acids may play a central role. Intrahepatic levels of bile acids rapidly increase after resection. Through the activation of farnesoid X receptor (FXR), bile acids regulate their hepatic metabolism and also promote hepatocellular proliferation. FXR is also expressed in enterocytes, where bile acids stimulate the expression of fibroblast growth factor 15/19 (FGF15/19), which is released to the portal blood. Through the activation of FGFR4 on hepatocytes FGF15/19 regulates bile acids synthesis and finely tunes liver regeneration as part of the “hepatostat”. Here we review the experimental evidences supporting the relevance of the FXR-FGF15/19-FGFR4 axis in liver regeneration and discuss potential therapeutic applications of FGF15/19 in the prevention of liver failure. This article is part of a Special Issue entitled: Cholangiocytes in Health and Disease edited by Jesus Banales, Marco Marzioni, Nicholas LaRusso and Peter Jansen.     

Prediction of protein loop geometries in solution

The ability to determine the structure of a protein in solution is a critical tool for structural biology, as proteins in their native state are found in aqueous environments. Using a physical chemistry based prediction protocol, we demonstrate the ability to reproduce protein loop geometries in experimentally derived solution structures. Predictions were run on loops drawn from (1)NMR entries in the Protein Databank (PDB), and from (2) the RECOORD database in which NMR entries from the PDB have been standardized and re-refined in explicit solvent. The predicted structures are validated by comparison with experimental distance restraints, a test of structural quality as defined by the WHAT IF structure validation program, root mean square deviation (RMSD) of the predicted loops to the original structural models, and comparison of precision of the original and predicted ensembles. Results show that for the RECOORD ensembles, the predicted loops are consistent with an average of 95%, 91%, and 87% of experimental restraints for the short, medium and long loops respectively. Prediction accuracy is strongly affected by the quality of the original models, with increases in the percentage of experimental restraints violated of 2% for the short loops, and 9% for both the medium and long loops in the PDB derived ensembles. We anticipate the application of our protocol to theoretical modeling of protein structures, such as fold recognition methods; as well as to experimental determination of protein structures, or segments, for which only sparse NMR restraint data is available.