Expression of a novel HsMCAK mRNA splice variant,tsMCAK gene,in human testis

Identification of specifically expressed genes in the adult or fetal testis is very important for the study of genes related to the development and function of the testis. In this study, a human adult testis cDNA microarray was constructed and hybridized with 33P-labeled human adult and embryo testis cDNA probes, respectively. After differential display analyzing, a number of new genes related to the development of testis and spermatogenesis had been identified. One of these new genes is tsMCAK. tsMCAK was expressed 2.62 folds more in human adult testis than fetal testis. The full length of tsMCAK is 2401 bp and contains a 2013 bp open reading frame, encoding a 671-amino-acid protein. Sequence analysis showed that it has a central kinesin motor domain and is homologous to HsMCAK gene of the somatic cells. Blasting human genome database localized tsMCAK to human chromosome 1P34 and further investigation showed that it is a splice variant of HsMCAK. The tissue distribution of tsMCAK was determined by RT-PCR and it is expressed highly and specifically in the testis. Southern blot studies of its expression in patients with infertility indicated its specific expression in spermatogenic cells and its correlation with male infertility. The above results suggested that tsMCAK is a candidate gene for the testis-specific KRPs and its specific expression in the testis was correlated with spermatogenesis and may be correlated with male infertility.     

Atomic force microscopic measurement of the interdomain angle in symmetric Holliday junctions

The Holliday junction is a key intermediate in genetic recombination. It consists of four DNA strands that associate by base pairing to produce four double helices flanking a junction point. In the presence of multivalent cations, the four helices, in turn, stack in pairs to form two double-helical domains. The angle between these domains has been shown in a number of solution studies to be approximately 60 degrees in junctions flanked by asymmetric sequences. However, the recently determined crystal structure of a symmetric junction [Eichman, B. F., Vargason, J. M., Mooers, B. H. M., and Ho, P. S. (2000) Proc. Natl. Acad. Sci. U.S.A. 97, 3971-3976] finds an angle closer to 40 degrees, possibly because of sequence effects. From the crystal structure alone, one cannot exclude the possibility that this unusual angle is a consequence of crystal packing effects. We have formed two-dimensional (2D) periodic arrays of DNA parallelograms with the same junction-flanking sequence used to produce the crystals; these parallelograms are free to adopt their preferred interdomain angle. Atomic force microscopy can be used to establish the interdomain angle in this system. We find that the angle in this junction is 43 degrees, in good agreement with the results of crystallography. We have used hydroxyl radical autofootprinting to establish that the branch point is at the same migratory position seen in the crystals.     

Crystal structure of the cytoskeleton-associated protein glycine-rich (CAP-Gly) domain

Cytoskeleton-associated proteins (CAPs) are involved in the organization of microtubules and transportation of vesicles and organelles along the cytoskeletal network. A conserved motif, CAP-Gly, has been identified in a number of CAPs, including CLIP-170 and dynactins. The crystal structure of the CAP-Gly domain of Caenorhabditis elegans F53F4.3 protein, solved by single wavelength sulfur-anomalous phasing, revealed a novel protein fold containing three beta-sheets. The most conserved sequence, GKNDG, is located in two consecutive sharp turns on the surface, forming the entrance to a groove. Residues in the groove are highly conserved as measured from the information content of the aligned sequences. The C-terminal tail of another molecule in the crystal is bound in this groove.     

GDNF acutely potentiates Ca2+ channels and excitatory synaptic transmission in midbrain dopaminergic neurons

Glial cell line-derived neurotrophic factor (GDNF) is best known for its long-term survival effect on dopaminergic neurons in the ventral midbrain. A recent study showed that acute application of GDNF to these neurons suppresses A-type potassium channels and potentiates neuronal excitability. Here we have characterized the acute effects of GDNF on Ca(2+) channels and synaptic transmission. GDNF rapidly and reversibly potentiated the high voltage-activated (HVA) Ca(2+) channel currents in cultured dopaminergic neurons. Analyses of channel kinetics indicate that GDNF decreased the activation time constant, increased the inactivation and deactivation time constants of HVA Ca(2+) channel currents. Ca(2+) imaging experiments demonstrate that GDNF facilitated Ca(2+) influx induced by membrane depolarization. To investigate the physiological consequences of the Ca(2+) channel modulation, we examined the acute effects of GDNF on excitatory synaptic transmission at synapses made by these dopaminergic neurons, which co-release the transmitter glutamate. Within 3 min of application, GDNF increased the amplitude of spontaneous and evoked excitatory autaptic- or multiple-postsynaptic currents. The frequency as well as the amplitude of miniature excitatory postsynaptic currents was also increased. These results reveal, for the first time, an acute effect of GDNF on synaptic transmission and its potential mechanisms, and suggest that an important function of GDNF for midbrain dopaminergic neurons is the acute modulation of transmission and ion channels.     

Humanin peptides block calcium influx of rat hippocampal neurons by altering fibrogenesis of Abeta(1-40)

Humanin peptides (including HN, HNG and other mutants) were reported previously that antagonize neurotoxicity caused by various familial Alzheimer's disease (FAD) genes and Abeta derivatives. Herein, we describe the aggregation dynamics and the representative morphological characteristics of Abeta(1-40) after different time of addition humanin peptides, which revealed that (a) the interactions of both HN and HNG with Abeta(1-40) induced quick and significant increase of light-scattering intensity, and (b) HNG also caused obvious morphological alteration from fibrillary to amorphous. In the meantime, the experiments also revealed that the interaction of HNG with Abeta(1-40) could decrease Abeta(1-40)-induced calcium rise, an initial event accompanying Abeta(1-40)-induced apoptosis of cultured neurons. Our results indicate that HNG can protect neurons by altering Abeta(1-40) morphology.     

About the effect of the amplitude of fluctuations on the coefficient of rubbing of the brownian oscillator in aqueous media

The methods of molecular dynamics and correlation functions of coordinates were applied to investigate molecular oscillations in water solution at a variation of the amplitude of fluctuations. It was established, that the amplitude of fluctuation exceeds the threshold value (in water of the order of 0.3-0.45 A), of the force of friction sharply increases, and the vibration mode changes to the mode of limited diffusion. The consistency of the results with the theory of M?ssbauer effect and the model of Brownian oscillator for conformational mobility in biopolymers is discussed.     

Probing of the spliceosome with site-specifically derivatized 5' splice site RNA oligonucleotides.

We have developed a site-specific chemical modification technique to incorporate a photoreactive azidophenacyl (APA) group at designated internal positions along the RNA phosphodiester backbone. Using this technique, we have analyzed interactions of the 5' splice site (5'SS) RNA within the spliceosome. Several crosslinked products can be detected within complex B using the derivatized 5'SS RNAs, including U6 snRNA, hPrp8p, and 114-, 90-, 70-, 54-, and 27-kDa proteins. The 5'SS RNAs derivatized at intron positions +4 to +8 crosslink to U6 snRNA, confirming the previously reported pairing interaction between these sequences. hPrp8p and p70 are crosslinked to the 5'SS RNA when the APA is placed within the 5' exon. Finally, a set of unidentified proteins, including p114, p54, and p27, is detected with the 5'SS RNA derivatized at intron positions +4 to +8. Introduction of the bulky APA group near the 5'SS junction (positions -2 to +3) strongly interferes with complex B formation and thus no APA crosslinks are observed at these positions. Together with our earlier observation that hPrp8p crosslinks to the GU dinucleotide at the 5' end of the intron, these results suggest that the inhibitory effect of APA results from steric hindrance of the hPrp8p:5'SS interaction. Unexpectedly, thio-modifications within the region of the 5'SS RNA that is involved in base pairing to U6 snRNA strongly stimulate complex B formation.     

应用离子注入技术对家蚕形态学及遗传学进行的初步研究(英文)

离子注入技术是将某种元素的原子进行电离,并使其在电场中加速,在获得较高的速度后射入固体材料表面。在离子注入过程中,被电离的离子在电场作用下加速运动,离子靠着本身获得的动能进入基体表面,在表层中运动的离子与基体原子作用损失能量后在一定的位置停留下来。该技术自６０年代问世以来,主要用于材料改性等方面。８０年代中期,我国学者开始将其用于农作物育种方面的研究,大大拓宽了离子注入技术的应用领域。所用实验材料的基因及表现型见Ｔａｂ３,我们将氢离子（Ｅ＝３５ＭｅＶ）注入处于胚胎发育后期的家蚕卵内（Ｔａｂ１）,观察其对家蚕形态及遗传方面的影响,结果表明：（１）在家蚕胚胎发育的已４期注入氢离子,其半致死剂量ＬＤ５０为１ｘ１０１０～１ｘ１０１１ｃｍ２这一区间之内;当剂量达到１ｘ１０１２ｃｍ２时,已全部致死（Ｆｉｇ．１＆Ｔａｂ．２）;（２）注入氢离子能够使家蚕在第１腹节上产生褐斑（Ｆｉｇ．２）的频率增高。并首次观察到因注入氢离子而导致家蚕出现非成对的褐斑（Ｆｉｇ．３＆Ｔａｂ．４）。（３）在氢离子注入剂量为１ｘ１０１０ｃｍ２时,能够诱变产生大量的嵌合体家蚕,并且诱变频率高达３８．５％（Ｆｉｇ．４＆Ｔａｂ．５）,这样高的     

Exosome and Exosomal MicroRNA:Trafficking,Sorting, and Function

Exosomes are 40–100 nm nano-sized vesicles that are released from many cell types into the extracellular space. Such vesicles are widely distributed in various body fluids. Recently,m RNAs and micro RNAs(mi RNAs) have been identified in exosomes, which can be taken up by neighboring or distant cells and subsequently modulate recipient cells. This suggests an active sorting mechanism of exosomal mi RNAs, since the mi RNA profiles of exosomes may differ from those of the parent cells. Exosomal mi RNAs play an important role in disease progression, and can stimulate angiogenesis and facilitate metastasis in cancers. In this review, we will introduce the origin and the trafficking of exosomes between cells, display current research on the sorting mechanism of exosomal mi RNAs, and briefly describe how exosomes and their mi RNAs function in recipient cells.Finally, we will discuss the potential applications of these mi RNA-containing vesicles in clinical settings.