Xist-mediated chromatin changes that establish silencing of an entire X chromosome in mammals

X chromosome inactivation (XCI) ensures an equal gene dosage between the sexes in placental mammals. Xist, a modular multi-domain X-encoded long non-coding RNA coats the X chromosome in cis during XCI. Xist recruits chromatin remodelers and repressor complexes ensuring silencing of the inactive X (Xi). Here, we review the recent work focused on the role of Xist functional repeats and interacting RNA-binding factors in the establishment of the silent state. Xist orchestrates recruitment of remodelers and repressors that first facilitate removal of the active chromatin landscape and subsequently direct the transition into a repressive heterochromatic environment. Some of these factors affect silencing on a chromosome-wide scale, while others display gene-specific silencing defects. The temporal order of recruitment shows each silencing step is party dependent on one another. After the Xi is established, many of the factors are dispensable, and a different repertoire of proteins ensure the silenced Xi is maintained and propagated.     

A possible mechanism of processive nucleotide and repeat additions by the telomerase

Telomerase is a specialized cellular ribonucleoprotein complex that can synthesize long stretches of a DNA primer by using an intrinsic RNA template sequence. This requires that the telomerase must be able to carry out both nucleotide and repeat additions. Here, based on available structures and experimental data, a model is presented to describe these two addition activities. In the model, the forward movement of the polymerase active site along the template during the processive nucleotide addition is rectified through the incorporation of a matched base, via the Brownian ratchet mechanism. The unpairing of the DNA:RNA hybrid and then repositioning of product 3′-end after each round of repeat synthesis, which are prerequisites for the processive repeat addition, are caused by a force acting on the primer. The force results from the conformational transition of the stem III pseudoknot, which is mechanically induced by the rotation of TERT fingers together with stem IV loop towards the polymerase active site upon a nucleotide binding. Based on the model, the dynamics of processive nucleotide and repeat additions by recombinant Tetrahymena telomerase is studied analytically, which gives good quantitative explanations to the previous experimental results. Moreover, some predicted results are presented. In particular, it is shown that the repeat addition processivity is mainly determined by the difference between the free-energy change required to disrupt the DNA:RNA hybrid and that required to unfold the stem III pseudoknot. A large difference in free energy corresponds to a low repeat addition processivity while a small difference in free energy corresponds to a high repeat addition processivity.     

Classification and nomenclature of endogenous retroviral sequences (ERVs): Problems and recommendations

The genomes of many species are crowded with repetitive mobile sequences. In the case of endogenous retroviruses (ERVs) there is, for various reasons, considerable confusion regarding names assigned to families/groups of ERVs as well as individual ERV loci. Human ERVs have been studied in greater detail, and naming of HERVs in the scientific literature is somewhat confusing not just to the outsider. Without guidelines, confusion for ERVs in other species will also probably increase if those ERVs are studied in greater detail. Based on previous experience, this review highlights some of the problems when naming and classifying ERVs, and provides some guidance for detecting and characterizing ERV sequences. Because of the close relationship between ERVs and exogenous retroviruses (XRVs) it is reasonable to reconcile their classification with that of XRVs. We here argue that classification should be based on a combination of similarity, structural features, (inferred) function, and previous nomenclature. Because the RepBase system is widely employed in genome annotation, RepBase designations should be considered in further taxonomic efforts. To lay a foundation for a phylogenetically based taxonomy, further analyses of ERVs in many hosts are needed. A dedicated, permanent, international consortium would best be suited to integrate and communicate our current and future knowledge on repetitive, mobile elements in general to the scientific community.     

Outbreak of Abortions and Infertility in Thoroughbred Mares Associated with Waterborne <Emphasis Type="Italic">Aeromonas hydrophila</Emphasis>

At a thoroughbred equine breeding farm near Hissar (Haryana), three mares aborted in their seventh month of pregnancy. The vaginal swabs of all aborted mares, and stomach contents, heart blood, liver, spleen and placenta of aborted fetuses yielded pure culture of Aeromonas hydrophila. In addition, A. hydrophila was also isolated from the vaginal swabs of three repeat breeding mares and faecal sample of a diarrheic foal. The source of infection was possibly water supply as all the water samples collected from taps, mother tank and storage tank were found to be positive for A. hydrophila. The antibiogram of all the isolates was similar showing resistance to ampicillin, carbenicillin, gentamicin, kanamycin and amikacin but sensitive to chloramphenicol, ciprofloxacin, cefuroxime, ceftriaxone, cotrimoxazole, cotrimazine, nitrofurantoin, streptomycin and tetracycline. All the 20 sera samples collected from three aborted and three repeat breeding, and eight in-contact mares, a diarrheic foal, three cows and two male buffaloes maintained at the same farm contained antibodies against A. hydrophila with titres ranging from 80 to 640. The water supply was instantly chlorinated using 0.05% sodium hypochlorite for three consecutive days and all the culturally positive mares were treated with intravaginal administration of 1 g ciprofloxacin, while the foal was given nitrofurantoin for three days. After one month, A. hydrophila could not be isolated either from mares or from their environment and antibody titre in all the seropositive animals showed a declining trend. Later, all the aborted and repeat breeding mares were confirmed to be pregnant. Thus, the present study indicated that water-borne A. hydrophila might be associated with equine abortions and infertility, and diarrhea in newborn foals.     

Molecular stiffness of selectins

During inflammation, selectin-ligand interactions provide forces for circulating leukocytes to adhere to vascular surfaces, which stretch the interacting molecules, suggesting that mechanical properties may be pertinent to their biological function. From mechanical measurements with atomic force microscopy, we analyzed the molecular characteristics of selectins complexed with ligands and antibodies. Respective stiffness of L-, E-, and P-selectins (4.2, 1.4, and 0.85 piconewton/nm) correlated inversely with the number (2, 6, and 9) of consensus repeats in the selectin structures that acted as springs in series to dominate their compliance. After reconstitution into a lipid bilayer, purified membrane P-selectin remained a dimer, capable of forming dimeric bonds with P-selectin glycoprotein ligand (PSGL)-1, endoglycan-Ig, and a dimeric form of a glycosulfopeptide modeled after the N terminus of PSGL-1. By comparison, purified membrane L- and E-selectin formed only monomeric bonds under identical conditions. Ligands and antibodies were much less stretchable than selectins. The length of endoglycan-Ig was found to be 51 ± 12 nm. These results provide a comprehensive characterization of the molecular stiffness of selectins and illustrate how mechanical measurements can be utilized for molecular analysis, e.g. evaluating the multimericity of selectins and determining the molecular length of endoglycan.     

Identification and characterization of the integrin alpha2beta1 binding motif in chondroadherin mediating cell attachment

Chondroadherin is a leucine-rich repeat protein known to mediate adhesion of isolated cells via the integrin α(2)β(1) and to interact with collagen. In this work, we show that cell adhesion to chondroadherin leads to activation of MAPKs but does not result in cell spreading and division. This is in contrast to the spreading and dividing of cells grown on collagen, although the binding is mediated via the same α(2)β(1) receptor. We identified a cell binding motif, CQLRGLRRWLEAK(318) by mass spectrometry after protease digestion of chondroadherin. Cells adhering to the synthetic peptide CQLRGLRRWLEAK(318) remained round, as was observed when they bound to the intact protein. The peptide added in solution was able to inhibit cell adhesion to the intact protein in a dose-dependent manner and was also verified to bind to the α(2)β(1) integrin. A cyclic peptide, CQLRGLRRWLEAKASRPDATC(326), mimicking the structural constraints of this sequence in the intact protein, showed similar efficiency in inhibiting binding to chondroadherin. The unique peptide motif responsible for cellular binding is primarily located in the octamer sequence LRRWLEAK(318). Binding of cells to the active peptide or to chondroadherin immobilized on cell culture plates rapidly induces intracellular signaling (i.e. ERK phosphorylation). Thus, chondroadherin interaction with cells may be central for maintaining the adult chondrocyte phenotype and cartilage homeostasis. The peptides, particularly the more stable cyclic peptide, open new opportunities to modulate cell behavior in situations of tissue pathology.     

The WD40 repeat protein WDR26 binds Gβγ and promotes Gβγ-dependent signal transduction and leukocyte migration

The Gβγ subunits of heterotrimeric G proteins transmit signals to control many cellular processes, including leukocyte migration. Gβγ signaling may regulate and be regulated by numerous signaling partners. Here, we reveal that WDR26, a member of the WD40 repeat protein family, directly bound free Gβγ in vitro, and formed a complex with endogenous Gβγ in Jurkat T cells stimulated by the chemokine SDF1α. Suppression of WDR26 by siRNAs selectively inhibited Gβγ-dependent phospholipase Cβ and PI3K activation, and attenuated chemotaxis in Jurkat T cells and differentiated HL60 cells in vitro and Jurkat T cell homing to lymphoid tissues in scid mice. Similarly, disruption of the WDR26/Gβγ interaction via expression of a WDR26 deletion mutant impaired Gβγ signaling and Jurkat T cell migration, indicating that the function of WDR26 depends on its binding to Gβγ. Additional data show that WDR26 also controlled RACK1, a negative regulator, in binding Gβγ and inhibiting leukocyte migration. Collectively, these experiments identify WDR26 as a novel Gβγ-binding protein that is required for the efficacy of Gβγ signaling and leukocyte migration.     

苹果叶绿体基因组特征分析

苹果（Malus×domestica）是最重要的温带水果之一。为了能更好的了解本种的分子生物学基础．对已发布的苹果叶绿体全基因组序列进行了结构特征分析。结果显示苹果的叶绿体基因组全长为160068bp,具有典型的被子植物叶绿体基因组的环状四分体结构,包含大单拷贝区（LSC）,小单拷贝区（SSC）和两个反向互补重复区（IRs）,长度分别为88184bp,19180bp和26352bp。基因组共有135个基因（20个基因分布在反向互补重复区,因此整个基因组包含115个不同的基因）。按照功能进行分类,这115个基因包括81个蛋白质编码基因,4个rRNA编码基因和30个tRNA基因。其中,ycf15．ycf68和infA三个基因包含多个终止密码子,推测可能为假基因。苹果的基因组结构．基因顺序．GC含量和密码子使用偏好均与典型的被子植物叶绿体基因组类似。在苹果的叶绿体基因组中,共检测到30个大于30bp的重复序列,其中包括21串联重复,6个正向重复和3个反向重复序列;并检测到237个简单重复序列（SSR）位点,大部分的SSR位点都偏向于A或者T组成。此外,每10000bp非编码区平均分布有24个SSR位点,而编码区平均有5个SSR位点,表明SSRs在叶绿体基因组上的分布是不均匀的。本文对苹果叶绿体基因组序列特征的报道,将有助于促进该种的居群遗传学、系统发育和叶绿体基因工程的研究。     

Molecular characterization of Lathyrus species using chloroplast DNA trnH-psbA

Lathyrus L. is an important genus contributing in human food, animal feed and fodder. The genetic variation is studied among and within six species sampled over a large geographical area: Lathyrus cicera, Lathyrus sativus, Lathyrus sylvestris, Lathyrus tuberosus, Lathyrus ochrus and Lathyrus aphaca. The phylogenetic relationship among these species was assessed using sequences of chloroplast DNA trnH-psbA (intergenic spacer). The highly polymorphic spacer' length was 330 bp. The phylogenetic analyses using Maximum Parsimony and Genetic Distances, agreed with the universal taxonomy of Kupicha. L. sativus and L. cicera could be considered as sister species, sharing a common ancestor.