Membrane Protein Biogenesis in Ffh- or FtsY-Depleted Escherichia coli

BACKGROUND: The Escherichia coli version of the mammalian signal recognition particle (SRP) system is required for biogenesis of membrane proteins and contains two essential proteins: the SRP subunit Ffh and the SRP-receptor FtsY. Scattered in vivo studies have raised the possibility that expression of membrane proteins is inhibited in cells depleted of FtsY, whereas Ffh-depletion only affects their assembly. These differential results are surprising in light of the proposed model that FtsY and Ffh play a role in the same pathway of ribosome targeting to the membrane. Therefore, we decided to evaluate these unexpected results systematically. METHODOLOGY/PRINCIPAL FINDINGS: We characterized the following aspects of membrane protein biogenesis under conditions of either FtsY- or Ffh-depletion: (i) Protein expression, stability and localization; (ii) mRNA levels; (iii) folding and activity. With FtsY, we show that it is specifically required for expression of membrane proteins. Since no changes in mRNA levels or membrane protein stability were detected in cells depleted of FtsY, we propose that its depletion may lead to specific inhibition of translation of membrane proteins. Surprisingly, although FtsY and Ffh function in the same pathway, depletion of Ffh did not affect membrane protein expression or localization. CONCLUSIONS: Our results suggest that indeed, while FtsY-depletion affects earlier steps in the pathway (possibly translation), Ffh-depletion disrupts membrane protein biogenesis later during the targeting pathway by preventing their functional assembly in the membrane

Proteins and DNA elements essential for the CRISPR adaptation process in Escherichia coli

The clustered regularly interspaced short palindromic repeats and their associated proteins (CRISPR/Cas) constitute a recently identified prokaryotic defense mechanism against invading nucleic acids. Activity of the CRISPR/Cas system comprises of three steps: (i) insertion of alien DNA sequences into the CRISPR array to prevent future attacks, in a process called ‘adaptation’, (ii) expression of the relevant proteins, as well as expression and processing of the array, followed by (iii) RNA-mediated interference with the alien nucleic acid. Here we describe a robust assay in Escherichia coli to explore the hitherto least-studied process, adaptation. We identify essential genes and DNA elements in the leader sequence and in the array which are essential for the adaptation step. We also provide mechanistic insights on the insertion of the repeat-spacer unit by showing that the first repeat serves as the template for the newly inserted repeat. Taken together, our results elucidate fundamental steps in the adaptation process of the CRISPR/Cas system

A Novel Translocation Breakpoint within the BPTF Gene Is Associated with a Pre-Malignant Phenotype

Partial gain of chromosome arm 17q is an abundant aberrancy in various cancer types such as lung and prostate cancer with a prominent occurrence and prognostic significance in neuroblastoma – one of the most common embryonic tumors. The specific genetic element/s in 17q responsible for the cancer-promoting effect of these aberrancies is yet to be defined although many genes located in 17q have been proposed to play a role in malignancy. We report here the characterization of a naturally-occurring, non-reciprocal translocation der(X)t(X;17) in human lung embryonal-derived cells following continuous culturing. This aberrancy was strongly correlated with an increased proliferative capacity and with an acquired ability to form colonies in vitro. The breakpoint region was mapped by fluorescence in situ hybridization (FISH) to the 17q24.3 locus. Further characterization by a custom-made comparative genome hybridization array (CGH) localized the breakpoint within the Bromodomain PHD finger Transcription Factor gene (BPTF), a gene involved in transcriptional regulation and chromatin remodeling. Interestingly, this translocation led to elevation in the mRNA levels of the endogenous BPTF. Knock-down of BPTF restricted proliferation suggesting a role for BPTF in promoting cellular growth. Furthermore, the BPTF chromosomal region was found to be amplified in various human tumors, especially in neuroblastomas and lung cancers in which 55% and 27% of the samples showed gain of 17q24.3, respectively. Additionally, 42% percent of the cancer cell lines comprising the NCI-60 had an abnormal BPTF locus copy number. We suggest that deregulation of BPTF resulting from the translocation may confer the cells with the observed cancer-promoting phenotype and that our cellular model can serve to establish causality between 17q aberrations and carcinogenesis

Modulated expression of WFDC1 during carcinogenesis and cellular senescence

Fibroblasts located adjacent to the tumor [cancer-associated fibroblasts (CAFs)] that constitute a large proportion of the cancer-associated stroma facilitate the transformation process. In this study, we compared the biological behavior of CAFs that were isolated from a prostate tumor to their normal-associated fibroblast (NAF) counterparts. CAFs formed more colonies when seeded at low cell density, exhibited a higher proliferation rate and were less prone to contact inhibition. In contrast to the general notion that high levels of α-smooth muscle actin serve as a marker for CAFs, we found that prostate CAFs express it at a lower level compared with prostate NAFs. Microarray analysis revealed a set of 161 genes that were altered in CAFs compared with NAFs. We focused on whey acidic protein four-disulfide core domain 1 (WFDC1), a known secreted protease inhibitor, and found it to be downregulated in the CAFs. WFDC1 expression was also dramatically downregulated in highly prolific mesenchymal cells and in various cancers including fibrosarcomas and in tumors of the lung, bladder and brain. Overexpression of WFDC1 inhibited the growth rate of the fibrosarcoma HT1080 cell line. Furthermore, WFDC1 level was upregulated in senescent fibroblasts. Taken together, our data suggest an important role for WFDC1 in inhibiting proliferation of both tumors and senescent cells. Finally, we suggest that the downregulation of WFDC1 might serve as a biomarker for cellular transformation

Distribution and Extinction of Ungulates during the Holocene of the Southern Levant

BACKGROUND: The southern Levant (Israel, Palestinian Authority and Jordan) has been continuously and extensively populated by succeeding phases of human cultures for the past 15,000 years. The long human impact on the ancient landscape has had great ecological consequences, and has caused continuous and accelerating damage to the natural environment. The rich zooarchaeological data gathered at the area provide a unique opportunity to reconstruct spatial and temporal changes in wild species distribution, and correlate them with human demographic changes. METHODOLOGY: Zoo-archaeological data (382 animal bone assemblages from 190 archaeological sites) from various time periods, habitats and landscapes were compared. The bone assemblages were sorted into 12 major cultural periods. Distribution maps showing the presence of each ungulate species were established for each period. CONCLUSIONS: The first major ungulate extinction occurred during the local Iron Age (1,200-586 BCE), a period characterized by significant human population growth. During that time the last of the largest wild ungulates, the hartebeest (Alcelaphus buselaphus), aurochs (Bos primigenius) and the hippopotamus (Hippopotamus amphibius) became extinct, followed by a shrinking distribution of forest-dwelling cervids. A second major wave of extinction occurred only in the 19th and 20th centuries CE. Furthermore, a negative relationship was found between the average body mass of ungulate species that became extinct during the Holocene and their extinction date. It is thus very likely that the intensified human activity through habitat destruction and uncontrolled hunting were responsible for the two major waves of ungulate extinction in the southern Levant during the late Holocene

The Human Cell Atlas.

The recent advent of methods for high-throughput single-cell molecular profiling has catalyzed a growing sense in the scientific community that the time is ripe to complete the 150-year-old effort to identify all cell types in the human body. The Human Cell Atlas Project is an international collaborative effort that aims to define all human cell types in terms of distinctive molecular profiles (such as gene expression profiles) and to connect this information with classical cellular descriptions (such as location and morphology). An open comprehensive reference map of the molecular state of cells in healthy human tissues would propel the systematic study of physiological states, developmental trajectories, regulatory circuitry and interactions of cells, and also provide a framework for understanding cellular dysregulation in human disease. Here we describe the idea, its potential utility, early proofs-of-concept, and some design considerations for the Human Cell Atlas, including a commitment to open data, code, and community

Moult Strategies Affect Age Differences in Autumn Migration Timing in East Mediterranean Migratory Passerines

<div><p>Adult passerines renew their flight feathers at least once every year. This complete moult occurs either in the breeding areas, just after breeding (summer moult), or, in some long-distance migratory species, at the non-breeding areas, after arrival to the southern wintering area at the end of autumn migration (winter moult). The aim of this study was to relate moult strategies with the DMD, the difference in median migration date, through Israel, between juveniles and adults. Our data on autumn migration timing in juveniles and adults was based on ringing data of 49,125 individuals belonging to 23 passerine species that breed in Europe and Western Asia and migrate through Israel. We found that DMD was associated with moult timing. In all species that perform a winter moult, adults preceded juveniles during autumn. Among migrants who perform a summer moult, we found evidence of both migration timing patterns: juveniles preceding adults or adults preceding juveniles. In addition, in summer moulters, we found a significant, positive correlation between mean breeding latitude and DMD. Although previous studies described that moult duration and extent can be affected by migration, we suggest that moult strategies affect both migration timing and migration strategy. These two moult strategies (summer or winter moult) also represent two unique migration strategies. Our findings highlight the evolutionary interplay between moult and migration strategies.</p></div

The difference in median migration date (DMD), through Israel, between juveniles and adults (days) in winter moulters (n = 8 species) and summer moulters (n = 15 species).

<p>The difference between the two groups was significant) Mann-Whitney U-Test, U = 3.00, Z = 3.69, P < 0.001).</p