L-asparaginase inhibits invasive and angiogenic activity and induces autophagy in ovarian cancer

Recent work identified L‐asparaginase (L‐ASP) as a putative therapeutic target for ovarian cancer. We suggest that L‐ASP, a dysregulator of glycosylation, would interrupt the local microenvironment, affecting the ovarian cancer cell—endothelial cell interaction and thus angiogenesis without cytotoxic effects. Ovarian cancer cell lines and human microvascular endothelial cells (HMVEC) were exposed to L‐ASP at physiologically attainable concentrations and subjected to analyses of endothelial tube formation, invasion, adhesion and the assessment of sialylated proteins involved in matrix‐associated and heterotypic cell adhesion. Marked reduction in HMVEC tube formation in vitro, HMVEC and ovarian cancer cell invasion, and heterotypic cell‐cell and cell‐matrix adhesion was observed (P < 0.05–0.0001). These effects were associated with reduced binding to ß1integrin, activation of FAK, and cell surface sialyl Lewis^X (sLe^x) expression. No reduction in HMVEC E‐selectin expression was seen consistent with the unidirectional inhibitory actions observed. L‐ASP concentrations were non‐toxic to either ovarian cancer or HMVEC lines in the time frame of the assays. However, early changes of autophagy were observed in both cell types with induction of ATG12, beclin‐1, and cleavage of LC‐3, indicating cell injury did occur. These data and the known mechanism of action of L‐ASP on glycosylation of nascent proteins suggest that L‐ASP reduces of ovarian cancer dissemination and progression through modification of its microenvironment. The reduction of ovarian cancer cell surface sLe^x inhibits interaction with HMVEC and thus HMVEC differentiation into tubes, inhibits interaction with the local matrix reducing invasive behaviour, and causes cell injury initiating autophagy in tumour and vascular cells.     

Infection of differentiated porcine airway epithelial cells by influenza virus: differential susceptibility to infection by porcine and avian viruses

Background

Swine are important hosts for influenza A viruses playing a crucial role in the epidemiology and interspecies transmission of these viruses. Respiratory epithelial cells are the primary target cells for influenza viruses.

Methodology/Principal Findings

To analyze the infection of porcine airway epithelial cells by influenza viruses, we established precision-cut lung slices as a culture system for differentiated respiratory epithelial cells. Both ciliated and mucus-producing cells were found to be susceptible to infection by swine influenza A virus (H3N2 subtype) with high titers of infectious virus released into the supernatant already one day after infection. By comparison, growth of two avian influenza viruses (subtypes H9N2 and H7N7) was delayed by about 24 h. The two avian viruses differed both in the spectrum of susceptible cells and in the efficiency of replication. As the H9N2 virus grew to titers that were only tenfold lower than that of a porcine H3N2 virus this avian virus is an interesting candidate for interspecies transmission. Lectin staining indicated the presence of both α-2,3- and α-2,6-linked sialic acids on airway epithelial cells. However, their distribution did not correlate with pattern of virus infection indicating that staining by plant lectins is not a reliable indicator for the presence of cellular receptors for influenza viruses.

Conclusions/Significance

Differentiated respiratory epithelial cells significantly differ in their susceptibility to infection by avian influenza viruses. We expect that the newly described precision-cut lung slices from the swine lung are an interesting culture system to analyze the infection of differentiated respiratory epithelial cells by different pathogens (viral, bacterial and parasitic ones) of swine.     

Dasty and UniProt DAS: a perfect pair for protein feature visualization

In this study, we present two freely available and complementary Distributed Annotation System (DAS) resources: a DAS reference server that provides up-to-date sequence and annotation from UniProt, with additional feature links and database cross-references from InterPro and a DAS client implemented using Java and Macromedia Flash that is optimized for the display of protein features.     

Structure and function of the human sperm-specific isoform of protein kinase A (PKA) catalytic subunit Cα2

Protein kinase A (PKA) exists as several tissue-specific isoforms that through phosphorylation of serine and threonine residues of substrate proteins act as key regulators of a number of cellular processes. We here demonstrate that the human sperm-specific isoform of PKA named Cα2 is important for sperm motility and thus male fertility. Furthermore, we report on the first three-dimensional crystal structure of human apo Cα2 to 2.1 ?. Apo Cα2 displays an open conformation similar to the well-characterized apo structure of murine Cα1. The asymmetric unit contains two molecules and the core of the small lobe is rotated by almost 13° in the A molecule relative to the B molecule. In addition, a salt bridge between Lys72 and Glu91 was observed for Cα2 in the apo-form, a conformation previously found only in dimeric or ternary complexes of Cα1. Human Cα2 and Cα1 share primary structure with the exception of the amino acids at the N-terminus coded for by an alternative exon 1. The N-terminal glycine of Cα1 is myristoylated and this aliphatic chain anchors the N-terminus to an intramolecular hydrophobic pocket. Cα2 cannot be myristoylated and the crystal structure revealed that the equivalent hydrophobic pocket is unoccupied and exposed. Nuclear magnetic resonance (NMR) spectroscopy further demonstrated that detergents with hydrophobic moieties of different lengths can bind deep into this uncovered pocket. Our findings indicate that Cα2 through the hydrophobic pocket has the ability to bind intracellular targets in the sperm cell, which may modulate protein stability, activity and/or cellular localization.     

Prenylflavonoid variation in Humulus lupulus: distribution and taxonomic significance of xanthogalenol and 4'-O-methylxanthohumol

The resins produced by either lupulin or leaf glands of over 120 plants of Humulus lupulus and one plant of H. japonicus (Cannabinaceae) were analyzed for the presence of prenylated flavonoids. The H. lupulus taxa investigated were H. lupulus var. lupulus from Europe, H. lupulus var. cordifolius from Japan, and H. lupulus from North America. Fifty-two of the plants examined were cultivars of European, American, and Japanese origin. Twenty-two flavonoids were detected in the glandular exudates of H. lupulus by HPLC-MS MS. Xanthohumol (3'-prenyl-6'-O-methylchalconaringenin) was the principal prenylflavonoid in all H. lupulus plants and was accompanied by 11 structurally similar chalcones. Ten flavonoids were identified as the flavanone isomers of these chalcones. Three other prenylchalcones were isolated from H. lupulus cv. 'Galena', one of which was identified as 3'-prenyl-4'-O-methylchalconaringenin (named 'xanthogalenol'). The distribution of three 4'-O-methylchalcones, i.e. xanthogalenol, 4'-O-methylxanthohumol, and 4',6'-di-O-methylchalconaringenin, was found to be limited to wild American plants from the Missouri-Mississippi river basin, H. lupulus var. cordifolius, and most of their descendents. These 4'-O-methylchalcones were absent from cultivars of European origin, and from wild hops from Europe and southwestern USA. The flavonoid dichotomy (presence versus absence of 4'-O-methylchalcones) indicates that there are at least two evolutionary lineages within H. lupulus (European and Japanese American), which is in agreement with morphological, molecular, and phytogeographical evidence. Leaf glands of H. japonicus from eastern Asia did not produce the H. lupulus prenylflavonoids.     

Surprisingly long survival of premature conclusions about naked mole-rat biology

Naked mole-rats express many unusual traits for such a small rodent. Their morphology, social behaviour, physiology, and ageing have been well studied over the past half-century. Many early findings and speculations about this subterranean species persist in the literature, although some have been repeatedly questioned or refuted. While the popularity of this species as a natural-history curiosity, and oversimplified story-telling in science journalism, might have fuelled the perpetuation of such misconceptions, an accurate understanding of their biology is especially important for this new biomedical model organism. We review 28 of these persistent myths about naked mole-rat sensory abilities, ecophysiology, social behaviour, development and ageing, and where possible we explain how these misunderstandings came about.     

Coronavirus Infection and Diversity in Bats in the Australasian Region

Following the SARS outbreak, extensive surveillance was undertaken globally to detect and identify coronavirus diversity in bats. This study sought to identify the diversity and prevalence of coronaviruses in bats in the Australasian region. We identified four different genotypes of coronavirus, three of which (an alphacoronavirus and two betacoronaviruses) are potentially new species, having less than 90% nucleotide sequence identity with the most closely related described viruses. We did not detect any SARS-like betacoronaviruses, despite targeting rhinolophid bats, the putative natural host taxa. Our findings support the virus-host co-evolution hypothesis, with the detection of Miniopterus bat coronavirus HKU8 (previously reported in Miniopterus species in China, Hong Kong and Bulgaria) in Australian Miniopterus species. Similarly, we detected a novel betacoronavirus genotype from Pteropus alecto which is most closely related to Bat coronavirus HKU9 identified in other pteropodid bats in China, Kenya and the Philippines. We also detected possible cross-species transmission of bat coronaviruses, and the apparent enteric tropism of these viruses. Thus, our findings are consistent with a scenario wherein the current diversity and host specificity of coronaviruses reflects co-evolution with the occasional host shift.     

Prevalence of chlamydiae in boars and semen used for artificial insemination

Although there are indications for venereal transmission of chlamydiae in pigs, direct diagnostic evidence on the presence of these bacteria in boars and boar semen in particular is still incomplete. We investigated boars from two studs (A, B) in semen (A: n = 174; B: n = 100) and faeces (A: n = 174; B: n = 24) for chlamydiae using ompA-PCR and partial ompA gene sequencing. Additionally, blood serum was examined for chlamydial antibodies using an indirect ELISA (A: n = 171; B: n = 62). Chlamydiae were found in 9 (5.2%) and 24 (24.0%) semen specimens, and in 71 (40.1%) and 2 (8.3%) faecal samples from boars of stud A and B, respectively. Regarding individual chlamydial species, Chlamydophila psittaci and Chlamydia suis were identified most frequently, with the former predominating in semen (in 23 out of 33 positive samples) and the latter in faeces (68/73). In contrast, Chlamydophila pecorum was found only sporadically. Chlamydial antibodies were detected in 80 (46.8%) and 6 (9.7%) boars of stud A and B, respectively. No correlation was observed between the data from serology and PCR of semen or faeces in either of the studs. In conclusion, detection of chlamydiae in semen of boars suggests a potential for venereal transmission. Whether the high overall prevalence of chlamydial infections reflects a general situation in boars needs to be investigated. Serological testing failed to identify boars shedding chlamydiae in their semen.