What leads from <Emphasis Type="Italic">dead-end?</Emphasis>

The 129 mouse strain develops congenital testicular germ cell tumors (TGCTs) at a low frequency. TGCTs in mice resemble the testicular tumors (teratomas) that occur in human infants. The genes that cause these tumors in 129 have not been identified. The defect at the Ter locus increases TGCT incidence such that 94% of 129-Ter/Ter males develop TGCTs. The primary effect of the Ter mutation is progressive loss of primordial germ cells (PGCs) during embryonic development. This results in sterility in adult Ter/Ter mice on all mouse strain backgrounds. However, on the 129 background, Ter causes tumor development in addition to sterility. Therefore, Ter acts as a modifier of 129-derived TGCT susceptibility genes. Ter was identified to be a mutation that inactivates the Dead-end1 (Dnd1) gene. In this perspective, I discuss the possible areas of future investigations to elucidate the mechanism of TGCT development due to Dnd1 inactivation. Received 29 September 2006; received after revision 29 January 2007; accepted 19 February 2007     

<Emphasis Type="Italic">Cis</Emphasis>–<Emphasis Type="Italic">trans</Emphasis> interactions of cell surface receptors: biological roles and structural basis

Cell surface receptors bind ligands expressed on other cells (in trans) in order to communicate with neighboring cells. However, an increasing number of cell surface receptors are found to also interact with ligands expressed on the same cell (in cis). These observations raise questions regarding the biological role of such cis interactions. Specifically, it is important to know whether cis and trans binding have distinct functional effects and, if so, how a single cell discriminates between interactions in cis versus trans. Further, what are the structural features that allow certain cell surface receptors to engage ligand both on the same as well as on an apposed cell membrane? Here, we summarize known examples of receptors that display cis–trans binding and discuss the emerging diversity of biological roles played by these unconventional two-way interactions, along with their structural basis.     

Beta-carotene affects gene expression in lungs of male and female <Emphasis Type="Italic">Bcmo1</Emphasis><Superscript>−/−</Superscript> mice in opposite directions

Molecular mechanisms triggered by high dietary beta-carotene (BC) intake in lung are largely unknown. We performed microarray gene expression analysis on lung tissue of BC supplemented beta-carotene 15,15′-monooxygenase 1 knockout (Bcmo1 ^−/−) mice, which are—like humans—able to accumulate BC. Our main observation was that the genes were regulated in an opposite direction in male and female Bcmo1 ^−/− mice by BC. The steroid biosynthetic pathway was overrepresented in BC-supplemented male Bcmo1 ^−/− mice. Testosterone levels were higher after BC supplementation only in Bcmo1 ^−/− mice, which had, unlike wild-type (Bcmo1 ^+/+) mice, large variations. We hypothesize that BC possibly affects hormone synthesis or metabolism. Since sex hormones influence lung cancer risk, these data might contribute to an explanation for the previously found increased lung cancer risk after BC supplementation (ATBC and CARET studies). Moreover, effects of BC may depend on the presence of frequent human BCMO1 polymorphisms, since these effects were not found in wild-type mice.     

"Add-on" domains of<Emphasis Type="Italic"> Drosophila</Emphasis> β1,4-<Emphasis Type="Italic">N</Emphasis>-acetylgalactosaminyltransferase B in the stem region and its pilot protein

The glycolipid specific Drosophila melanogaster β1,4-N-acetylgalactosaminyltransferase B (β4GalNAcTB) depends on a zinc finger DHHC protein family member named GalNAcTB pilot (GABPI) for activity and translocation to the Golgi. The six-membrane spanning protein actually lacks the cysteine in the cytoplasmic DHHC motif, displaying DHHS instead. Here we show that the whole conserved region around the DHHS sequence, which is essential for palmitoylation in DHHC proteins, is not required for GABPI to interact with β4GalNAcTB. In contrast, the two luminal loops between transmembrane domain 3–4 and 5–6 contain conserved amino acids, which are crucial for activity. Besides the dependence on GABPI, β4GalNAcTB requires its exceptional short stem region for activity. A few hydrophobic amino acids positioned close to the transmembrane domain are essential for the interaction with GABPI. Along with its catalytic domain, β4GalNAcTB, thus, requires an area in its own stem region and two small luminal loops of GABPI as "add-on" domains. Moreover, some inactive GABPI mutants could be rescued by fusion with β4GalNAcTB, indicating their importance in direct GABPI-β4GalNAcTB interaction.     

The role of the <Emphasis Type="Italic">ATP2C1</Emphasis> gene in Hailey–Hailey disease

Hailey–Hailey disease (HHD) is a rare autosomal dominant acantholytic dermatosis, characterized by a chronic course of repeated and exacerbated skin lesions in friction regions. The pathogenic gene of HHD was reported to be the ATPase calcium-transporting type 2C member 1 gene (ATP2C1) located on chromosome 3q21–q24. Its function is to maintain normal intracellular concentrations of Ca²⁺/Mn²⁺ by transporting Ca²⁺/Mn²⁺ into the Golgi apparatus. ATP2C1 gene mutations are reportedly responsible for abnormal cytosolic Ca²⁺/Mn²⁺ levels and the clinical manifestations of HHD. Environmental factors and genetic modifiers may also affect the clinical variability of HHD. This article aims to critically discuss the clinical and pathological features of HHD, differential diagnoses, and genetic and functional studies of the ATP2C1 gene in HHD. Further understanding the role of the ATP2C1 gene in the pathogenesis of HHD by genetic, molecular, and animal studies may contribute to a better clinical diagnosis and provide new strategies for the treatment and prevention of HHD.     

Proposition II (Book I) of Newton’s <Emphasis Type="Italic">Principia</Emphasis>

After preparing the way with comments on evanescent quantities and then Newton’s interpretation of his second law, this study of Proposition II (Book I)— Proposition II Every body that moves in some curved line described in a plane and, by a radius drawn to a point, either unmoving or moving uniformly forward with a rectilinear motion, describes areas around that point proportional to the times, is urged by a centripetal force tending toward that same point. —asks and answers the following questions: When does a version of Proposition II first appear in Newton’s work? What revisions bring that initial version to the final form in the 1726 Principia? What, exactly, does this proposition assert? In particular, what does Newton mean by the motion of a body “urged by a centripetal force”? Does it assert a true mathematical claim? If not, what revision makes it true? Does the demonstration of Proposition II persuade? Is it as convincing, for example, as the most convincing arguments of the Principia? If not, what revisions would make the demonstration more persuasive? What is the importance of Proposition II, to the physics of Book III and the mathematics of Book I?     

<Emphasis Type="Italic">Legionella pneumophila</Emphasis> — a human pathogen that co-evolved with fresh water protozoa

The bacterial pathogen Legionella pneumophila is found ubiquitously in fresh water environments where it replicates within protozoan hosts. When inhaled by humans it can replicate within alveolar macrophages and cause a severe pneumonia, Legionnaires disease. Yet much needs to be learned regarding the mechanisms that allow Legionella to modulate host functions to its advantage and the regulatory network governing its intracellular life cycle. The establishment and publication of the complete genome sequences of three clinical L. pneumophila isolates paved the way for major breakthroughs in understanding the biology of L. pneumophila. Based on sequence analysis many new putative virulence factors have been identified foremost among them eukaryotic-like proteins that may be implicated in many different steps of the Legionella life cycle. This review summarizes what is currently known about regulation of the Legionella life cycle and gives insight in the Legionella-specific features as deduced from genome analysis. Received 1 September 2006; received after revision 10 October 2006; accepted 22 November 2006     

The in vivo role of α-mannosidase IIx and its role in processing of <Emphasis Type="Italic">N</Emphasis>-glycans in spermatogenesis

The surfaces of mammalian cells are covered by a variety of carbohydrates linked to proteins and lipids. N-glycans are commonly found carbohydrates in plasma membrane proteins. The structure and biosynthetic pathway of N-glycans have been analyzed extensively. However, functional analysis of cell surface N-glycans is just under way with recent studies of targeted disruption of genes involved in N-glycan synthesis. This review briefly introduces the potential role of processing -mannosidases in N-glycan biosynthesis and recent findings derived from the -mannosidase IIx (MX) gene knockout mouse, which shows male infertility. Thus, the MX gene knockout experiment unveiled a novel function of specific N-glycan, which is N-acetylglucosamine-terminated and fucosylated triantennary structure, in the adhesion between germ cells and Sertoli cells. Analysis of the MX gene knockout mouse is a good example of a multidisciplinary approach leading to a novel discovery in the emerging field of glycobiology.Received 29 November 2002; received after revision 30 December 2002; accepted 20 January 2003     

BlaB,a protein involved in the regulation of <Emphasis Type="Italic">Streptomyces cacaoi</Emphasis> β-lactamases,is a penicillin-binding protein

Streptomyces cacaoi -lactamase genes are controlled by two regulators named blaA and blaB. Whereas BlaA has been identified as a LysR-type activator, the function of BlaB is still unknown. Its primary structure is similar to that of the serine penicillin-recognizing enzymes (PREs). Indeed, the SXXK and KTG motifs are perfectly conserved in BlaB, whereas the common SXN element found in PREs is replaced by a SDG motif. Site-directed mutations were introduced in these motifs and they all disturb -lactamase regulation. A water-soluble form of BlaB was also overexpressed in the Streptomyces lividans TK24 cytoplasm and purified. To elucidate the activity of BlaB, several compounds recognized by PREs were tested. BlaB could be acylated by some of them, and it can therefore be considered as a penicillin-binding protein. BlaB is devoid of -lactamase, D-aminopeptidase, DD-carboxypeptidase or thiolesterase activity.Received 13 January 2003; received after revision 9 April 2003; accepted 11 April 2003     

<Emphasis Type="Italic">Trichomonas vaginalis</Emphasis> degrades nitric oxide and expresses a flavorubredoxin-like protein: a new pathogenic mechanism?

Besides possessing many physiological roles, nitric oxide (NO) produced by the immune system in infectious diseases has antimicrobial effects. Trichomoniasis, the most widespread non-viral sexually transmitted disease caused by the microaerophilic protist Trichomonas vaginalis, often evolves into a chronic infection, with the parasite able to survive in the microaerobic, NO-enriched vaginal environment. We relate this property to the finding that T. vaginalis degrades NO under anaerobic conditions, as assessed amperometrically. This activity, which is maximal (133 ± 41 nmol NO/10⁸ cells per minute at 20°C) at low NO concentrations ( 1.2 M), was found to be: (i) NADH dependent, (ii) cyanide insensitive and (iii) inhibited by O₂. These features are consistent with those of the Escherichia coli A-type flavoprotein (ATF), recently discovered to be endowed with NO reductase activity. Using antibodies against the ATF from E. coli, a protein band was immunodetected in the parasite grown in a standard medium. If confirmed, the expression of an ATF in eukaryotes suggests that the genes coding for ATFs were transferred during evolution from anaerobic Prokarya to pathogenic protists, to increase their fitness for the microaerobic, parasitic life style. Thus the demonstration of an ATF in T. vaginalis would appear relevant to both pathology and evolutionary biology. Interestingly, genomic analysis has recently demonstrated that Giardia intestinalis and other pathogenic protists have genes coding for ATFs.Received 1 November 2003; received after revision 5 January 2004; accepted 13 January 2004     

Muscle LIM protein promotes expression of the acetylcholine receptor <Emphasis Type="Italic">γ</Emphasis>-subunit gene cooperatively with the myogenin-E12 complex

Muscle LIM protein (MLP, also referred to as CRP3) is a muscle-specific LIM-only protein, which consists of two LIM motifs. MLP functions as a positive regulator during myogenesis. Here we report that MLP serves as a cofactor regulating the expression of the nicotinic acetylcholine receptor (AChR) -subunit gene in skeletal muscle cells. We found that MLP promoted the expression of the AChR -subunit gene in C2C12 myotubes, but not in C2C12 myoblasts or NIH3T3 fibroblasts. Furthermore, we showed that MLP interacted with myogenin in vivo and enhanced the binding ability of the myogenin-E12 heterodimer to the E boxes in the AChR -subunit gene promoter. Together, these results suggest that MLP promotes the specific expression of the AChR -subunit gene cooperatively with the myogenin-E12 complex during myogenesis.Received 17 May 2004; received after revision 22 July 2004; accepted 26 July 2004     

<Emphasis Type="Italic">O</Emphasis>-GlcNAc transferase associates with the MCM2–7 complex and its silencing destabilizes MCM–MCM interactions

O-GlcNAcylation of proteins is governed by O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA). The homeostasis of O-GlcNAc cycling is regulated during cell cycle progression and is essential for proper cellular division. We previously reported the O-GlcNAcylation of the minichromosome maintenance proteins MCM2, MCM3, MCM6 and MCM7. These proteins belong to the MCM2–7 complex which is crucial for the initiation of DNA replication through its DNA helicase activity. Here we show that the six subunits of MCM2–7 are O-GlcNAcylated and that O-GlcNAcylation of MCM proteins mainly occurs in the chromatin-bound fraction of synchronized human cells. Moreover, we identify stable interaction between OGT and several MCM subunits. We also show that down-regulation of OGT decreases the chromatin binding of MCM2, MCM6 and MCM7 without affecting their steady-state level. Finally, OGT silencing or OGA inhibition destabilizes MCM2/6 and MCM4/7 interactions in the chromatin-enriched fraction. In conclusion, OGT is a new partner of the MCM2–7 complex and O-GlcNAcylation homeostasis might regulate MCM2–7 complex by regulating the chromatin loading of MCM6 and MCM7 and stabilizing MCM/MCM interactions.     

The epididymis-specific antimicrobial peptide β-defensin 15 is required for sperm motility and male fertility in the rat (<Emphasis Type="Italic">Rattus norvegicus</Emphasis>)

Spermatozoa acquire forward motility and fertilizing capacity during their transit through the epididymis. The maturation process involves modifications of the sperm surface by different proteins that are secreted by a series of specialized regions in the epididymal epithelium. Here we show that the rat epididymis-specific β-defensin 15 (Defb15) exhibits an androgen-dependent expression pattern, and it can bind to the acrosomal region of caput sperm. Coculture of caput spermatozoa with Defb15 antibody in vitro resulted in a significant decline in sperm motility. Moreover, the total and progressive motility of the spermatozoa dramatically decreased in rats when Defb15 was downregulated by lentivirus-mediated RNAi in vivo. Remarkably, knock down of Defb15 led to a reduction in fertility and embryonic development failure. In addition, the recombinant Defb15 showed antimicrobial activity in a dose-dependent fashion. These results suggest that Defb15 plays a dual role in both sperm maturation and pathogen defense in rat epididymis.     

Crystal structure of <Emphasis Type="Italic">Enterobacter cloacae</Emphasis> 908R class C β-lactamase bound to iodo-acetamido-phenyl boronic acid,a transition-state analogue

The structures of the class C -lactamase from Enterobacter cloacae 908R alone and in complex with a boronic acid transition-state analogue were determined by X-ray crystallography at 2.1 and 2.3 Å, respectively. The structure of the enzyme resembles those of other class C -lactamases. The structure of the complex with the transition-state analogue, iodo-acetamido-phenyl boronic acid, shows that the inhibitor is covalently bound to the active-site serine (Ser64). Binding of the inhibitor within the active site is compared with previously determined structures of complexes with other class C enzymes. The structure of the boronic acid adduct indicates ways to improve the affinity of this class of inhibitors. This structure of 908R class C -lactamase in complex with a transition-state analogue provides further insights into the mechanism of action of these hydrolases.Received 16 May 2003; accepted 4 June 2003     

<Emphasis Type="Italic">Clostridium difficile</Emphasis>-related postinfectious IBS: a case of enteroglial microbiological stalking and/or the solution of a conundrum?

Post-infectious irritable bowel syndrome is a well-defined pathological entity that develops in about one-third of subjects after an acute infection (bacterial, viral) or parasitic infestation. Only recently it has been documented that an high incidence of post-infectious irritable bowel syndrome occurs after Clostridium difficile infection. However, until now it is not known why in some patients recovered from this infection the gastrointestinal disturbances persist for months or years. Based on our in vitro studies on enteric glial cells exposed to the effects of C. difficile toxin B, we hypothesize that persistence of symptoms up to the development of irritable bowel syndrome might be due to a disturbance/impairment of the correct functions of the enteroglial intestinal network.     

CD8<Superscript>+</Superscript> Tregs in autoimmunity: learning “self”-control from experience

Autoreactive CD8⁺ regulatory T cells (Tregs) play important roles as modulators of immune responses against self, and numerical and functional defects in CD8⁺ Tregs have been linked to autoimmunity. Several subsets of CD8⁺ Tregs have been described. However, the origin of these T cells and how they participate in the natural progression of autoimmunity remain poorly defined. We discuss several lines of evidence suggesting that the autoimmune process itself promotes the development of autoregulatory CD8⁺ T cells. We posit that chronic autoantigenic exposure fosters the differentiation of non-pathogenic autoreactive CD8⁺ T cells into antigen-experienced, memory-like autoregulatory T cells, to generate a “negative feedback” regulatory loop capable of countering pathogenic autoreactive effectors. This hypothesis predicts that approaches capable of boosting autoregulatory T cell memory will be able to blunt autoimmunity without compromising systemic immunity.