Disruption of the dynamic sub-cellular localization of the Xenopus tumorhead protein causes embryonic lethality at the early gastrula transition

Abstract The Xenopus laevis tumorhead (TH) protein, a positive regulator of cell proliferation during embryogenesis, shuttles from the cell periphery into the nucleus during embryogenesis. In these studies, we performed a detailed analysis of TH's subcellular localization pattern to characterize its dynamic behavior. We found that TH exhibits distinct patterns of localization in different germ layers. At the blastula stage, TH is present in the apical cell periphery of prospective mesodermal and ectodermal cells. At the gastrula stage, TH is distributed throughout the entire cytoplasm of prospective mesodermal and ectodermal cells, whereas it shows nuclear localization in presumptive endodermal cells. TH moves into the nucleus of mesodermal and ectodermal cells during the neurula and early tailbud stages. To understand if TH is regulated by changes in its subcellular localization, we used a TH mutant containing signals for farnesylation and palmitoylation to tether the protein to the plasma membrane. Ubiquitous overexpression of this mutant causes embryonic lethality at the early gastrula transition. Further examination using TUNEL assays indicated that wild-type TH overexpression induces apoptosis during gastrulation, and that this effect is exacerbated by the overexpression of the membrane-bound TH mutant. Taken together, our results suggest that changes in the sub-cellular localization of the TH protein are important for its function because blocking the nuclear translocation of overexpressed TH increases apoptosis and causes embryos to die. Our data also suggest that TH plays a role outside the nucleus when it is present at the cell periphery.     

Serious doubts over “Eggs forever?”

A recent commentary in this journal by Byskov et al. (2005) claims that, despite published results from numerous independent lines of investigation from our laboratory and others, there does not "exist any evidence for neo-folliculogenesis in the adult mammalian ovary." While we agree with Byskov et al. that our work represents a radical departure from the age-old dogma that mammalian females permanently lose the capacity for oocyte and follicle production during the perinatal period, careful examination of all of the available data leaves no doubt that adult female mammals retain the capacity for oogenesis and folliculogenesis. These findings do not change the fact that exhaustion of the oocyte pool occurs with advancing chronological age--a process responsible for driving the menopause in women--but rather question the basic mechanism underlying age-related ovarian failure. In this regard, studies of aging male mice have demonstrated that testicular atrophy is associated with a dramatic decline in the number, activity and quality of germline stem cells that maintain spermatogenesis during adulthood (Zhang et al., 2006). Therefore, to the contrary of the opinion of Byskov et al. that such a process would be "considered exceptional among stem cells," it is certainly reasonable to hypothesize that a similar deterioration of female germline stem cell function underlies the decline in oocyte quality and the onset of ovarian failure in aging females. Further, while we accept that a departure from conventional thinking can take years to gain widespread acceptance, we feel this resistance to change should not be construed as the sole means to voice opinions about the validity of our data or the maturity of our principal conclusion.     

Hermes RNA-binding protein targets RNAs-encoding proteins involved in meiotic maturation, early cleavage, and germline development

The early development of metazoans is mainly regulated by differential translation and localization of maternal mRNAs in the embryo. In general, these processes are orchestrated by RNA-binding proteins interacting with specific sequence motifs in the 3'-untranslated region (UTR) of their target RNAs. Hermes is an RNA-binding protein, which contains a single RNA recognition motif (RRM) and is found in various vertebrate species from fish to human. In Xenopus laevis, Hermes mRNA and protein are localized in the vegetal region of oocytes. A subpopulation of Hermes protein is concentrated in a specific structure in the vegetal cortex, called the germ plasm (believed to contain determinants of the germ cell fate) where Hermes protein co-localizes with Xcat2 and RINGO/Spy mRNAs. The level of total Hermes protein decreases during maturation. The precocious depletion of Hermes protein by injection of Hermes antisense morpholino oligonucleotide (HE-MO) accelerates the process of maturation and results in cleavage defects in vegetal blastomeres of the embryo. It is known that several maternal mRNAs including RINGO/Spy and Mos are regulated at the translational level during meiotic maturation and early cleavage in Xenopus. The ectopic expression of RINGO/Spy or Mos causes resumption of meiotic maturation and cleavage arrests, which resemble the loss of Hermes phenotypes. We found that the injection of HE-MO enhances the acceleration of maturation caused by the injection of RINGO/Spy mRNA, and that Hermes protein is present as mRNP complex containing RINGO/Spy, Mos, and Xcat2 mRNAs in vivo. We propose that as an RNA-binding protein, Hermes may be involved in maturation, cleavage events at the vegetal pole and germ cell development by negatively regulating the expression of RINGO/Spy, Mos, and Xcat2 mRNAs.     

Long-term hypoxia modulates expression of key genes regulating adrenomedullary function in the late gestation ovine fetus

We previously communicated that long-term hypoxia (LTH) resulted in a selective reduction in plasma epinephrine following acute stress in fetal sheep. The present study tested the hypothesis that LTH selectively reduces adrenomedullary expression of phenylethanolamine-N-methyltransferase (PNMT), the rate-limiting enzyme for epinephrine synthesis. We also examined the effect of LTH on adrenomedullary nicotinic, muscarinic, and glucocorticoid receptor (GR) expression. Ewes were maintained at high altitude (3,820 m) from 30 to 138 days gestation (dGA); adrenomedullary tissue was collected from LTH and age-matched, normoxic control fetuses at 139-141 dGA. Contrary to our hypothesis, in addition to PNMT, adrenomedullary expression (mRNA, protein) of tyrosine hydroxylase (TH) and dopamine beta-hydroxylase (DBH) were reduced in the LTH fetus. Immunocytochemistry indicated that TH and DBH expression was lower throughout the medulla, while PNMT appeared to reflect a reduction in PNMT-expressing cells. Nicotinic receptor alpha 1, 2, 3, 5, 6, 7, beta 1, 2, and 4 subunits were expressed in the medulla of LTH and control fetuses. Messenger RNA for alpha 1 and 7 and beta 1 and 2 subunits was lower in LTH fetuses. Muscarinic receptors M1, M2, and M3 as well as the GR were also expressed, and no differences were noted between groups. In summary, LTH in fetal sheep has a profound effect on expression of key enzymes mediating adrenomedullary catecholamine synthesis. Further, LTH impacts nicotinic receptor subunit expression potentially altering cholinergic neurotransmission within the medulla. These findings have important implications regarding fetal cardiovascular and metabolic responses to stress in the LTH fetus.     

Lysigenous aerenchyma formation in Arabidopsis is controlled by LESION SIMULATING DISEASE1

Aerenchyma tissues form gas-conducting tubes that provide roots with oxygen under hypoxic conditions. Although aerenchyma have received considerable attention in Zea mays, the signaling events and genes controlling aerenchyma induction remain elusive. Here, we show that Arabidopsis thaliana hypocotyls form lysigenous aerenchyma in response to hypoxia and that this process involves H(2)O(2) and ethylene signaling. By studying Arabidopsis mutants that are deregulated for excess light acclimation, cell death, and defense responses, we find that the formation of lysigenous aerenchyma depends on the plant defense regulators LESION SIMULATING DISEASE1 (LSD1), ENHANCED DISEASE SUSCEPIBILITY1 (EDS1), and PHYTOALEXIN DEFICIENT4 (PAD4) that operate upstream of ethylene and reactive oxygen species production. The obtained results indicate that programmed cell death of lysigenous aerenchyma in hypocotyls occurs in a similar but independent manner from the foliar programmed cell death. Thus, the induction of aerenchyma is subject to a genetic and tissue-specific program. The data lead us to conclude that the balanced activities of LSD1, EDS1, and PAD4 regulate lysigenous aerenchyma formation in response to hypoxia.     

The creatine kinase system and pleiotropic effects of creatine

The pleiotropic effects of creatine (Cr) are based mostly on the functions of the enzyme creatine kinase (CK) and its high-energy product phosphocreatine (PCr). Multidisciplinary studies have established molecular, cellular, organ and somatic functions of the CK/PCr system, in particular for cells and tissues with high and intermittent energy fluctuations. These studies include tissue-specific expression and subcellular localization of CK isoforms, high-resolution molecular structures and structure–function relationships, transgenic CK abrogation and reverse genetic approaches. Three energy-related physiological principles emerge, namely that the CK/PCr systems functions as (a) an immediately available temporal energy buffer, (b) a spatial energy buffer or intracellular energy transport system (the CK/PCr energy shuttle or circuit) and (c) a metabolic regulator. The CK/PCr energy shuttle connects sites of ATP production (glycolysis and mitochondrial oxidative phosphorylation) with subcellular sites of ATP utilization (ATPases). Thus, diffusion limitations of ADP and ATP are overcome by PCr/Cr shuttling, as most clearly seen in polar cells such as spermatozoa, retina photoreceptor cells and sensory hair bundles of the inner ear. The CK/PCr system relies on the close exchange of substrates and products between CK isoforms and ATP-generating or -consuming processes. Mitochondrial CK in the mitochondrial outer compartment, for example, is tightly coupled to ATP export via adenine nucleotide transporter or carrier (ANT) and thus ATP-synthesis and respiratory chain activity, releasing PCr into the cytosol. This coupling also reduces formation of reactive oxygen species (ROS) and inhibits mitochondrial permeability transition, an early event in apoptosis. Cr itself may also act as a direct and/or indirect anti-oxidant, while PCr can interact with and protect cellular membranes. Collectively, these factors may well explain the beneficial effects of Cr supplementation. The stimulating effects of Cr for muscle and bone growth and maintenance, and especially in neuroprotection, are now recognized and the first clinical studies are underway. Novel socio-economically relevant applications of Cr supplementation are emerging, e.g. for senior people, intensive care units and dialysis patients, who are notoriously Cr-depleted. Also, Cr will likely be beneficial for the healthy development of premature infants, who after separation from the placenta depend on external Cr. Cr supplementation of pregnant and lactating women, as well as of babies and infants are likely to be of benefit for child development. Last but not least, Cr harbours a global ecological potential as an additive for animal feed, replacing meat- and fish meal for animal (poultry and swine) and fish aqua farming. This may help to alleviate human starvation and at the same time prevent over-fishing of oceans.     

Frequent coexistence of anti-topoisomerase I and anti-U1RNP autoantibodies in African American patients associated with mild skin involvement: a retrospective clinical study

Introduction  

The presence of anti-topoisomerase I (topo I) antibodies is a classic scleroderma (SSc) marker presumably associated with a unique clinical subset. Here the clinical association of anti-topo I was reevaluated in unselected patients seen in a rheumatology clinic setting.     

Mapping of interactions between human macrophages and Staphylococcus aureus reveals an involvement of MAP kinase signaling in the host defense

Staphylococcus aureus is a dangerous opportunistic human pathogen that causes serious invasive diseases when it reaches the bloodstream. Recent studies have shown that S. aureus is highly resistant to killing by professional phagocytes and that such cells even provide a favorable environment for intracellular survival of S. aureus. Importantly, the reciprocal interactions between phagocytes and S. aureus have remained largely elusive. Here we have employed kinase profiling to define the nature and time resolution of the human THP-1 macrophage response toward S. aureus and proteomics to identify the response of S. aureus toward macrophages. The results of these studies reveal major macrophage signaling pathways triggered by S. aureus and proteomic signatures of the responses of S. aureus to macrophages. We also identify human proteins bound to S. aureus that have potential roles in bacterial killing and internalization. Most noticeably, our observations challenge the classical concept that macrophage responses are mainly mediated through Toll-like receptor 2 and NF-κB signaling and highlight the important role of the stress-activated MAP kinase signaling in orchestrating the host defense.