氟化物对桑蚕幼虫中肠ATP酶活性影响的电镜细胞化学定位

氟化物对桑蚕幼虫中肠ＡＴＰ酶活性影响的电镜细胞化学定位陈玉银,包焕盛,吴玉澄（浙江农业大学杭州３１００Ｎ）桑蚕Ｂｏｍｂｙｘｍｏｒｉ（Ｌ．）氟中毒导致蚕茧严重减产是目前蚕业界最为关切的问题,有关桑蚕幼虫氟中毒的机理研究也从各方面展开［１１］。已知桑蚕氟...     

Epidermal growth factor improves intestinal morphology by stimulating proliferation and differentiation of enterocytes and mTOR signaling pathway in weaning piglets

Epidermal growth factor(EGF) has been shown to improve piglet intestinal morphology and epithelial recovery. In an attempt to further understand the mechanisms behind these improvements, this study tested the hypothesis that dietary EGF may affect intestinal morphology by stimulating the proliferation and differentiation of enterocytes in weaning piglets. In piglets receiving200 μg kg–1 EGF, crypt depth and villus height increased(P0.05). Adding 400 μg kg–1 EGF increased villus height-to-crypt depth ratio(P0.05), but reduced crypt depth(P0.05). Dietary supplementation with 200 μg kg–1 EGF significantly increased the number of Ki67-positive cells(P0.01) and tended to increase the mRNA level of proliferating cell nuclear antigen(P0.10).However, this supplementation decreased the expression level of intestinal fatty acid-binding protein(P0.05). Piglets fed with400 μg kg–1 EGF had an increased mRNA level of intestinal alkaline phosphatase(P0.05). The phosphorylation of m TOR(mammalian target of rapamycin) was observed in the 200 μg kg–1 EGF group. These results suggest that dietary supplementation with a low level of EGF improved piglet intestinal morphology through stimulating the proliferation and differentiation of enterocytes, and the mTOR signaling pathway may partly be involved in this process.     

In vivo genetic ablation of the periotic mesoderm affects cell proliferation survival and differentiation in the cochlea

Tbx1 is required for ear development in humans and mice. Gene manipulation in the mouse has discovered multiple consequences of loss of function on early development of the inner ear, some of which are attributable to a cell autonomous role in maintaining cell proliferation of epithelial progenitors of the cochlear and vestibular apparata. However, ablation of the mesodermal domain of the gene also results in severe but more restricted abnormalities. Here we show that Tbx1 has a dynamic expression during late development of the ear, in particular, is expressed in the sensory epithelium of the vestibular organs but not of the cochlea. Vice versa, it is expressed in the condensed mesenchyme that surrounds the cochlea but not in the one that surrounds the vestibule. Loss of Tbx1 in the mesoderm disrupts this peri-cochlear capsule by strongly reducing the proliferation of mesenchymal cells. The organogenesis of the cochlea, which normally occurs inside the capsule, was dramatically affected in terms of growth of the organ, as well as proliferation, differentiation and survival of its epithelial cells. This model provides a striking demonstration of the essential role played by the periotic mesenchyme in the organogenesis of the cochlea.     

Neutralization of acidic residues in helix II stabilizes the folded conformation of acyl carrier protein and variably alters its function with different enzymes

Acyl carrier protein (ACP), a small protein essential for bacterial growth and pathogenesis, interacts with diverse enzymes during the biosynthesis of fatty acids, phospholipids, and other specialized products such as lipid A. NMR and hydrodynamic studies have previously shown that divalent cations stabilize native helical ACP conformation by binding to conserved acidic residues at two sites (A and B) at either end of the "recognition" helix II. To examine the roles of these amino acids in ACP structure and function, site-directed mutagenesis was used to replace individual site A (Asp-30, Asp-35, Asp-38) and site B (Glu-47, Glu-53, Asp-56) residues in recombinant Vibrio harveyi ACP with the corresponding amides, along with combined mutations at each site (SA, SB) or both sites (SA/SB). Like native V. harveyi ACP, all individual mutants were unfolded at neutral pH but adopted a helical conformation in the presence of millimolar Mg(2+) or upon fatty acylation. Mg(2+) binding to sites A or B independently stabilized native ACP conformation, whereas mutant SA/SB was folded in the absence of Mg(2+), suggesting that charge neutralization is largely responsible for ACP stabilization by divalent cations. Asp-35 in site A was critical for holo-ACP synthase activity, while acyl-ACP synthetase and UDP-N-acetylglucosamine acyltransferase (LpxA) activities were more affected by mutations in site B. Both sites were required for fatty acid synthase activity. Overall, our results indicate that divalent cation binding site mutations have predicted effects on ACP conformation but unpredicted and variable consequences on ACP function with different enzymes.     

The renewal and differentiation of Isl1+ cardiovascular progenitors are controlled by a Wnt/beta-catenin pathway

Isl1(+) cardiovascular progenitors and their downstream progeny play a pivotal role in cardiogenesis and lineage diversification of the heart. The mechanisms that control their renewal and differentiation are largely unknown. Herein, we show that the Wnt/beta-catenin pathway is a major component by which cardiac mesenchymal cells modulate the prespecification, renewal, and differentiation of isl1(+) cardiovascular progenitors. This microenvironment can be reconstituted by a Wnt3a-secreting feeder layer with ES cell-derived, embryonic, and postnatal isl1(+) cardiovascular progenitors. In vivo activation of beta-catenin signaling in isl1(+) progenitors of the secondary heart field leads to their massive accumulation, inhibition of differentiation, and outflow tract (OFT) morphogenic defects. In addition, the mitosis rate in OFT myocytes is significantly reduced following beta-catenin deletion in isl1(+) precursors. Agents that manipulate Wnt signals can markedly expand isl1(+) progenitors from human neonatal hearts, a key advance toward the cloning of human isl1(+) heart progenitors.     

Identification of a key residue in the conformational stability of acyl carrier protein

Conformational flexibility of acyl carrier protein (ACP) is important for its ability to interact with multiple enzymes in bacterial fatty acid metabolism. We have recently shown that, unlike the prototypical ACP from Escherichia coli, the more acidic Vibrio harveyi ACP is largely unfolded at physiological pH. Mutations D18K, A75H and A75H/D18K were made in recombinant V. harveyi ACP (rACP) to determine the importance of basic residues Lys-18 and His-75 in maintaining the native conformation of E. coli ACP. Both D18K and A75H ACPs were fatty acylated by acyl-ACP synthetase, showing that neither mutation grossly alters tertiary structure. Circular dichroism (CD) indicated that rACP refolded upon addition of MgCl(2) at 100-fold lower concentrations (<1 mM) than KCl, suggesting that divalent cations stabilize rACP by interaction at specific sites. Surprisingly, mutants A75H and A75H/D18K exhibited native-like conformation in the absence of MgCl(2), while the D18K mutant was comparable to rACP. Moreover, the alpha-helical content of A75H, A75H/D18K and E. coli ACPs was more sensitive than that of rACP or D18K ACP to modification by the histidine-selective reagent diethylpyrocarbonate. Together, these results suggest that the partial positive charge of His-75 may be important in maintaining the conformational stability of E. coli ACP at a neutral pH.     

Genetic pathways to mammalian heart development: Recent progress from manipulation of the mouse genome

Mammalian heart development requires multiple genetic networks, only some of which are becoming known in all their complexity. Substantial new information has become available thanks to an expanding toolkit that offers more and more mouse gene manipulation options, and that is taking the mouse closer to more powerful invertebrate genetic models. We review examples of recent data with a cardiac-lineage-based view of heart development, especially outflow tract and right ventricle. The medical significance of these studies is not only relevant to congenital heart disease, but also to the biology of cardiac cell regeneration.     

Mapping the protein interaction network in methicillin-resistant Staphylococcus aureus

Mortality attributable to infection with methicillin-resistant Staphylococcus aureus (MRSA) has now overtaken the death rate for AIDS in the United States, and advances in research are urgently needed to address this challenge. We report the results of the systematic identification of protein-protein interactions for the hospital-acquired strain MRSA-252. Using a high-throughput pull-down strategy combined with quantitative proteomics to distinguish specific from nonspecific interactors, we identified 13,219 interactions involving 608 MRSA proteins. Consecutive analyses revealed that this protein interaction network (PIN) exhibits scale-free organization with the characteristic presence of highly connected hub proteins. When clinical and experimental antimicrobial targets were queried in the network, they were generally found to occupy peripheral positions in the PIN with relatively few interacting partners. In contrast, the hub proteins identified in this MRSA PIN that are essential for network integrity and stability have largely been overlooked as drug targets. Thus, this empirical MRSA-252 PIN provides a rich source for identifying critical proteins essential for network stability, many of which can be considered as prospective antimicrobial drug targets.