Synthesis and antiproliferative activity of C-homo-lactam derivatives of 7-deoxycholic acid

Using deoxycholic acid as starting materials, a series of 12a-aza-C-homo-12-one 7-deoxycholic acid derivatives were synthesized The antiproliferative activity of the synthesized compounds against some carcinoma cell lines was investigated. The results showed that some 12-oxy-12a-aza-C-homo-7-deoxycholic acid derivatives displayed distinct cytotoxicity to HeLa (human cervical carcinoma) and Tu 686 (laryngocarcinoma) tumor cell lines. In particular, the IC₅₀ values of the compounds 6 and 7 against Tu 686 cells are 16.7 and 19.8 μM/L respectively. The information obtained from the studies may be useful for the design of novel chemotherapeutic drugs.     

Methyl-β-cyclodextrin potentiates the BITC-induced anti-cancer effect through modulation of the Akt phosphorylation in human colorectal cancer cells

ABSTRACT

Methyl-β-cyclodextrin (MβCD) is an effective agent for the removal of plasma membrane cholesterol. In this study, we investigated the modulating effects of MβCD on the antiproliferation induced by benzyl isothiocyanate (BITC), an ITC compound mainly derived from papaya seeds. We confirmed that MβCD dose-dependently increased the cholesterol level in the medium, possibly through its removal from the plasma membrane of human colorectal cancer cells. The pretreatment with a non-toxic concentration (2.5 mM) of MβCD significantly enhanced the BITC-induced cytotoxicity and apoptosis induction, which was counteracted by the cholesterol supplementation. Although BITC activated the phosphoinositide 3-kinase (PI3K)/Akt pathway, MβCD dose-dependently inhibited the phosphorylation level of Akt. On the contrary, the treatment of MβCD enhanced the phosphorylation of mitogen activated protein kinases, but did not potentiate their BITC-induced phosphorylation. These results suggested that MβCD might potentiate the BITC-induced anti-cancer by cholesterol depletion and thus inhibition of the PI3K/Akt-dependent survival pathway.

Abbreviations: CDs: cyclodextrins; MβCD: methyl-β-cyclodextrin; ITCs: isothiocyanates; BITC: benzyl isothiocyanate; PI3K: phosphoinositide 3-kinase; PDK1: phosphoinositide-dependent kinase-1; MAPK: mitogen activated protein kinase; ERK1/2: extracellular signal-regulated kinase1/2; JNK: c-Jun N-terminal kinase; PI: propidium iodide; FBS: fatal bovine serum; TLC: thin-layer chromatography; PBS(-): phosphate-buffered saline without calcium and magnesium; MEK: MAPK/ERK kinase; PIP2: phosphatidylinositol-4,5-bisphosphate; PIP3: phosphatidylinositol-3,4,5-trisphosphate     

Angelman Syndrome Protein Ube3a Regulates Synaptic Growth and Endocytosis by Inhibiting BMP Signaling in Drosophila

Altered expression of the E3 ubiquitin ligase UBE3A, which is involved in protein degradation through the proteasome-mediated pathway, is associated with neurodevelopmental and behavioral defects observed in Angelman syndrome (AS) and autism. However, little is known about the neuronal function of UBE3A and the pathogenesis of UBE3A-associated disorders. To understand the in vivo function of UBE3A in the nervous system, we generated multiple mutations of ube3a, the Drosophila ortholog of UBE3A. We found a significantly increased number of total boutons and satellite boutons in conjunction with compromised endocytosis in the neuromuscular junctions (NMJs) of ube3a mutants compared to the wild type. Genetic and biochemical analysis showed upregulation of bone morphogenetic protein (BMP) signaling in the nervous system of ube3a mutants. An immunochemical study revealed a specific increase in the protein level of Thickveins (Tkv), a type I BMP receptor, but not other BMP receptors Wishful thinking (Wit) and Saxophone (Sax), in ube3a mutants. Ube3a was associated with and specifically ubiquitinated lysine 227 within the cytoplasmic tail of Tkv, and promoted its proteasomal degradation in Schneider 2 cells. Negative regulation of Tkv by Ube3a was conserved in mammalian cells. These results reveal a critical role for Ube3a in regulating NMJ synapse development by repressing BMP signaling. This study sheds new light onto the neuronal functions of UBE3A and provides novel perspectives for understanding the pathogenesis of UBE3A-associated disorders.     

Drosophila S6 Kinase Like Inhibits Neuromuscular Junction Growth by Downregulating the BMP Receptor Thickveins

Synaptic connections must be precisely controlled to ensure proper neural circuit formation. In Drosophila melanogaster, bone morphogenetic protein (BMP) promotes growth of the neuromuscular junction (NMJ) by binding and activating the BMP ligand receptors wishful thinking (Wit) and thickveins (Tkv) expressed in motor neurons. We report here that an evolutionally conserved, previously uncharacterized member of the S6 kinase (S6K) family S6K like (S6KL) acts as a negative regulator of BMP signaling. S6KL null mutants were viable and fertile but exhibited more satellite boutons, fewer and larger synaptic vesicles, larger spontaneous miniature excitatory junctional potential (mEJP) amplitudes, and reduced synaptic endocytosis at the NMJ terminals. Reducing the gene dose by half of tkv in S6KL mutant background reversed the NMJ overgrowth phenotype. The NMJ phenotypes of S6KL mutants were accompanied by an elevated level of Tkv protein and phosphorylated Mad, an effector of the BMP signaling pathway, in the nervous system. In addition, Tkv physically interacted with S6KL in cultured S2 cells. Furthermore, knockdown of S6KL enhanced Tkv expression, while S6KL overexpression downregulated Tkv in cultured S2 cells. This latter effect was blocked by the proteasome inhibitor MG132. Our results together demonstrate for the first time that S6KL regulates synaptic development and function by facilitating proteasomal degradation of the BMP receptor Tkv.     

Molecular phylogenetic relationships of China Seas groupers based on cytochrome <Emphasis Type="Italic">b</Emphasis> gene fragment sequences

The classification and evolutionary relationships are important issues in the study of the groupers. Cytochrome b gene fragment of twenty-eight grouper species within six genera of subfamily Epinephelinae was amplified using PCR techniques and the sequences were analyzed to derive the phylogenetic relationships of the groupers from the China Seas. Genetic information indexes, including Kimura-2 parameter genetic distance and T _S/T _V ratios, were generated by using a variety of biology softwares. With Niphon spinosus, Pagrus major and Pagrus auriga as the designated outgroups, phylogenetic trees, which invoke additional homologous sequences of other Epinephelus fishes from GenBank, were constructed based on the neighbor-joining (NJ), maximum-parsimony (MP), maximum-likelihood (ML) and minimum-evolution (ME) methods. Several conclusions were drawn from the DNA sequences analysis: (1) genus Plectropomus, which was early diverged, is the most primitive group in the subfamily Epinephelinae; (2) genus Variola is more closely related to genus Cephalopolis than the other four genera; (3) genus Cephalopolis is a monophyletic group and more primitive than genus Epinephelus; (4) Promicrops lanceolatus and Cromileptes altivelis should be included in genus Epinephelus; (5) there exist two sister groups in genus Epinephelus.     

红汁乳菇醇提物对水稻、稗草的化感影响

研究了红汁乳菇子实体乙醇抽提物对水稻、稗草的化感影响。用不同浓度的红汁乳菇子实体乙醇抽提物处理水稻和稗草种子 ,测量处理后不同时间的水稻和稗草幼苗的根长和苗高 ,按照 Willamson,G.B.的方法计算化感作用效应 (RI)。结果表明 :红汁乳菇乙醇抽提物对稗草幼苗的生长有显著的抑制作用 ,能够导致稗草叶片发黄 ,缩短稗草的根和苗 ,甚至延迟部分种子的萌发。其中 8.5 mg/ ml的乙醇抽提物处理稗草种子后 3d、8d,对根长的化感作用效应分别为 - 0 .98、- 0 .5 8;对苗高的化感作用效应分别为 - 0 .5 0、- 0 .5 1。 6 .5 m g/ ml和 10 .5 mg/ ml处理组的结果与 8.5 mg/ ml处理组的结果在 p=0 .0 5水平上差异不显著。对水稻幼苗的生长表现出一定的调节作用 ,在较低浓度下 (≤ 4 .5 0 mg/ m l)对水稻幼苗有一定促进作用 ,如 4 .5 0 m g/ ml处理水稻种子后 3d、8d,对苗高的化感作用效应分别为 0 .2 5、0 .10 ;在较高浓度下 (如≥ 6 .5 0 mg/ ml)对水稻根长和苗高均为抑制作用 ,6 .5 0 mg/ ml时抑制作用最强 ,处理水稻种子后 3d、8d对幼苗的根长的化感作用效应分别为 - 0 .93、- 0 .4 8;对苗高的化感作用效应分别为 - 0 .5 0、- 0 .2 2 ,在 p=0 .0 5水平上与其它处理组差异显著     

A nonstructural protein encoded by a rice reovirus induces an incomplete autophagy to promote viral spread in insect vectors

Viruses can hijack autophagosomes as the nonlytic release vehicles in cultured host cells. However, how autophagosome-mediated viral spread occurs in infected host tissues or organs in vivo remains poorly understood. Here, we report that an important rice reovirus, rice gall dwarf virus (RGDV) hijacks autophagosomes to traverse multiple insect membrane barriers in the midgut and salivary gland of leafhopper vector to enhance viral spread. Such virus-containing double-membraned autophagosomes are prevented from degradation, resulting in increased viral propagation. Mechanistically, viral nonstructural protein Pns11 induces autophagy and embeds itself in the autophagosome membranes. The autophagy-related protein 5 (ATG5)-ATG12 conjugation is essential for initial autophagosome membrane biogenesis. RGDV Pns11 specifically interacts with ATG5, both in vitro and in vivo. Silencing of ATG5 or Pns11 expression suppresses ATG8 lipidation, autophagosome formation, and efficient viral propagation. Thus, Pns11 could directly recruit ATG5-ATG12 conjugation to induce the formation of autophagosomes, facilitating viral spread within the insect bodies. Furthermore, Pns11 potentially blocks autophagosome degradation by directly targeting and mediating the reduced expression of N-glycosylated Lamp1 on lysosomal membranes. Taken together, these results highlight how RGDV remodels autophagosomes to benefit viral propagation in its insect vector.