Laboratory of Molecular Biology

About: Laboratory of Molecular Biology is a facility organization based out in Cambridge, Cambridgeshire, United Kingdom. It is known for research contribution in the topics: Gene & RNA. The organization has 19395 authors who have published 24236 publications receiving 2101480 citations.

The binding site for C1q on IgG

Abstract: In humoral defence, pathogens are cleared by antibodies acting as adaptor molecules: they bind to antigen and trigger clearance mechanisms such as phagocytosis, antibody-dependent cell-mediated cytotoxicity and complement lysis. The first step in the complement cascade is the binding of C1q to the antibody. There are six heads on C1q, connected by collagen-like stems to a central stalk, and the isolated heads bind to the Fc portion of antibody rather weakly, with an affinity of 100 microM (ref. 3). Binding of antibody to multiple epitopes on an antigenic surface, aggregates the antibody and this facilitates the binding of several C1q heads, leading to an enhanced affinity of about 10 nM (ref. 1). Within the Fc portion of the antibody, C1q binds to the CH2 domain. The interaction is sensitive to ionic strength, and appears to be highly conserved throughout evolution as C1q reacts with IgG from different species (for example see ref. 8). By systematically altering surface residues in the mouse IgG2b isotype, we have localized the binding site for C1q to three side chains, Glu 318, Lys 320 and Lys 322. These residues are relatively conserved in other antibody isotypes, and a peptide mimic of this sequence is able to inhibit complement lysis. We propose that this sequence motif forms a common core in the interactions of IgG and C1q.

Hypoxia promotes expansion of the CD133-positive glioma stem cells through activation of HIF-1alpha.

Abstract: Hypoxia contributes to the progression of a variety of cancers by activating adaptive transcriptional programs that promote cell survival, motility and tumor angiogenesis. Although the importance of hypoxia and subsequent hypoxia-inducible factor-1alpha (HIF-1alpha) activation in tumor angiogenesis is well known, their role in the regulation of glioma-derived stem cells is unclear. In this study, we show that hypoxia (1% oxygen) promotes the self-renewal capacity of CD133-positive human glioma-derived cancer stem cells (CSCs). Propagation of the glioma-derived CSCs in a hypoxic environment also led to the expansion of cells bearing CXCR4 (CD184), CD44(low) and A2B5 surface markers. The enhanced self-renewal activity of the CD133-positive CSCs in hypoxia was preceded by upregulation of HIF-1alpha. Knockdown of HIF-1alpha abrogated the hypoxia-mediated CD133-positive CSC expansion. Inhibition of the phosphatidylinositol 3-kinase(PI3K)-Akt or ERK1/2 pathway reduced the hypoxia-driven CD133 expansion, suggesting that these signaling cascades may modulate the hypoxic response. Finally, CSCs propagated at hypoxia robustly retained the undifferentiated phenotype, whereas CSCs cultured at normoxia did not. These results suggest that response to hypoxia by CSCs involves the activation of HIF-1alpha to enhance the self-renewal activity of CD133-positive cells and to inhibit the induction of CSC differentiation. This study illustrates the importance of the tumor microenvironment in determining cellular behavior.

Antagonist binding profiles of five cloned human muscarinic receptor subtypes.

Abstract: A variety of muscarinic antagonists are currently used as tools to pharmacologically subclassify muscarinic receptors into M1, M2 and M3 subtypes. In the present study, we have determined the affinity profiles of several of these antagonists at five cloned human muscarinic receptors (m1-m5) stably expressed in Chinese hamster ovary cells (CHO-K1). At all five receptors, the (R)-enantiomers of trihexyphenidyl and hexbutinol displayed considerably higher affinities (up to 525-fold) than their corresponding (S)-isomers. The stereoselectivity ratios [inhibition constant(S)/inhibition constant(R)] for both pairs of enantiomers were lowest at m2 receptors, suggesting that less stringent configurational demands are made by this receptor subtype. The "M1-selective" antagonist (R)-trihexyphenidyl displayed high affinities for m1 and m4 receptors. The "M2-selective" antagonists himbacine, (+-)-5,11-dihydro-11- ([(2-[(dipropylamino)methyl]-1- piperidinyl)ethyl)amino]carbonyl)-6H-pyrido(2,3-b)(1,4)benzodiazepine-6- one (AF-DX 384), 11-[4-[4-(diethylamino)butyl]-1-piperidinyl)acetyl)-5,11- dihydro-6H-pyrido(2,3-b) (1,4)benzodiazepine-6-one (AQ-RA 741) and (+)-(11-[2-[(diethylamino) methyl]-1-piperidinyl)acetyl)-5,11-di-hydro-6H-pyrido(2,3-b)(1,4) benzodiazepine-6-one [AF-DX 250; the (+)-enantiomer of AF-DX 116] exhibited high affinities for m2 and m4, intermediate affinities for m1 and m3 and low affinities for m5 receptors. This selectivity profile was most prominent for AQ-RA 741, which displayed 195- and 129-fold higher affinities for m2 and m4 receptors than for m5 receptors.(ABSTRACT TRUNCATED AT 250 WORDS)