Cofilin activity during insulin-like growth factor I-stimulated neuroblastoma cell motility

Insulin-like growth factor I (IGF-I) is a potent stimulator of neuroblastoma cell motility. Cell motility requires lamellipodium extension at the leading edge of the cell through organized actin polymerization, and IGF-I stimulates lamellipodial elaboration in human neuroblastoma cells. Rac is a Rho GTPase that stimulates lamellipodial formation via the regulation of actin polymerization. In this study, we show that IGF-I-stimulated phosphatidylinositol 3-kinase (PI-3K) activity promotes rac activation and subsequent activation of the down- stream effectors LIM kinase and cofilin. Overexpression of wild-type LIM kinase and wild-type Xenopus ADF/cofilin (XAC) suppresses IGF-I-stimulated motility in SH-SY5Y cells, while expression of dominant negative LIM kinase and constitutively active XAC increases SH-SY5Y motility in the absence of IGF-I stimulation. These results suggest that regulation by cofilin of actin depolymerization is important in the process of neuroblastoma cell motility, and IGF-I regulates cofilin activity in part through PI-3K, rac, and LIM kinase.Received 18 October 2004; received after revision 3 December 2004; accepted 16 December 2004     

Tyrosinase inhibitors from natural and synthetic sources: structure, inhibition mechanism and perspective for the future

Tyrosinase is known to be a key enzyme in melanin biosynthesis, involved in determining the color of mammalian skin and hair. Various dermatological disorders, such as melasma, age spots and sites of actinic damage, arise from the accumulation of an excessive level of epidermal pigmentation. In addition, unfavorable enzymatic browning of plant-derived foods by tyrosinase causes a decrease in nutritional quality and economic loss of food products. The inadequacy of current conventional techniques to prevent tyrosinase action encourages us to seek new potent tyrosinase inhibitors. This article overviews the various inhibitors obtained from natural and synthetic sources with their industrial importance.Received 9 February 2005; received after revision 4 April 2005; accepted 14 April 2005     

Licensing of natural killer cells by host major histocompatibility complex class I molecules

Self versus non-self discrimination is a central theme in biology from plants to vertebrates, and is particularly relevant for lymphocytes that express receptors capable of recognizing self-tissues and foreign invaders. Comprising the third largest lymphocyte population, natural killer (NK) cells recognize and kill cellular targets and produce pro-inflammatory cytokines. These potentially self-destructive effector functions can be controlled by inhibitory receptors for the polymorphic major histocompatibility complex (MHC) class I molecules that are ubiquitously expressed on target cells. However, inhibitory receptors are not uniformly expressed on NK cells, and are germline-encoded by a set of polymorphic genes that segregate independently from MHC genes. Therefore, how NK-cell self-tolerance arises in vivo is poorly understood. Here we demonstrate that NK cells acquire functional competence through 'licensing' by self-MHC molecules. Licensing involves a positive role for MHC-specific inhibitory receptors and requires the cytoplasmic inhibitory motif originally identified in effector responses. This process results in two types of self-tolerant NK cells--licensed or unlicensed--and may provide new insights for exploiting NK cells in immunotherapy. This self-tolerance mechanism may be more broadly applicable within the vertebrate immune system because related germline-encoded inhibitory receptors are widely expressed on other immune cells.     

Recognition of bacterial glycosphingolipids by natural killer T cells

Natural killer T (NKT) cells constitute a highly conserved T lymphocyte subpopulation that has the potential to regulate many types of immune responses through the rapid secretion of cytokines. NKT cells recognize glycolipids presented by CD1d, a class I-like antigen-presenting molecule. They have an invariant T-cell antigen receptor (TCR) alpha-chain, but whether this invariant TCR recognizes microbial antigens is still controversial. Here we show that most mouse and human NKT cells recognize glycosphingolipids from Sphingomonas, Gram-negative bacteria that do not contain lipopolysaccharide. NKT cells are activated in vivo after exposure to these bacterial antigens or bacteria, and mice that lack NKT cells have a marked defect in the clearance of Sphingomonas from the liver. These data suggest that NKT cells are T lymphocytes that provide an innate-type immune response to certain microorganisms through recognition by their antigen receptor, and that they might be useful in providing protection from bacteria that cannot be detected by pattern recognition receptors such as Toll-like receptor 4.     

Recognition of transmembrane helices by the endoplasmic reticulum translocon

Membrane proteins depend on complex translocation machineries for insertion into target membranes. Although it has long been known that an abundance of nonpolar residues in transmembrane helices is the principal criterion for membrane insertion, the specific sequence-coding for transmembrane helices has not been identified. By challenging the endoplasmic reticulum Sec61 translocon with an extensive set of designed polypeptide segments, we have determined the basic features of this code, including a 'biological' hydrophobicity scale. We find that membrane insertion depends strongly on the position of polar residues within transmembrane segments, adding a new dimension to the problem of predicting transmembrane helices from amino acid sequences. Our results indicate that direct protein-lipid interactions are critical during translocon-mediated membrane insertion.     

Somatic misexpression of germline P granules and enhanced RNA interference in retinoblastoma pathway mutants

Caenorhabditis elegans homologues of the retinoblastoma (Rb) tumour suppressor complex specify cell lineage during development. Here we show that mutations in Rb pathway components enhance RNA interference (RNAi) and cause somatic cells to express genes and elaborate perinuclear structures normally limited to germline-specific P granules. Furthermore, particular gene inactivations that disrupt RNAi reverse the cell lineage transformations of Rb pathway mutants. These findings suggest that mutations in Rb pathway components cause cells to revert to patterns of gene expression normally restricted to germ cells. Rb may act by a similar mechanism to transform mammalian cells.     

Crystal structure of the intensely sweet protein monellin

Two unusual proteins, discovered in African berries, possess the interesting property of having a very high specificity for the sweet receptors. These proteins, monellin and thaumatin, are approximately 100,000 times sweeter than sugar on a molar basis and several thousand times sweeter on a weight basis. Neither contains carbohydrates or modified amino acids. Several interesting observations have been made about the two proteins: native conformations are important for the sweet taste, although both proteins are intensely sweet, there are no statistically significant sequence similarities between them; and despite the absence of sequence similarity, antibodies against thaumatin compete for monellin (as well as many other sweet compounds, but not for chemically modified non-sweet monellin) and vice versa. To understand the structural basis of these observations we determined the crystal structure of thaumatin, and report here the structure of monellin at 3 A resolution. Monellin consists of two peptide chains, the A chain of 44 residues and the B chain of 50 residues. We find no similarity between the backbone structure of monellin and that of thaumatin.