RESUMEN
Cdc42 GTPase is required for polarization in eukaryotic cells, but its spatial regulation is poorly understood. In Schizosaccharomyces pombe, Cdc42p is activated by Scd1p and Gef1p, two guanine-nucleotide exchange factors. Two-hybrid screening identified Hob3p as a Gef1p binding partner. Hob3p is a BAR domain-containing protein ortholog of human Bin3. Hob3p also interacts directly with Cdc42p independently of Gef1p. Hob3p, Cdc42p and Gef1p form a complex, and Hob3p facilitates Gef1p-Cdc42p interaction and activation. Hob3p forms a ring in the division area, similar to that of Gef1p. This localization requires actin polymerization and Cdc15p but is independent of the septation initiation network. Hob3p is required for the concentration of Cdc42p to the division area. The actomyosin ring contraction is slower in hob3Delta than in wild-type cells, and this contributes to its cytokinesis defect. Moreover, this report extends previous evidence that human Bin3 suppresses the cytokinesis phenotype of hob3Delta cells, showing that Bin3 can partially recover the GTP-Cdc42p level and its localization. These results suggest that Hob3p is required to recruit and activate Cdc42p at the cell division site and that this function might be conserved in other eukaryotes.
Asunto(s)
Proteínas Portadoras/metabolismo , Citocinesis , Proteínas de Microfilamentos/metabolismo , Proteínas de Schizosaccharomyces pombe/metabolismo , Schizosaccharomyces/citología , Homología de Secuencia de Aminoácido , Proteína de Unión al GTP cdc42/metabolismo , Actinas/metabolismo , Actomiosina/metabolismo , Polaridad Celular , Activación Enzimática , Unión Proteica , Transporte de ProteínasRESUMEN
Electrospray ionization mass spectrometry (ESI-MS) and tandem mass spectrometry (ESI-MS/MS) have been used to differentiate the 2- and 4-methylpyridyl isomers of free-base and metallated cationic beta-vinylpyridylporphyrins. The analysis by ESI-MS/MS of the deuterated analogs and semiempirical calculations of structural and electronic parameters were also undertaken. The two free-base isomers are easily differentiated by ESI-MS/MS but the presence of a metallic center renders differentiation of the metallated isomers less effective. The data acquired show that of all the studied compounds, the free-base 2-methylpyridyl isomer, which was operative in the in vitro photoinactivation of Herpes simples virus, has a different gas-phase behavior. Local distortion of the macrocycle due to the presence of the beta-vinylpyridyl substituent occurs for all the compounds, but a different electron density distribution can account for the observed gas-phase behavior of this potential virus photoinactivator.
Asunto(s)
Antivirales/química , Porfirinas/química , Espectrometría de Masa por Ionización de Electrospray/métodos , Compuestos de Vinilo/química , Antivirales/análisis , Cationes/química , Isomerismo , Fotoquímica , Porfirinas/análisis , Compuestos de Vinilo/análisis , Inactivación de VirusRESUMEN
Neutral and cationic pyrrolidine-fused chlorins and isobacteriochlorins derived from meso-tetrakis(pentafluorophenyl)porphyrin undergo cycloreversion reactions in the gas phase, either when desorbed from a liquid matrix by ion bombardment or when electrosprayed. Cycloreversion occurs through loss of either neutral or charged moieties, with and without hydrogen and methyl radical migration, and both as high- and low-energy collision processes. For the doubly charged isobacteriochlorin, one-electron reduction with methyl loss occurs under ion bombardment and electrospray, through hypervalent pyrrolidinium radical formation.
