Novel 19 F-MRS ß-galactosidase reporter molecules incorporated nitrogen mustard analogues.

In this study, we propose a novel molecular platform-integrated fluorinated antitumor nitrogen mustards for 19 F-MRS assay of ß-galactosidase (ß-gal) activity. Following this idea, we have designed, synthesized, and characterized 2-fluoro-4-[bis(2'-chloroethyl)amino]phenyl ß-D-galactopyranoside 5, 2-fluoro-4-{bis[2'-O-(ß-D-galactopyranosyl)ethyl]amino}phenyl ß-D-galactopyranoside 8, 2-fluoro-4-{bis[[1″-(ß-D-galactopyranosyl)-1″, 2″, 3″-triazol-4″-yl]methyl] amino}phenyl ß-D-galactopyranoside 14 and 2-fluoro-4-{bis[[1″-(ß-D-glucopyranosyl)-1″, 2″, 3″-triazol-4″-yl]methyl]amino}phenyl ß-D-galactopyranoside 15 through glycosylation and click reaction strategies, and their structures were confirmed by NMR and HRMS or elemental analysis data. Among them, 2-fluoro-4-[bis(2'-chloroethyl)amino]phenyl ß-D-galacto-pyranoside 5 was found very sensitive to ß-gal (E801A) in PBS at 37°C with big ΔδF response. Here, we demonstrated the feasibility of this platform for assessing ß-gal activity in solution, and in vitro with lacZ-transfected human MCF7 breast and PC3 prostate tumor cells, by the characterization of ß-gal-responsive 19 F-chemical shift changes ΔδF and hydrolytic kinetics.

Activation of the NaCl- and drought-induced RD29A and RD29B promoters by constitutively active Arabidopsis MAPKK or MAPK proteins.

Mitogen-activated protein (MAP) kinases mediate cellular responses to a wide variety of stimuli. Activation of a MAP kinase (MAPK) occurs after phosphorylation by an upstream MAP kinase kinase (MAPKK). The Arabidopsis thaliana genome encodes 10 MKKs, but few of these have been shown directly to activate any of the 20 Arabidopsis MAPKs (AtMPKs) and NaCl-, drought- or abscisic acid (ABA)-induced genes RD29A or RD29B. We have constructed the constitutively activated form for nine of the 10 AtMKK proteins, and tested their ability to activate the RD29A and RD29B promoters and also checked the ability of the nine activated AtMKK proteins to phosphorylate 11 of the AtMPK proteins in transient assays. The results show that three proteins, AtMKK1, AtMKK2 and AtMKK3, could activate the RD29A promoter, while these three and two additional AtMKK6/8 proteins could activate the RD29B promoter. Four other proteins, AtMKK7/AtMKK9 and AtMKK4/AtMKK5, can cause hypersensitive response (HR) in tobacco leaves using transient analysis. The activation of the RD29A promoter correlated with four uniquely activated AtMPK proteins. A novel method of activating AtMPK proteins by fusion to a cis-acting mutant of a human MAPK kinase MEK1 was used to confirm that specific members of the AtMPK gene family can activate the RD29A stress pathway.