Calcium/calmodulin inhibition of the Arabidopsis BRASSINOSTEROID-INSENSITIVE 1 receptor kinase provides a possible link between calcium and brassinosteroid signalling

The receptor kinase BRI1 (BRASSINOSTEROID-INSENSITIVE 1) is a key component in BR (brassinosteroid) perception and signal transduction, and has a broad impact on plant growth and development. In the present study, we demonstrate that Arabidopsis CaM (calmodulin) binds to the recombinant cytoplasmic domain of BRI1 in a Ca2+-dependent manner in vitro. In silico analysis predicted binding to Helix E of the BRI1 kinase subdomain VIa and a synthetic peptide based on this sequence interacted with Ca2+/CaM. Co-expression of CaM with the cytoplasmic domain of BRI1 in Escherichia coli strongly reduced autophosphorylation of BRI1, in particular on tyrosine residues, and also reduced the BRI1-mediated transphosphorylation of E. coli proteins on tyrosine, threonine and presumably serine residues. Several isoforms of CaM and CMLs (CaM-like proteins) were more effective (AtCaM6, AtCaM7 and AtCML8, where At is Arabidopsis thaliana) than others (AtCaM2, AtCaM4 and AtCML11) when co-expressed with BRI1 in E. coli. These results establish a novel assay for recombinant BRI1 transphosphorylation activity and collectively uncover a possible new link between Ca2+ and BR signalling

Down regulation of NM23.H1 gene inhibits cell proliferation

nm23 gene expression is strictly related to the state of cell growth. The level of its expression parallels the fraction of thymidine-incorporating cells (S-phase cells) in neoplastic mammary tissues and in the synchronously cycling fraction of MCF 1OA cells. nm23.h1 reaches a peak of expression in the S-phase, and is present at very low level during the GO/G1 phase. Two strategies are used to demonstrate the direct involvement of the nm23.h1 gene in the process of cell proliferation. The first consists of transient inhibition of nm23.h1 expression by using anti-sense oligonucleotide treatment; weak inhibitory effect on cell proliferation is observed. The second strategy involves the stable inhibition of nm23.h1 expression by transfection of MCF1OA cells with a plasmid vector expressing the human nm23.h1 anti-sense mRNA. The anti-sense-transfected cells show consistently slower proliferative activity than the control

Down-regulation of the nm23.h1 gene inhibits cell proliferation

nm23 gene expression is strictly related to the state of cell growth. The level of its expression parallels the fraction of thymidine-incorporating cells (S-phase cells) in neoplastic mammary tissues and in the synchronously cycling fraction of MCF10A cells. nm23.h1 reaches a peak of expression in the S-phase, and is present at very low level during the G(0)/G(1) phase. Two strategies are used to demonstrate the direct involvement of the nm23.h1 gene in the process of cell proliferation. The first consists of transient inhibition of nm23.h1 expression by using anti-sense oligonucleotide treatment; weak inhibitory effect on cell proliferation is observed. The second strategy involves the stable inhibition of nm23.h1 expression by transfection of MCF10A cells with a plasmid vector expressing the human nm23.h1 anti-sense mRNA. The anti-sense-transfected cells show consistently slower proliferative activity than the control. (C) 1997 Wiley-Liss, Inc