Physical and genetic interactions link the yeast protein Zds1p with mRNA nuclear export

Eukaryotic gene expression requires the export of mRNA from the nucleus to the cytoplasm. The DEAD box protein Dbp5p is an essential export factor conserved from yeast to man. A fraction of Dbp5p forms a complex with nucleoporins of the cytoplasmic filaments of the nuclear pore complex. Gfd1p was identified originally as a multicopy suppressor of the rat8-2 ts allele of DBP5. Here we reported that Dbp5p and Gfd1p interact with Zds1p, a protein previously identified as a multicopy suppressor in several yeast genetic screens. By using the two-hybrid system, we showed that Zds1p interacts in vivo with both Gfd1p and Dbp5p. In vitro binding experiments revealed that Gfd1p and Dbp5p bind directly to the C-terminal part of Zds1p. In addition, ZDS1 interacted genetically with mutant alleles of genes encoding key factors in mRNA export, including DBP5 and MEX67. Furthermore, deletion of ZDS1 or of both ZDS1 and the closely related ZDS2 exacerbated the poly(A)+ export defects shown by dbp5-2 and mex67-5 mutants. We proposed that Zds1p associates with the complex formed by Dbp5p, Gfd1p, and nucleoporins at the cytosolic fibrils of the nuclear pore complex and is required for optimal mRNA export.     

转录因子对B细胞发育的调控作用

Ｂ细胞同其他血细胞一样,也是由多能造血干细胞发育为祖细胞,再发育成 末成熟细胞的。发育过程中,向Ｂ细胞发育的前 细胞逐渐推动向其他谱系发育的潜能,Ｂ细胞谱系特征性基因开始表达。近来的研究表明,Ｂ细胞发育过程中有多种转录因子参与,它们调节多种重要基因的转录,保证Ｂ细胞的正常发育。本文综述了与Ｂ细胞发育相关转录因子的最新研究进展。     

Dual role of BKI1 and 14-3-3 s in brassinosteroid signaling to link receptor with transcription factors

The plasma membrane-localized plant steroid hormone receptor, BRASSINOSTEROID INSENSITIVE 1 (BRI1), is quiescent in the absence of steroids, largely due to a negative regulator, BRI1 KINASE INHIBITOR 1 (BKI1). Here, we report that the steroid-induced, plasma membrane-dissociated and phosphorylated BKI1 also plays positive roles in BR signaling by interacting with a subset of 14-3-3 proteins. The cytosolic fraction of BKI1 carboxyl terminal region enhances BR signaling. Mutations of two serine residues in this region lead to reduced phosphorylation by the BRI1 kinase and constitutive plasma membrane localization. The 14-3-3 proteins can interact with the phosphorylated BKI1 through a motif that contains the two phosphorylation sites to release inhibition of BRI1 by BKI1. Meanwhile, the cytosolic BKI1 antagonizes the 14-3-3?s and enhances accumulation of BRI1 EMS SUPPRESSOR 1 (BES1)/BRASSINAZOLE RESISTANT 1 (BZR1) in the nucleus to regulate BR-responses.     

Cellular interactions with the extracellular matrix are coupled to diverse transmembrane signaling pathways

Extracellular matrix (ECM) glycoproteins such as laminin, fibronectin, or collagen IV play a major role in cell behavior regulation. The molecular mechanisms taking place at the interface between the ECM and the cell surface are now rather well defined; however, very little is known about intracellular signals induced by these interactions. In order to get insights into the transduction pathways involved in cell-ECM interactions we have investigated the effects of several intracellular kinase inhibitors. Calmodulin-dependent kinase inhibitors, W-7 and sphingosine, have negative effects on cell-matrix interactions. They inhibit adhesion of several cell lines to laminin (IC50 = 4-10 microM), fibronectin and collagen IV (IC50 = 7-25 microM). The effects are immediate, reversible, and also cell specific, certain combinations of cell line-substrate being irresponsive to these inhibitors. In contrast, two inhibitors, H-7 and staurosporine, for which protein kinase C is a common target, increase two- to fourfold the attachment of HT1080, OVCAR-4, and B16F10 cells to laminin but not to fibronectin. Another inhibitor, HA-1004, known to inhibit protein kinase A at low concentrations, has an activating effect only at high concentration (> 200 microM) when it becomes an inhibitor of protein kinase C. These inhibitors are without effect on RuGli and Saos-2 cell adhesion on the three substrates. Altogether these results suggest that calmodulin-dependent kinases and protein kinase C could be separately involved in ECM-induced cellular responses. However, the effects of kinase inhibitors are substrate-specific and cell type-specific, suggesting that the intracellular signals induced by the extracellular matrix vary with the nature of integrin involved in signal transmission.     

A genetic library screen for signaling proteins that interact with phosphorylated T cell costimulatory receptors

In the past decade, the fundamental importance and therapeutic potential of costimulatory signals for lymphocyte activation have spurred a large amount of work in immunology, infection, cancer, autoimmune diseases, etc. However, the mechanisms behind T cell costimulation remain unclear, partly due to the lack of suitable techniques. There is an urgent need for functional genomic research to develop comprehensive approaches to direct identification of protein-protein interactions that are dependent on the posttranslational modification of one component of the complex, particularly in the field of T cell immunology. Using inducible costimulator (ICOS) as a model, we failed to find any proteins that associated with the cytoplasmic tail of ICOS by the yeast two-hybrid approach. Therefore, we have developed a new yeast three-hybrid system that facilitates the rapid screening of cDNA libraries to find signaling molecules that interact with phosphorylated T cell costimulatory receptors. We demonstrate the utility of this technique to detect the interaction between ICOS and the p85 regulatory subunit of phosphatidylinositol 3-kinase (PI3K). The p85 unit of PI3K is the only signaling molecule identified so far that interacts with ICOS. This system may be of great help in dissecting the mechanisms of T cell costimulation and could be applied to other receptors.