No abstract

Selenite is a predominant form of selenium (Se) available to plants, especially in anaerobic soils, but the molecular mechanism of selenite uptake by plants is not well understood.ltn1, a rice mutant previously shown to have increased phosphate (Pi) uptake, was found to exhibit higher selenite uptake than the wild-type in both concentration- and time-dependent selenite uptake assays. Respiratory inhibitors significantly inhibited selenite uptake in the wildtype and the ltn1 mutant, indicating that selenite uptake was coupled with H+ and energy-dependent. Selenite uptake was greatly enhanced under Pi-starvation conditions, suggesting that Pi transporters are involved in selenite uptake.OsPT2, the most abundantly expressed Pi transporter in the roots, is also significantly up-regulated in ltn1 and dramatically induced by Pi starvation. OsPT2-overexpressing and knockdown plants displayed significantly increased and decreased rates of selenite uptake, respectively, suggesting that OsPT2 plays a crucial role in selenite uptake. Se content in rice grains also increased significantly in OsPT2-overexpressing plants.These data strongly demonstrate that selenite and Pi share similar uptake mechanisms and that OsPT2 is involved in selenite uptake, which provides a potential strategy for breeding Se-enriched rice varieties.

The tumor suppressor Merlin/NF2 functions upstream of the core Hippo pathway kinases Lats1/2 and Mst1/2, as well as the nuclear E3 ubiquitin ligase CRL4DCAF1. Numerous mutations of Merlin have been identified in Neurofibromatosis type 2 and other cancer patients. Despite more than two decades of research, the upstream regulator of Merlin in the Hippo pathway remains unknown. Here we show by high-resolution crystal structures that the Lats1/2-binding site on the Merlin FERM domain is physically blocked by Merlin's auto-inhibitory tail. Angiomotin binding releases the auto-inhibition and promotes Merlin's binding to Lats1/2. Phosphorylation of Ser518 outside the Merlin's auto-inhibitory tail does not obviously alter Merlin's conformation, but instead prevents angiomotin from binding and thus inhibits Hippo pathway kinase activation. Cancer-causing mutations clustered in the angiomotin-binding domain impair angiomotin-mediated Merlin activation. Our findings reveal that angiomotin and Merlin respectively interface cortical actin filaments and core kinases in Hippo signaling, and allow construction of a complete Hippo signaling pathway.

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The comparative typing of matched tumor and blood DNAs at dinucleotide repeat (microsatellite) loci has revealed in tumor DNA the presence of alleles that are not observed in normal DNA. The occurrence of these additional alleles is possibly due to replication errors (RERs). Although this observation has led to the recognition of a subtype of colorectal cancer with a high incidence of RERs (caused by a deficiency in DNA mismatch repair), a thorough analysis of the RER frequency in a consecutive series of colorectal cancers had not been reported. It is shown here that the extensive typing of 88 colorectal tumors reveals a bimodal distribution for the frequency of RER at microsatellite loci. Within the major mode (75 tumors, RER؊ subtype), the probability that a locus exhibited instability did not differ significantly among loci and tumors, being 0.02. The subsequent development of a statistical test for an operational discrimination between the RER؊ and RER؉ subtypes indicated that the probability of misclassification did not exceed 0.001 in this series. The frequency of K-ras mutation was found to be equivalent in the two subtypes. However, in the RER؉ tumors, the p53 gene mutation was less frequently detected, the adenomatous polyposis coli (APC) mutation was rare, and the biallelic inactivation of either of these genes was not observed. Furthermore, the concomitant occurrence of APC and tumor growth factor ␤ receptor type II gene alterations was found only once. These data suggest that the repertoires of genes that are frequently altered in RER؉ and RER؊ tumors may be more different than previously thought.

The c-Myc oncoprotein is a general transcription factor whose target genes dictate the c-Myc phenotype. One such target of c-Myc, 'onzin', is normally expressed at high levels in myeloid cells and is dramatically downregulated in response to c-Myc overexpression. We show here that short hairpin interfering RNA-mediated knockdown of endogenous onzin results in a reduced growth rate and a proapoptotic phenotype. In contrast, onzin overexpression in fibroblasts is associated with an increased growth rate, resistance to apoptotic stimuli, loss of the G2/M checkpoint, and tumorigenic conversion. Onzin-overexpressing cells fail to induce p53 in response to apoptotic stimuli and contain higher levels of the active, phosphorylated forms of Akt1 and, more strikingly, of Mdm2. Using yeast two-hybrid and coimmunoprecipitation assays, we show that onzin directly interacts with both proteins. Green fluorescent protein tagging also confirms directly that Akt1 and Mdm2 colocalize with onzin, although the precise subcellular distribution of each protein is dependent on its relative abundance. Collectively, our results identify onzin as a novel regulator of several p53-dependent aspects of the c-Myc phenotype via its dramatic effect on Mdm2. This is reminiscent of the c-Mycp19 ARF -| Mdm2 pathway and might function as a complementary arm to ensure the proper cellular response to oncogenic and/or apoptotic stimuli.

Friend Leukemia Virus Induced erythroleukemia-1 (Fli-1), an ETS transcription factor, was isolated a quarter century ago through a retrovirus mutagenesis screen. Fli-1 has since been recognized to play critical roles in normal development and homeostasis. For example, it transcriptionally regulates genes that drive normal hematopoiesis and vasculogenesis. Indeed, Fli-1 is one of 10 key regulators of hematopoietic stem/progenitor cell maintenance and differentiation. Aberrant expression of Fli-1 also underlies a number of virally induced leukemias, including Friend virus-induced erythroleukemia and various types of human cancers, and it is the target of chromosomal translocations in childhood Ewing’s sarcoma. Abnormal expression of Fli-1 is important in the aetiology of auto-immune diseases such as Systemic Lupus Erythematosus (SLE) and Systemic Sclerosis (SSc). These studies establish Fli-1 as a strong candidate for drug development. Despite difficulties in targeting transcription factors, recent studies identified small molecule inhibitors for Fli-1. Here we review past and ongoing research on Fli-1 with emphasis on its mechanistic function in autoimmune disease and malignant transformation. The significance of identifying Fli-1 inhibitors and their clinical applications for treatment of disease and cancer with deregulated Fli-1 expression are discussed.

The Crumbs (Crb) complex, formed by Crb, PALS1, and PATJ, is evolutionarily conserved in metazoans and acts as a master cellgrowth and -polarity regulator at the apical membranes in polarized epithelia. Crb intracellular functions, including its direct binding to PALS1, are mediated by Crb's highly conserved 37-residue cytoplasmic tail. However, the mechanistic basis governing the highly specific Crb-PALS1 complex formation is unclear, as reported interaction between the Crb tail (Crb-CT) and PALS1 PSD-95/DLG/ZO-1 (PDZ) domain is weak and promiscuous. Here we have discovered that the PDZ-Src homolgy 3 (SH3)-Guanylate kinase (GK) tandem of PALS1 binds to Crb-CT with a dissociation constant of 70 nM, which is ∼100-fold stronger than the PALS1 PDZ-Crb-CT interaction. The crystal structure of the PALS1 PDZ-SH3-GK-Crb-CT complex reveals that PDZ-SH3-GK forms a structural supramodule with all three domains contributing to the tight binding to Crb. Mutations disrupting the tertiary interactions of the PDZ-SH3-GK supramodule weaken the PALS1-Crb interaction and compromise PALS1-mediated polarity establishment in MadinDarby canine kidney (MDCK) cysts. We further show that specific target binding of other members of membrane-associated guanylate kinases (MAGUKs) (e.g., CASK binding to neurexin) also requires the presence of their PDZ-SH3-GK tandems.cell polarity | supramodule | MAGUK | PBM | Stardust